Diagnostic Pathology: Transplant Pathology [2 ed.] 0323553575, 9780323553575

Table of contents :
Front Cover
Diagnostic Pathology: Transplant Pathology
Copyright Page
Dedication
Contributing Authors
Preface
Acknowledgments
Sections
Table of Contents
SECTION 1:
Immunology
Chapter
1. Immune Response in Organ Transplantation
INTRODUCTION
INNATE IMMUNE RESPONSES IN SOT
INFECTION, TISSUE DAMAGE, AND IMMUNE
RESPONSE IN SOT
NK CELLS AND IMMUNE RESPONSE IN SOT
B-CELL IMMUNE RESPONSES IN SOT
NF-ΚB AND T CELLS IN SOT
IMMUNOSUPPRESSIVE THERAPY IN SOT
SUMMARY
SELECTED REFERENCES
Chapter 2. Regulatory Immune Cells and Transplant Tolerance
TERMINOLOGY
INTRODUCTION
IMMUNOLOGICAL TOLERANCE
TRANSPLANT TOLERANCE
SELECTED REFERENCES
Chapter 3. NK Cells
NK-CELL BIOLOGY
NK CELLS IN SOLID ORGAN
TRANSPLANTATION
NKT CELLS IN SOT
SUMMARY
SELECTED REFERENCES
Chapter 4. Complement
ROLE OF COMPLEMENT IN ATYPICAL
HEMOLYTIC UREMIC SYNDROME AND
THROMBOTIC MICROANGIOPATHY
SELECTED REFERENCES
Chapter 5. Laboratory-Based Immune Monitoring in Organ Transplantation
INTRODUCTION
LABORATORY APPROACHES TO IMMUNE
MONITORING
IMMUNE RESPONSE BIOMARKERS
INFECTIONS IN SOT
SUMMARY
SELECTED REFERENCES
SECTION 2:
HLA Testing
Chapter 6. Human Leukocyte Antigen System
TERMINOLOGY
INTRODUCTION
GENETICS
HUMAN LEUKOCYTE ANTIGEN PROPERTIES
STRUCTURE
NOMENCLATURE
ANTIHUMAN LEUKOCYTE ANTIGEN
ANTIBODIES
SELECTED REFERENCES
Chapter 7. Histocompatibility Testing
INTRODUCTION
HLA TYPING
ANTI-HLA ANTIBODY TESTING
CROSSMATCH TESTING
OTHER TESTING
SELECTED REFERENCES
Chapter 8. Transplantation and HLA
INTRODUCTION
IMMUNE RESPONSES TO ALLOGRAFTS
HLA MATCHING
ANTI-HLA ANTIBODIES
SELECTED REFERENCES
Chapter 9. ABO Blood Group Antigens and Transplantation
ABO BLOOD GROUP SYSTEM
ABO AND TRANSPLANTATION
SELECTED REFERENCES
Chapter 10. Apheresis and Transplantation
INTRODUCTION
INDICATIONS FOR THERAPEUTIC APHERESIS
IN TRANSPLANTATION
SELECTED REFERENCES
Chapter 11. HLA and Transfusion
INTRODUCTION
HUMAN LEUKOCYTE ANTIGEN-RELATED
IMMUNOLOGICAL TRANSFUSION
REACTIONS
SELECTED REFERENCES
SECTION 3:
Immunosuppressive Drugs
Chapter 12. Mechanism of Action of Immunosuppressive Drugs
MECHANISMS OF REJECTION
IMMUNOSUPPRESSIVE DRUGS
SELECTED REFERENCES
Chapter 13. Therapeutic Drug Monitoring in Transplant Patients
TERMINOLOGY
CLINICAL ISSUES
ETIOLOGY/PATHOGENESIS
PHARMACOLOGIC AGENTS
SELECTED REFERENCES
Chapter 14. History of Immunosuppression Drugs in Transplantation
HISTORICAL OVERVIEW OF
IMMUNOSUPPRESSION
DRUGS IN CLINICAL USE (1962-1994)
MODERN ERA (1995-PRESENT)
ANTIBODY THERAPIES
SELECTED REFERENCES
SECTION 4:
Kidney Transplantation
Chapter 15. History of Kidney Transplantation
CHRONOLOGY AND EVOLUTION
LABORATORY TESTING
ALLOANTIBODY TESTING
OTHER ENDEAVORS
SELECTED REFERENCES
Chapter 16. Pathologic Classification of Renal Allograft Diseases
TERMINOLOGY
DEFINITIONS
CATEGORY 1: ALLOIMMUNE RESPONSES
CATEGORY 2: DRUG TOXICITY AND
HYPERSENSITIVITY
CATEGORY 3: INFECTION
CATEGORY 4: ANATOMIC COMPLICATIONS
CATEGORY 5: RECURRENT AND DE NOVO
DISEASES
CATEGORY 6: DONOR DISEASE
CATEGORY 7: OTHER DISEASES
NOTES
SELECTED REFERENCES
Chapter 17. Evaluation of the End-Stage Kidney
TERMINOLOGY
EPIDEMIOLOGY
ETIOLOGY/PATHOGENESIS
MACROSCOPIC
MICROSCOPIC
SELECTED REFERENCES
Chapter 18. Evaluation of Allograft Kidney
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL IMPLICATIONS
BIOPSY PROCESSING
EVALUATION OF BIOPSY
SPECIAL CONSIDERATIONS IN TRANSPLANT
BIOPSIES
NEW APPROACHES
PROTOCOL BIOPSIES (SURVEILLANCE
BIOPSIES)
BANFF CLASSIFICATION
BANFF SCORING CATEGORIES
SELECTED REFERENCES
Chapter 19. Evaluation of the Donor Kidney
SURGICAL/CLINICAL CONSIDERATIONS
SPECIMEN EVALUATION
EVALUATION OF DONOR BIOPSY
REPORTING
PITFALLS
SELECTED REFERENCES
Chapter 20. Evaluation of Transplant Nephrectomy
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL IMPLICATIONS
MACROSCOPIC
MICROSCOPIC
SELECTED REFERENCES
Chapter 21. Evaluation of Fibrosis
TERMINOLOGY
VALUE
MECHANISMS
METHODS FOR ASSESSMENT
SELECTED REFERENCES
Chapter 22. Protocol Biopsies
TERMINOLOGY
CLINICAL IMPLICATIONS
MICROSCOPIC
DIAGNOSES
SELECTED REFERENCES
Chapter 23. Accommodation
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 24. Tolerance
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Surgical Complications
Chapter 25. Acute Allograft Ischemia
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 26. Urine Leak
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 27. Lymphocele
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 28. Renal Artery or Vein Thrombosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 29. Transplant Renal Artery Stenosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Allograft Rejection
Chapter 30. Hyperacute Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 31. Acute T-Cell-Mediated Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 32. Chronic T-Cell-Mediated Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 33. Acute Antibody-Mediated Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 34. Chronic Antibody-Mediated Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
REPORTING
SELECTED REFERENCES
Recurrent Diseases
Chapter 35. Recurrent Diseases in the Allograft
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
De Novo Diseases
Chapter 36. De Novo Focal Segmental Glomerulosclerosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 37. De Novo Membranous Glomerulonephritis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 38. Anti-GBM Disease in Alport Syndrome
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 39. Hyperperfusion Injury
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 40. Engraftment Syndrome
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 41. Kidney Diseases in Nonrenal Transplant Recipients
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 42. Graft-vs.-Host Glomerulopathies
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Drug Toxicities
Chapter 43. Calcineurin Inhibitor Toxicity
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 44. mTOR Inhibitor Toxicity
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Infections
Chapter 45. Acute Pyelonephritis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 46. Polyomavirus Nephritis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 47. Adenovirus, Kidney
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 48. Cytomegalovirus Infection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 49. Histoplasmosis
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 50. Candidiasis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 51. Cryptococcosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 52. Mucormycosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 53. Aspergillosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 54. Coccidioidomycosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 55. Paracoccidioidomycosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 56. Microsporidiosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 57. Tuberculosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 58. Malakoplakia
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 59. Nocardiosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
SECTION 5:
Liver Transplantation
Chapter 60. Pathologic Classification of Liver Allograft Diseases
TERMINOLOGY
ALLOIMMUNE RESPONSE
NONALLOIMMUNE DISEASES
RECURRENT DISEASES
INFECTIONS
SELECTED REFERENCES
Chapter 61. Gross Evaluation of Failed Native Liver
TERMINOLOGY
CLINICAL IMPLICATIONS
MACROSCOPIC
SELECTED REFERENCES
Chapter 62. Evaluation of Failed Liver Allograft
CLINICAL IMPLICATIONS
MACROSCOPIC
MICROSCOPIC
SELECTED REFERENCES
Chapter 63. Evaluation of the Donor Liver
TERMINOLOGY
CLINICAL IMPLICATIONS
MICROSCOPIC
SELECTED REFERENCES
Chapter 64 . History of Liver Transplantation
TERMINOLOGY
CHRONOLOGY AND EVOLUTION
ORGAN ALLOCATION POLICY
LIVER ALLOGRAFT PATHOLOGY
SELECTED REFERENCES
Posttransplant Surgical Complications
Chapter 65. Preservation Injury
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 66. Hepatic Artery Thrombosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 67. Portal Vein Thrombosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 68. Bile Duct Stricture, Leak, Sludge, Biloma
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 69. Hepatic Venous Outflow Obstruction
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 70. Hyperperfusion Syndrome
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Allograft Rejection
Chapter 71. T-Cell-Mediated Rejection, Liver
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 72. Antibody-Mediated Rejection, Liver
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 73. Chronic (Ductopenic) Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Recurrent Diseases in Liver Allograft
Chapter 74. Recurrent Hepatitis B Virus
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 75. Recurrent Hepatitis C Virus
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 76. Fibrosing Cholestatic HBV or HCV Hepatitis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 77. Recurrent Autoimmune Hepatitis
TERMINOLOGY
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 78. Recurrent Primary Biliary Cholangitis
TERMINOLOGY
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 79. Recurrent Primary Sclerosing Cholangitis
TERMINOLOGY
CLINICAL ISSUES
IMAGING
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 80. Recurrent Fatty Liver Disease
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Infections
Chapter 81. Cytomegalovirus
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 82. Herpes Simplex Virus
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 83. Adenovirus, Liver
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 84. Hepatitis E Virus
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 85. Epstein-Barr Virus, Liver
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 86. Fungal Infections, Liver
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
SELECTED REFERENCES
Late-Graft Dysfunction
Chapter 87. Plasma Cell-Rich Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 88. Graft-vs.-Host Disease, Liver
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
SECTION 6:
Heart Transplantation
Chapter 89. Pathologic Classification of Cardiac Allograft Diseases
TERMINOLOGY
CLINICAL ISSUES
PATHOLOGIC ISSUES
SELECTED REFERENCES
Chapter 90. Evaluation of Failed Native and Transplanted Heart
CLINICAL IMPLICATIONS
MACROSCOPIC
MICROSCOPIC
SELECTED REFERENCES
Chapter 91. History of Heart Transplantation
TERMINOLOGY
CHRONOLOGY AND EVOLUTION
TREATMENT ISSUES
SELECTED REFERENCES
Evaluation of Explanted Heart
Chapter 92. Ischemic Heart Disease
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 93. Dilated Cardiomyopathy
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 94. Hypertrophic Cardiomyopathy
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 95. Congenital Heart Disease
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
Chapter 96. Sarcoidosis, Heart
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 97. Arrhythmogenic Right Ventricular Cardiomyopathy
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 98. Other Causes of End-Stage Heart Disease
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Allograft Rejection
Chapter 99. Acute Cellular Rejection, Heart
TERMINOLOGY
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 100. Antibody-Mediated Rejection, Heart
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 101. Chronic Allograft Vasculopathy, Heart
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Noninfectious Lesions
Chapter 102. Quilty Lesions
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 103. Site of Previous Biopsy
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Infections
Chapter 104. Myocarditis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
SECTION 7:
Lung Transplantation
Chapter 105. Pathologic Classification of Lung Allograft Diseases
TERMINOLOGY
ALLOIMMUNE RESPONSE
NONALLOIMMUNE DISEASES
INDICATIONS FOR LUNG TRANSPLANTATION
SELECTED REFERENCES
Chapter 106. Examination of Native and Transplanted Lungs
TERMINOLOGY
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 107. History of Lung Transplantation
TERMINOLOGY
LANDMARK EVENTS IN LUNG
TRANSPLANTATION
SELECTED REFERENCES
Evaluation of Failed Native Lung
Chapter 108. Emphysema
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 109. Cystic Fibrosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 110. Idiopathic Pulmonary Fibrosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 111. Connective Tissue Disease-Associated Lung Disease
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 112. Sarcoidosis, Lung
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 113. Pulmonary Arterial Hypertension
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 114. Other Causes of End-Stage Lung Disease
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Surgical Complications
Chapter 115. Surgical Aspects and Complications, Lung
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
SELECTED REFERENCES
Allograft Rejection
Chapter 116. Pathologic Classification of Rejection
TERMINOLOGY
CLINICAL ISSUES
ETIOLOGY/PATHOGENESIS
CLINICAL IMPLICATIONS
MACROSCOPIC FINDINGS
MICROSCOPIC FINDINGS
SELECTED REFERENCES
Chapter 117. Antibody-Mediated Rejection, Lung
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 118. Acute Cellular Rejection, Grade A
TERMINOLOGY
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 119. Acute Cellular Rejection, Grade B
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 120. Chronic Allograft Dysfunction, Lung
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Noninfectious Lesions
Chapter 121. Organizing Pneumonia
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 122. Microaspiration
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Infections
Chapter 123. Bacterial Infections
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 124. Viral Infections
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 125. Fungal Infections, Lung
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
SECTION 8:
Intestinal Transplantation
Chapter 126. Pathologic Classification of Intestinal Allograft Diseases
TERMINOLOGY
DEFINITIONS
ALLOIMMUNE RESPONSE
NONALLOIMMUNE DISEASES
MULTIVISCERAL TRANSPLANT ISSUES
RECURRENT INTESTINAL DISEASES
RETRANSPLANTATION
SELECTED REFERENCES
Chapter 127. Indications and Evaluation of Explant
TERMINOLOGY
EPIDEMIOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL IMPLICATIONS
MACROSCOPIC
MICROSCOPIC
SELECTED REFERENCES
Chapter 128. Reperfusion Injury
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 129. History of Intestinal and Multivisceral Transplantation
TERMINOLOGY
CHRONOLOGY AND EVOLUTION
SELECTED REFERENCES
Allograft Rejection/Immunological Injury
Chapter 130. Acute Antibody-Mediated Rejection, Intestine
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 131. Acute Cellular Rejection, Intestine
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 132. Chronic Rejection, Intestine
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 133. Stomach Rejection
CLINICAL ISSUES
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 134. Colon Rejection
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 135. Graft-vs.-Host Disease, Intestine
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Infections
Chapter 136. Bacterial and Fungal Infections
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 137. Adenovirus, Intestine
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 138. Rotavirus, Cytomegalovirus, and Herpes Simplex Virus
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 139. Epstein-Barr Virus, Intestine
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
SECTION 9:
Pancreas Transplantation
Chapter 140. Pathologic Classification of Pancreas Allograft Diseases
TERMINOLOGY
ALLOIMMUNE RESPONSES
DRUG TOXICITY
NONALLOIMMUNE DISEASES
ISLET CELL TRANSPLANTATION
NOTES
SELECTED REFERENCES
Chapter 141. Clinical Considerations in Pancreas Transplant Evaluation
TERMINOLOGY
CLINICAL IMPLICATIONS
OUTCOMES
ASSESSMENT
DIAGNOSTIC UTILITY OF PANCREAS
TRANSPLANT BIOPSY
COMPREHENSIVE HISTOLOGIC
EXAMINATION
HUMAN ISLET TRANSPLANTATION
SELECTED REFERENCES
Chapter 142. History of Pancreas Transplantation
TERMINOLOGY
CHRONOLOGY AND EVOLUTION
PROGRESS IN TREATMENT
PANCREAS TRANSPLANT BIOPSY
HUMAN ISLET TRANSPLANTATION
SELECTED REFERENCES
Surgical Complications
Chapter 143. Surgical Aspects and Complications, Pancreas
TERMINOLOGY
HISTORY
ETIOLOGY/PATHOGENESIS
CLINICAL IMPLICATIONS
IMAGE FINDINGS
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Allograft Rejection
Chapter 144. Acute Cellular Rejection, Pancreas
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 145. Antibody-Mediated Rejection, Pancreas
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 146. Chronic Allograft Rejection/Graft Sclerosis
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Graft Dysfunction
Chapter 147. Recurrent Diabetes Mellitus
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
SELECTED REFERENCES
Chapter 148. Islet Cell Toxicity and Islet Amyloid Deposition
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MICROSCOPIC
SELECTED REFERENCES
Infections
Chapter 149. Intraabdominal and Opportunistic Infections
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
SECTION 10:
Vascularized Composite
Allotransplantation
Chapter 150. History of Vascularized Composite Allotransplantation
TERMINOLOGY
CHRONOLOGY AND EVOLUTION
CLINICAL ISSUES
SUMMARY
SELECTED REFERENCES
Allograft Rejection
Chapter 151. Acute T-Cell- and Antibody-Mediated Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
Chapter 152. Chronic Rejection
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
SELECTED REFERENCES
SECTION 11:
Posttransplant
Neoplastic Disorders
Chapter 153. Posttransplant Lymphoproliferative Disease
TERMINOLOGY
ETIOLOGY/PATHOGENESIS
CLINICAL ISSUES
IMAGING
MACROSCOPIC
MICROSCOPIC
ANCILLARY TESTS
DIFFERENTIAL DIAGNOSIS
DIAGNOSTIC CHECKLIST
REPORTING
SELECTED REFERENCES
INDEX

Citation preview

SECOND EDITION

CHANG | COLVIN Abraham | Alpert | Bracamonte | Cendales | Cornell | Farris | Gandhi Hart | Husain | Kambham | Langman | Meehan | Pogoriler Ranganathan | Seshan | Sharma | Wang | Westerhoff | Yeh | Yerian

ii

SECOND EDITION

Anthony Chang, MD Professor of Pathology Director, UChicago MedLabs Director, Renal Pathology and Renal Pathology Fellowship Associate Director, Pathology Residency Program The University of Chicago Chicago, Illinois

Robert B. Colvin, MD Benjamin Castleman Distinguished Professor of Pathology Department of Pathology Harvard Medical School Massachusetts General Hospital Boston, Massachusetts

iii

1600 John F. Kennedy Blvd. Ste 1800 Philadelphia, PA 19103-2899

DIAGNOSTIC PATHOLOGY: TRANSPLANT PATHOLOGY, SECOND EDITION

ISBN: 978-0-323-55357-5

Copyright © 2019 by Elsevier. All rights reserved. No part of this publication may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopying, recording, or any information storage and retrieval system, without permission in writing from the publisher. Details on how to seek permission, further information about the Publisher’s permissions policies and our arrangements with organizations such as the Copyright Clearance Center and the Copyright Licensing Agency, can be found at our website: www. elsevier.com/permissions. This book and the individual contributions contained in it are protected under copyright by the Publisher (other than as may be noted herein).

Notices Knowledge and best practice in this field are constantly changing. As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary. Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein. In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility. With respect to any drug or pharmaceutical products identified, readers are advised to check the most current information provided (i) on procedures featured or (ii) by the manufacturer of each product to be administered, to verify the recommended dose or formula, the method and duration of administration, and contraindications. It is the responsibility of practitioners, relying on their own experience and knowledge of their patients, to make diagnoses, to determine dosages and the best treatment for each individual patient, and to take all appropriate safety precautions. To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors, assume any liability for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions, or ideas contained in the material herein. Publisher Cataloging-in-Publication Data Names: Chang, Anthony. | Colvin, Robert B. Title: Diagnostic pathology. Transplant pathology / [edited by] Anthony Chang and Robert B. Colvin. Other titles: Transplant pathology. Description: Second edition. | Salt Lake City, UT : Elsevier, Inc., [2018] | Includes bibliographical references and index. Identifiers: ISBN 978-0-323-55357-5 Subjects: LCSH: Transplantation of organs, tissues, etc.--Complications--Handbooks, manuals, etc. | Transplantation immunology--Handbooks, manuals, etc. | MESH: Organ Transplantation--Atlases. | Transplantation Immunology--Atlases. Classification: LCC RD120.7.D53 2018 | NLM WO 517 | DDC 617.9’54--dc23 International Standard Book Number: 978-0-323-55357-5 Cover Designer: Tom M. Olson, BA Printed in Canada by Friesens, Altona, Manitoba, Canada Last digit is the print number: 9 8 7 6 5 4 3 2 1

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Dedication The authors thank our teachers, students, and families who gave us guidance, inspiration, and love. We owe you everything. You know who you are! We also thank the many investigators, pathologists, and clinicians who created the knowledge we have summarized, and the patients whose biopsies revealed so many of the insights. Bob and Tony

v

Contributing Authors Roshini Sarah Abraham, PhD, D(ABMLI) Professor of Laboratory Medicine & Pathology and Medicine Consultant and Director Cellular and Molecular Immunology Laboratory Department of Laboratory Medicine and Pathology Mayo Clinic Rochester, Minnesota

Lindsay Alpert, MD

Assistant Professor of Pathology The University of Chicago Medical Center Chicago, Illinois

Erika R. Bracamonte, MD

Associate Professor of Pathology Department of Pathology The University of Arizona College of Medicine Banner University Medical Center Tucson, Arizona

Linda Cendales, MD

Associate Professor of Surgery Duke Health Scholar Director, Vascularized Composite Allotransplantation Program Department of Surgery Duke University Durham, North Carolina

Lynn D. Cornell, MD

Consultant, Division of Anatomic Pathology Associate Professor of Laboratory Medicine and Pathology Mayo Clinic College of Medicine and Science Rochester, Minnesota

A. Brad Farris, III, MD

Director, Laboratory of Nephropathology and Electron Microscopy Associate Professor of Pathology Department of Pathology Emory University School of Medicine Atlanta, Georgia

vi

Manish J. Gandhi, MD

Associate Professor of Laboratory Medicine and Pathology Director, Tissue Typing Laboratory Associate Director, Blood Component Laboratory Consultant, Division of Transfusion Medicine Mayo Clinic Rochester, Minnesota

John Hart, MD

Professor of Pathology Sections of Surgical Pathology & Hepatology University of Chicago Medical Center Chicago, Illinois

Aliya N. Husain, MD Professor of Pathology The University of Chicago Chicago, Illinois

Neeraja Kambham, MD

Professor of Pathology Co-Director, Renal Pathology & EM Laboratory Department of Pathology Stanford University Stanford, California

Loralie Langman, PhD

Department of Laboratory Medicine and Pathology Professor of Laboratory Medicine and Pathology Mayo Clinic College of Medicine Rochester, Minnesota

Shane M. Meehan, MBBCh Renal Pathology Service Sharp Memorial Hospital San Diego, California

Jennifer Pogoriler, MD, PhD Assistant Professor of Pathology and Laboratory Medicine Pathology and Laboratory Medicine Children’s Hospital of Philadelphia Philadelphia, Pennsylvania

Sarangarajan Ranganathan, MD Professor of Pathology Medical Director, Anatomic Pathology Children’s Hospital of Pittsburgh of UPMC Pittsburgh, Pennsylvania

Surya V. Seshan, MD

Professor of Clinical Pathology Weill Cornell Medical College Cornell University New York, New York

Aarti Sharma, MD

Resident, Department of Pathology The University of Chicago Chicago, Illinois

Hanlin L. Wang, MD, PhD

Professor Director of Gastrointestinal Pathology Department of Pathology and Laboratory Medicine David Geffen School of Medicine University of California at Los Angeles Los Angeles, California

Maria Westerhoff, MD Associate Professor Department of Pathology University of Michigan Ann Arbor, Michigan

Matthew M. Yeh, MD, PhD

Professor of Pathology Adjunct Professor of Medicine Director, Gastrointestinal and Hepatic Pathology Program University of Washington School of Medicine Seattle, Washington

Lisa Yerian, MD

Medical Director, Continuous Improvement Assistant Professor of Pathology Cleveland Clinic Cleveland, Ohio

Additional Contributors Joseph Ahn, MD, MS, FACG Vijayalakshmi Ananthanarayanan, MD Ilyssa O. Gordon, MD, PhD Richard Masia, MD, PhD Rish K. Pai, MD, PhD Ivy A. Rosales, MD vii

viii

Preface Solid organ transplantation represents a monumental achievement of modern medicine and has extended the lives of countless patients. What was once a Herculean challenge is now fairly routine with excellent outcomes, but this could not be possible without major advances in surgical techniques and immunosuppressive agents, as well as improved understanding of transplant immunology and pathology. Diagnostic Pathology: Transplant Pathology, second edition represents the collective knowledge of over 20 experts of the various transplant organs (kidney, liver, heart, lung, pancreas, intestine, and vascularized composite allografts). For the practicing pathologist, transplant pathology provides the ultimate diagnostic challenge, as numerous injuries, including donor disease, surgical complications, allograft rejection, drug toxicity, opportunistic infections, &/or recurrent or de novo diseases can occur simultaneously or at different times. The content within this textbook serves to guide you through this complex realm. The numerous images and bullet point format aim to aid with current and future dilemmas provided at the microscope by your daily practice. The fascinating world of transplant pathology awaits in the following pages.

Anthony Chang, MD Professor of Pathology Director, UChicago MedLabs Director, Renal Pathology and Renal Pathology Fellowship Associate Director, Pathology Residency Program The University of Chicago Chicago, Illinois

Robert B. Colvin, MD Benjamin Castleman Distinguished Professor of Pathology Department of Pathology Harvard Medical School Massachusetts General Hospital Boston, Massachusetts

ix

x

Acknowledgments Lead Editor Matt W. Hoecherl, BS

Text Editors Arthur G. Gelsinger, MA Rebecca L. Bluth, BA Nina I. Bennett, BA Terry W. Ferrell, MS Megg Morin, BA

Image Editors Jeffrey J. Marmorstone, BS Lisa A. M. Steadman, BS

Illustrations Richard Coombs, MS Lane R. Bennion, MS Laura C. Wissler, MA

Art Direction and Design Tom M. Olson, BA Laura C. Wissler, MA

Production Coordinators Angela M. G. Terry, BA Emily C. Fassett, BA

xi

xii

Sections SECTION 1: Immunology SECTION 2: HLA Testing SECTION 3: Immunosuppressive Drugs SECTION 4: Kidney Transplantation SECTION 5: Liver Transplantation SECTION 6: Heart Transplantation SECTION 7: Lung Transplantation SECTION 8: Intestinal Transplantation SECTION 9: Pancreas Transplantation SECTION 10: Vascularized Composite Allotransplantation SECTION 11: Posttransplant Neoplastic Disorders

xiii

TABLE OF CONTENTS

4 8 14 20 24

SECTION 1: IMMUNOLOGY

80

Immune Response in Organ Transplantation Roshini Sarah Abraham, PhD, D(ABMLI) Regulatory Immune Cells and Transplant Tolerance Roshini Sarah Abraham, PhD, D(ABMLI) NK Cells Roshini Sarah Abraham, PhD, D(ABMLI) Complement Roshini Sarah Abraham, PhD, D(ABMLI) Laboratory-Based Immune Monitoring in Organ Transplantation Roshini Sarah Abraham, PhD, D(ABMLI)

86

SECTION 2: HLA TESTING 32 34 36 40 42 44

48 50 54

Human Leukocyte Antigen System Manish J. Gandhi, MD Histocompatibility Testing Manish J. Gandhi, MD Transplantation and HLA Manish J. Gandhi, MD ABO Blood Group Antigens and Transplantation Manish J. Gandhi, MD Apheresis and Transplantation Manish J. Gandhi, MD HLA and Transfusion Manish J. Gandhi, MD

60 62 68 74

xiv

94 96

SURGICAL COMPLICATIONS 100 104 106 108 110

112 116 128

Mechanism of Action of Immunosuppressive Drugs Loralie Langman, PhD and Lynn D. Cornell, MD Therapeutic Drug Monitoring in Transplant Patients Loralie Langman, PhD History of Immunosuppression Drugs in Transplantation Loralie Langman, PhD

132

History of Kidney Transplantation Lynn D. Cornell, MD Pathologic Classification of Renal Allograft Diseases Robert B. Colvin, MD Evaluation of the End-Stage Kidney Shane M. Meehan, MBBCh Evaluation of Allograft Kidney Lynn D. Cornell, MD Evaluation of the Donor Kidney Lynn D. Cornell, MD

Acute Allograft Ischemia A. Brad Farris, III, MD Urine Leak A. Brad Farris, III, MD Lymphocele A. Brad Farris, III, MD Renal Artery or Vein Thrombosis A. Brad Farris, III, MD Transplant Renal Artery Stenosis A. Brad Farris, III, MD

ALLOGRAFT REJECTION

SECTION 3: IMMUNOSUPPRESSIVE DRUGS

SECTION 4: KIDNEY TRANSPLANTATION 58

90

Evaluation of Transplant Nephrectomy Shane M. Meehan, MBBCh Evaluation of Fibrosis A. Brad Farris, III, MD Protocol Biopsies Lynn D. Cornell, MD Accommodation Lynn D. Cornell, MD Tolerance Robert B. Colvin, MD

140

Hyperacute Rejection Lynn D. Cornell, MD Acute T-Cell-Mediated Rejection Lynn D. Cornell, MD and Robert B. Colvin, MD Chronic T-Cell-Mediated Rejection Lynn D. Cornell, MD and Robert B. Colvin, MD Acute Antibody-Mediated Rejection Lynn D. Cornell, MD Chronic Antibody-Mediated Rejection Lynn D. Cornell, MD

RECURRENT DISEASES 152

Recurrent Diseases in the Allograft Anthony Chang, MD and Lynn D. Cornell, MD

DE NOVO DISEASES 160 162 166 168

De Novo Focal Segmental Glomerulosclerosis Anthony Chang, MD De Novo Membranous Glomerulonephritis Anthony Chang, MD Anti-GBM Disease in Alport Syndrome Anthony Chang, MD Hyperperfusion Injury Anthony Chang, MD

TABLE OF CONTENTS 170 174 178

Engraftment Syndrome A. Brad Farris, III, MD Kidney Diseases in Nonrenal Transplant Recipients Robert B. Colvin, MD Graft-vs.-Host Glomerulopathies Anthony Chang, MD

DRUG TOXICITIES 180 186

Calcineurin Inhibitor Toxicity Shane M. Meehan, MBBCh mTOR Inhibitor Toxicity Lynn D. Cornell, MD

INFECTIONS 188 192 200 204 208 210 212 214 216 218 220 222 224 226 230

Acute Pyelonephritis Neeraja Kambham, MD Polyomavirus Nephritis Anthony Chang, MD Adenovirus, Kidney Anthony Chang, MD Cytomegalovirus Infection Anthony Chang, MD Histoplasmosis Anthony Chang, MD Candidiasis Anthony Chang, MD Cryptococcosis Anthony Chang, MD Mucormycosis Anthony Chang, MD Aspergillosis Anthony Chang, MD Coccidioidomycosis Anthony Chang, MD Paracoccidioidomycosis Anthony Chang, MD Microsporidiosis Robert B. Colvin, MD Tuberculosis Neeraja Kambham, MD Malakoplakia Neeraja Kambham, MD Nocardiosis Neeraja Kambham, MD

SECTION 5: LIVER TRANSPLANTATION 234 236 240 242 246

Pathologic Classification of Liver Allograft Diseases John Hart, MD and Lindsay Alpert, MD Gross Evaluation of Failed Native Liver John Hart, MD and Lindsay Alpert, MD Evaluation of Failed Liver Allograft John Hart, MD and Lindsay Alpert, MD Evaluation of the Donor Liver John Hart, MD History of Liver Transplantation Anthony Chang, MD and John Hart, MD

POSTTRANSPLANT SURGICAL COMPLICATIONS 248 250 252 254 256 260

Preservation Injury Hanlin L. Wang, MD, PhD Hepatic Artery Thrombosis Lisa Yerian, MD Portal Vein Thrombosis Lisa Yerian, MD Bile Duct Stricture, Leak, Sludge, Biloma Hanlin L. Wang, MD, PhD Hepatic Venous Outflow Obstruction Hanlin L. Wang, MD, PhD Hyperperfusion Syndrome Hanlin L. Wang, MD, PhD

ALLOGRAFT REJECTION 262 266

270

T-Cell-Mediated Rejection, Liver Lisa Yerian, MD Antibody-Mediated Rejection, Liver Hanlin L. Wang, MD, PhD, Robert B. Colvin, MD, and Richard Masia, MD, PhD Chronic (Ductopenic) Rejection Matthew M. Yeh, MD, PhD

RECURRENT DISEASES IN LIVER ALLOGRAFT 272 274 278 280 282 284 286

Recurrent Hepatitis B Virus Rish K. Pai, MD, PhD and Lindsay Alpert, MD Recurrent Hepatitis C Virus Rish K. Pai, MD, PhD and Lindsay Alpert, MD Fibrosing Cholestatic HBV or HCV Hepatitis Rish K. Pai, MD, PhD and Lindsay Alpert, MD Recurrent Autoimmune Hepatitis Rish K. Pai, MD, PhD and Lindsay Alpert, MD Recurrent Primary Biliary Cholangitis Rish K. Pai, MD, PhD and Lindsay Alpert, MD Recurrent Primary Sclerosing Cholangitis Rish K. Pai, MD, PhD and Lindsay Alpert, MD Recurrent Fatty Liver Disease Rish K. Pai, MD, PhD and Lindsay Alpert, MD

INFECTIONS 288 290 292 294 296 300

Cytomegalovirus Maria Westerhoff, MD and Joseph Ahn, MD, MS, FACG Herpes Simplex Virus Maria Westerhoff, MD and Joseph Ahn, MD, MS, FACG Adenovirus, Liver Maria Westerhoff, MD and Joseph Ahn, MD, MS, FACG Hepatitis E Virus Maria Westerhoff, MD and Joseph Ahn, MD, MS, FACG Epstein-Barr Virus, Liver Maria Westerhoff, MD and Joseph Ahn, MD, MS, FACG Fungal Infections, Liver Maria Westerhoff, MD and Joseph Ahn, MD, MS, FACG

LATE-GRAFT DYSFUNCTION 302

Plasma Cell-Rich Rejection Lisa Yerian, MD

xv

TABLE OF CONTENTS 304

Graft-vs.-Host Disease, Liver Lisa Yerian, MD

EVALUATION OF FAILED NATIVE LUNG 368

SECTION 6: HEART TRANSPLANTATION 308

310 312

Pathologic Classification of Cardiac Allograft Diseases Aliya N. Husain, MD Evaluation of Failed Native and Transplanted Heart Jennifer Pogoriler, MD, PhD History of Heart Transplantation Aliya N. Husain, MD and Aarti Sharma, MD

EVALUATION OF EXPLANTED HEART 314 318 322 326 330 332 336

Ischemic Heart Disease Aliya N. Husain, MD and Aarti Sharma, MD Dilated Cardiomyopathy Jennifer Pogoriler, MD, PhD Hypertrophic Cardiomyopathy Jennifer Pogoriler, MD, PhD and Aliya N. Husain, MD Congenital Heart Disease Jennifer Pogoriler, MD, PhD and Aliya N. Husain, MD Sarcoidosis, Heart Aliya N. Husain, MD and Aarti Sharma, MD Arrhythmogenic Right Ventricular Cardiomyopathy Jennifer Pogoriler, MD, PhD and Aliya N. Husain, MD Other Causes of End-Stage Heart Disease Jennifer Pogoriler, MD, PhD and Aliya N. Husain, MD

ALLOGRAFT REJECTION 342 346 348

Acute Cellular Rejection, Heart Aliya N. Husain, MD and Jennifer Pogoriler, MD, PhD Antibody-Mediated Rejection, Heart Aliya N. Husain, MD and Aarti Sharma, MD Chronic Allograft Vasculopathy, Heart Aliya N. Husain, MD and Aarti Sharma, MD

NONINFECTIOUS LESIONS 350 352

Quilty Lesions Aliya N. Husain, MD and Aarti Sharma, MD Site of Previous Biopsy Aliya N. Husain, MD and Vijayalakshmi Ananthanarayanan, MD

370 372 374 376 378 380

SURGICAL COMPLICATIONS 384

Myocarditis Jennifer Pogoriler, MD, PhD and Aliya N. Husain, MD

388 390 392 394

364 366

xvi

Pathologic Classification of Lung Allograft Diseases Aliya N. Husain, MD and Aarti Sharma, MD Examination of Native and Transplanted Lungs Aliya N. Husain, MD History of Lung Transplantation Aliya N. Husain, MD and Aarti Sharma, MD

Pathologic Classification of Rejection Aliya N. Husain, MD and Aarti Sharma, MD Antibody-Mediated Rejection, Lung Aliya N. Husain, MD and Ilyssa O. Gordon, MD, PhD Acute Cellular Rejection, Grade A Aliya N. Husain, MD Acute Cellular Rejection, Grade B Aliya N. Husain, MD Chronic Allograft Dysfunction, Lung Aliya N. Husain, MD

NONINFECTIOUS LESIONS 396 398

Organizing Pneumonia Aliya N. Husain, MD Microaspiration Aliya N. Husain, MD

INFECTIONS 400 402 406

Bacterial Infections Aliya N. Husain, MD and Ilyssa O. Gordon, MD, PhD Viral Infections Aliya N. Husain, MD and Aarti Sharma, MD Fungal Infections, Lung Aliya N. Husain, MD

SECTION 8: INTESTINAL TRANSPLANTATION

SECTION 7: LUNG TRANSPLANTATION 362

Surgical Aspects and Complications, Lung Aliya N. Husain, MD and Aarti Sharma, MD

ALLOGRAFT REJECTION 386

INFECTIONS 354

Emphysema Aliya N. Husain, MD Cystic Fibrosis Aliya N. Husain, MD Idiopathic Pulmonary Fibrosis Aliya N. Husain, MD and Aarti Sharma, MD Connective Tissue Disease-Associated Lung Disease Ilyssa O. Gordon, MD, PhD and Aliya N. Husain, MD Sarcoidosis, Lung Aliya N. Husain, MD Pulmonary Arterial Hypertension Aliya N. Husain, MD and Aarti Sharma, MD Other Causes of End-Stage Lung Disease Aliya N. Husain, MD and Aarti Sharma, MD

412

414 418

Pathologic Classification of Intestinal Allograft Diseases Sarangarajan Ranganathan, MD Indications and Evaluation of Explant Sarangarajan Ranganathan, MD Reperfusion Injury Sarangarajan Ranganathan, MD

TABLE OF CONTENTS 422

History of Intestinal and Multivisceral Transplantation Sarangarajan Ranganathan, MD

INFECTIONS 504

ALLOGRAFT REJECTION/IMMUNOLOGICAL INJURY 424 428 436 440 444 448

Acute Antibody-Mediated Rejection, Intestine Sarangarajan Ranganathan, MD Acute Cellular Rejection, Intestine Sarangarajan Ranganathan, MD Chronic Rejection, Intestine Sarangarajan Ranganathan, MD Stomach Rejection Sarangarajan Ranganathan, MD Colon Rejection Sarangarajan Ranganathan, MD Graft-vs.-Host Disease, Intestine Sarangarajan Ranganathan, MD

SECTION 10: VASCULARIZED COMPOSITE ALLOTRANSPLANTATION 512

456 460

464

Bacterial and Fungal Infections Sarangarajan Ranganathan, MD Adenovirus, Intestine Sarangarajan Ranganathan, MD Rotavirus, Cytomegalovirus, and Herpes Simplex Virus Sarangarajan Ranganathan, MD Epstein-Barr Virus, Intestine Sarangarajan Ranganathan, MD

History of Vascularized Composite Allotransplantation Linda Cendales, MD

ALLOGRAFT REJECTION 514

522

INFECTIONS 452

Intraabdominal and Opportunistic Infections Surya V. Seshan, MD

Acute T-Cell- and Antibody-Mediated Rejection A. Brad Farris, III, MD, Ivy A. Rosales, MD, and Robert B. Colvin, MD Chronic Rejection A. Brad Farris, III, MD, Robert B. Colvin, MD, and Ivy A. Rosales, MD

SECTION 11: POSTTRANSPLANT NEOPLASTIC DISORDERS 526

Posttransplant Lymphoproliferative Disease A. Brad Farris, III, MD

SECTION 9: PANCREAS TRANSPLANTATION 472

474

480

Pathologic Classification of Pancreas Allograft Diseases Surya V. Seshan, MD Clinical Considerations in Pancreas Transplant Evaluation Surya V. Seshan, MD History of Pancreas Transplantation Surya V. Seshan, MD

SURGICAL COMPLICATIONS 482

Surgical Aspects and Complications, Pancreas Surya V. Seshan, MD

ALLOGRAFT REJECTION 488 492 496

Acute Cellular Rejection, Pancreas Erika R. Bracamonte, MD Antibody-Mediated Rejection, Pancreas Erika R. Bracamonte, MD and Surya V. Seshan, MD Chronic Allograft Rejection/Graft Sclerosis Erika R. Bracamonte, MD

GRAFT DYSFUNCTION 498 502

Recurrent Diabetes Mellitus Surya V. Seshan, MD Islet Cell Toxicity and Islet Amyloid Deposition Surya V. Seshan, MD

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SECOND EDITION

CHANG | COLVIN Abraham | Alpert | Bracamonte | Cendales | Cornell | Farris | Gandhi Hart | Husain | Kambham | Langman | Meehan | Pogoriler Ranganathan | Seshan | Sharma | Wang | Westerhoff | Yeh | Yerian

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SECTION 1

Immunology

Immune Response in Organ Transplantation Regulatory Immune Cells and Transplant Tolerance NK Cells Complement Laboratory-Based Immune Monitoring in Organ Transplantation

4 8 14 20 24

Immunology

Immune Response in Organ Transplantation

INTRODUCTION Defining Immune Response in Solid Organ Transplantation • Solid organ transplantation (SOT) involves graft-host interaction resulting in alloimmune response • Interaction has 3 key phases ○ Induction: Antigen recognition ○ Effector: Direct allograft injury ○ Resolution: Decrease in immune response to allograft • T cells defined as key players in SOT ○ Most immunosuppressive therapies target modulation of T-cell immune responses • Advances in transplantation immunology reveal role for B cells, NK cells, and components of innate immune response in maintaining allograft rejection ○ Complement also plays role in allograft responses and priming alloreactive T cells ○ NK cells have dual roles in SOT – Produce inflammatory mediators – Regulate immune responses ○ B cells play role in both acute and chronic antibodymediated rejection • Nonimmunologic tissue damage (i.e., ischemia-reperfusion injury) and infections ○ Enhance alloreactivity and promote rejection episodes ○ Mediated by production of damage-associated molecular patterns (DAMPs) and pathogen associated molecular patterns (PAMPs) ○ Mediated by modifications of alloreactive T-cell repertoire • Early inflammatory response to tissue injury depends upon adaptive immunity

INNATE IMMUNE RESPONSES IN SOT Innate Immunity and Complement in Allograft Rejection and Tolerance • Innate, antigen-independent proinflammatory events occur soon after SOT

○ May be regulated and enhanced by graft-specific adaptive immune response • Innate immune responses primarily mediated by PAMPs or pathogen-recognition receptors (PRRs) ○ PAMPs or PRRs recognize pathogen-derived molecules and host-derived molecules from damaged/stressed tissues ○ Ischemic and surgical trauma can trigger release of endogenous molecules capable of activating PRRs ○ Toll-like receptors (TLRs) are important PRRs that activate innate immunity and direct adaptive immune responses – TLRs are expressed on following cells □ Dendritic cells (DCs) □ B cells □ Mast cells □ T cells □ Endothelial cells □ Organ parenchymal cells – TLR expression modulated by inflammatory mediators and other localized or systemic activation signals – TLR stimulation results in activation of key transcription factors, such as NF-κB – Results in production of many mediators and augmented functions □ Proinflammatory cytokines □ Chemokines □ Antimicrobial peptides □ Adhesion molecules □ Enhanced antigen presentation □ Upregulation of costimulatory molecules on antigen-presenting cells (APCs) ○ Other PRRs include – Nucleotide-binding oligomerization domain (NOD)like receptors (NLRs) □ Direct role as intracellular sensors of cellular stress □ Components of inflammasome, which control activation of proinflammatory cytokines, IL-1β, and IL-18 – RIG-like helicases (RLHs)

Danger-Associated Molecular Signals (Left) Damage-associated molecular patterns (DAMPs) can accumulate in a donor organ through ischemiareperfusion injury or time from death. DAMPs act via toll-like receptors (TLRs) or specific receptors to activate innate immunity. (Right) TLRs recognize DAMPs, while complement receptors recognize complement effector molecules. Stressinduced signals through pattern-recognition receptors (PRR) mediate injury to tissue.

4

Innate Immune Response and Mechanisms of Injury

Immune Response in Organ Transplantation

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○

○

INFECTION, TISSUE DAMAGE, AND IMMUNE RESPONSE IN SOT Infections and Alloreactivity • Infections (viral) in pretransplant period can induce memory T cells that cross react with allogeneic MHC through direct allorecognition (heterologous immunity) ○ Alloreactive memory T cells are more important in allograft rejection because of – Propensity for rapid expansion – Production of inflammatory and cytotoxic effector mediators ○ Pretransplant frequency of donor-specific memory T cells (producing IFN-γ) in renal transplant patients correlates with risk of posttransplant rejection episodes • Posttransplant bacterial, fungal, and viral infections also associate with development of acute or chronic rejection, depending on type of SOT and infection • Activation of PRRs by infection results in eventual T-cell activation and differentiation, with downstream activation of other hematopoietic cells that may participate in rejection • Different pathogens elicit different classes of immune response • Type I IFNs produced during viral and intracellular bacterial infections and stimulate immune responses ○ Treatment of recurrent hepatitis C viral infections with IFN-α in transplant patients facilitates viral clearance but increases risk of allograft rejection • Besides bystander effects of infections (via cytokines) on Tcell activation, infections directly influence uptake and presentation of alloantigens • Certain infections (e.g., CMV) may have immunosuppressive effect systemically, which predisposes to opportunistic superinfections • New therapeutic approaches focus on blocking innate immune responses that stimulate alloreactivity but not protective immunity to infections ○ e.g., rapamycin (sirolimus) increases magnitude and quality of effector and memory CD8 T-cell responses to infection but inhibits alloreactive CD8 T-cell responses

Immunology

○

– Receptor for advanced glycation end products (RAGE) – Scavenger receptors – Complement receptors – Mannose-binding lectin Cells of innate immune compartment are critical in alloantigen presentation – Mature, activated DCs perform many functions □ Secretion of proinflammatory cytokines □ Upregulation of MHC class II on cells □ Increased expression of T-cell costimulatory molecules □ In absence of "danger" signals, DCs remain immature and mediate tolerance via induction of anergy or apoptosis on cognate interaction with antigen-specific T cells – Macrophages do not effectively prime naive T cells but play key role in immediate posttransplant period □ Donor and recipient macrophages infiltrate allografts and proliferate □ Absolute numbers of macrophages decrease in absence of rejection – In acute rejection, macrophage infiltration accounts for 40-60% of cellular infiltrate and performs following roles □ Production of proinflammatory cytokines □ Phagocytose necrotic cell debris □ Production of reactive oxygen species (ROS) □ Presents antigen to effector T cells – Neutrophils also mediate tissue injury via cytotoxic and proinflammatory mechanisms – NK cells are important innate lymphocytes, and while unlikely to solely mediate allograft rejection, they act as facilitators by amplifying early graft inflammation and supporting T-cell alloreactivity Complement activation, when unchecked, can result in tissue (allograft) injury – Plays key role in antibody-mediated rejection by promoting alloantigen-specific B-cell maturation and reducing threshold for B-cell stimulation by antigens Peripheral synthesis of complement characteristic of newly transplanted organs – Determines allograft response to surgical and other stressors – Contributes to T-cell priming and shaping of adaptive immune response related to transplant rejection Roles of complement – Complement triggered by binding of alloantibodies to donor organ endothelial cells – Significant variability in endothelial cell response to pathogenic antibody and complement – 3 possible outcomes □ Immediate thrombosis and allograft infarct □ Lesser injury with gradual decline of allograft function due to complement deposition □ No injury – All these phenomena can contribute to allograft damage and rejection

Danger Signals and Role in SOT • PAMPs allow innate recognition of invading pathogens • DAMPs also activate innate immunity ○ Via recognition of endogenous cellular stress "danger" signals • DAMPs can be subclassified based on physiological presence and properties ○ Intracellular molecules, including nucleic acids, and heatshock proteins (HSPs) – Not accessible normally to immune system – Only released into extracellular environment or expressed on cell surface after cell damage ○ Extracellular molecules, including extracellular matrix components, altered by cellular stress or injury • Process of organ harvesting and transplantation alone can generate DAMPs and activate innate immunity

5

Immunology

Immune Response in Organ Transplantation ○ Most DAMPs signal through TLRs, resulting in NF-κB activation and activation of inflammatory response genes, production of inflammatory cytokines, neutrophil recruitment, preparation of APC, and upregulation of costimulatory molecules and MHC – Other than TLRs, DAMPs also signal through nonPRRs, e.g. ligation of RAGE, which can act in concert with TLR activation to modulate inflammatory response • DAMPs can drive fibrosis-based tissue restructuring in allograft ○ Generate different downstream signals compared to PAMPs • Effect of DAMPs on rates of rejection and graft dysfunction provide avenues for therapeutic intervention ○ Either via decreasing DAMP expression, enhancing clearance, or blocking signaling • Important DAMPs in SOT, depending on transplant organ: HMGB1, ATP, HSPs, nucleic acids, Heparan sulfate, hyaluronan, fibronectin, haptoglobin

NK CELLS AND IMMUNE RESPONSE IN SOT NK-Cell-Mediated Responses in Rejection and Tolerance • Activated NK-cell production of IFN-γ augments early adaptive immune responses ○ NK-produced IFN-γ postulated to provide costimulation to antigen-experienced T cells – NK cells serve as bridge between innate and adaptive immunity in rejection • NK activating receptors, such as NKG2D, promote rejection by recognition of activating ligands on allografts • Mature DCs resist NK-mediated cytotoxicity through upregulation of MHC class I ○ Result in enhanced Th1-specific allogeneic responses by selecting mature DCs • Immature DCs eliminated by NK cell killing • Counterpoint to NK cell function in rejection is their role in tolerance induction ○ Both activating and inhibitory NK cell receptors required for mediating tolerance ○ Secretion of IL-10 in response to infection is example of NK cell regulation in systemic inflammation • NK cell regulation of immune response requires both cytotoxicity and cytokine production • NK cell immune responses regulated in turn by Tregs ○ TGF-β produced by Tregs suppresses cytotoxic profile of NK cells, inhibits granzyme A and B and CD16 expression, and downregulates activating receptor, NKG2D – Depletion of Tregs enhances NK-cell proliferation and cytotoxicity • Unique role of NK cells determine if downstream T-cell response skews toward tolerance or rejection, based on their effector function

B-CELL IMMUNE RESPONSES IN SOT Acute and Chronic Antibody-Mediated Rejection and Immunosuppression • Role of donor-specific alloantibodies (DSA) well established in allograft rejection 6

○ Non-donor-specific HLA antibodies reported and appear earlier compared to DSA • Weaning IS or homeostatic repopulation after immune cell depletion can lower threshold for B-cell activation ○ May facilitate DSA production, C4d(+) injury, and chronic rejection • Several commonly used transplant IS agents influence B-cell activation and DSA production to varying degrees ○ Alemtuzumab, monoclonal anti-CD52 antibody when used as induction agent, especially in absence of CNI immunosuppression – Causes B-cell suppression for 3-12 months – But associates with increased incidence of antibodymediated rejection in patients with preformed DSA ○ Belatacept, CTLA4 fusion protein – Binds CD80/CD86 – Blocks interaction with CD28 (inhibiting T-cell costimulation) – Decreases incidence of de novo alloantibody production ○ Bortezomib, proteasome inhibitor – Shown in early studies to reverse antibody-mediated rejection and decrease DSA after plasmapheresis • Additional studies needed to achieve robust elimination of DSA and durable allograft maintenance ○ Especially in subset of patients who have high levels of DSA pretransplant or either unresponsive or transiently responsive to treatment

NF-ΚB AND T CELLS IN SOT Role of NF-κB and T Cells in Transplant Immune Response • NF-κB, pleiotropic transcription factor, ubiquitously expressed ○ Post SOT, NF-κB in allograft parenchymal cells activated due to ischemic reperfusion injury (IRI) – Also expressed in intragraft-infiltrating cells (activated allogeneic T cells) during acute allograft rejection – Produces proinflammatory cytokines ○ NF-κB activation is not single event but happens repeatedly in cyclical manner – Endogenous DAMPs (intracellular proteins, DNA, RNA and nucleotides that perpetuate noninfectious inflammatory response) produced during IRI □ Result in TLR-2/TLR-4-dependent activation of NFκB □ Induces expression of proinflammatory cytokines such as IL-1 and TNF in myeloid cells and other cells, depending on nature of immune response □ Downstream signaling of these cytokines □ Activation of NF-κB (2nd wave) – This round of NF-κB upregulates effector molecules (IL-8, MCP-1) □ Attracts leukocytes to site of inflammation □ Promotes tissue injury (release of ROS) • T-cell activation through T-cell receptor signals NF-κB activation and eventually IL2 gene transcription and T-cell proliferation • Alloantigen recognition and T-cell activation in allograft rejection includes

Immune Response in Organ Transplantation

IMMUNOSUPPRESSIVE THERAPY IN SOT Modulation of Immune Response • IS in current era primarily antibody-based (but CNI and mTOR inhibitors also used) ○ Both monoclonal and polyclonal antibodies used to prevent and treat allograft rejection ○ Reduces incidence of acute rejection episodes ○ Improves short- and long-term graft survival but increased risk for both opportunistic infections and posttransplant malignancies • Immunosuppression used to target or control 1 or more of 3 alloimmune response phases (signal 1-3) • Induction immunosuppression based on type of SOT ○ 70-80% of SOT use induction with exception of liver transplant ○ Only 20% of liver transplants use induction • Immune response in SOT can be summarized in 3-signal model ○ Signal 1: Initiation of T-cell activation by alloantigen presentation on MHC class II by APCs

○ Signal 2: Expansion of T-cell activation with costimulatory signals (CD80/CD86 interaction with CD28) ○ Signal 3: Culmination of T-cell activation with stimulation of calcium-calcineurin pathway – Results in production of IL-2 by activated T cells – IL-2 binds IL-2R on activated T cells and triggers mTOR – Induces T-cell proliferation and further cytokine production • IS regimens typically consist of triple drug-maintenance therapy acting at different levels of immune response cascade ○ Induction agents act at signal 1 by effacing lymphocyte depletion (e.g., ATG, alemtuzumab) ○ Calcineurin inhibitors (CNI) and IL-2R antagonists act at signal 1 and 3 (e.g., cyclosporine, tacrolimus) – Inhibit cytokine production and IL-2-mediated activation of mTOR ○ Antiproliferative agents [e.g., mycophenolate mofetil (MMF), azathioprine] act downstream of signal 3 – Inhibit purine synthesis and work in concert with mTOR inhibitors (e.g., sirolimus) – Inhibits T-cell proliferation ○ Costimulation inhibitors (e.g., belatacept) act on signal 2 – Block binding of CD80 and CD86 to CD28 receptor

Immunology

○ Direct (intact donor HLA presenting "normal" antigen to recipient T cells) pathway ○ Indirect (alloantigen presentation by recipient DCs) pathway • NF-κB activation via canonical pathway also critical for thymic generation of natural Tregs, which suppresses allograft immune responses and prolongs allograft survival ○ Canonical pathway for NF-kB activation includes induction by physiological NF-κB stimuli and represented by TNFR1 signaling through binding of TRADD, recruitment of FADD and TRAF2 – IκBα phosphorylated in IKKβ-and NEMO-dependent manner, resulting in nuclear translocation of of p65containing heterodimers ○ Animal model data suggests that NF-κB may be important for Treg development but inhibitory for Treg function ○ Inhibition of upstream molecule of NF-κB pathway results in enhanced Treg suppressor function – Highlights potential role of this pharmacologic agent in improving graft survival • NF-κB occupies central role in allotransplantation ○ Modulates deleterious effects of IRI ○ Promotes survival of activated T cells ○ Facilitates differentiation of Th1, Th2, and Th17 effector cells ○ Promotes memory T-cell generation ○ Also involved in DC maturation and may downmodulate peripheral development of Tregs • NF-κB role in alloimmunity suggests specific blocking of Tcell NF-κB may prevent allograft rejection ○ NF-κB appears to be viable therapeutic target in transplantation – Risk:benefit ratio has to be assessed and toxicity of such therapies reduced – For more immunogenic transplanted organs, additional or alternate therapies may be needed since allograft rejection can take place in context of NF-κBimpaired T cells

SUMMARY Manipulation of Immune Response in SOT • Success of allotransplantation depends on optimizing immunosuppression to curb allograft-damaging immune responses while maintaining effective immunity against pathogens • Immune response (IR) against transplanted organs originates from several genetic barriers ○ Blood group antigens ○ HLA ○ Minor histocompatibility antigens ○ IR gene polymorphisms that modulate risk of infection or rejection • Control of IR in SOT requires multipronged approach ○ Minimize immunosuppression (personalized strategies) ○ Identify regimens for tolerance induction ○ Preserve immunity to pathogens ○ All these aspects may require combination of pharmacological intervention along with adoptive immunotherapies, which include adoptive transfer of ex vivo-expanded Tregs, antiviral cytotoxic T cells, and other types of regulatory immune cells

SELECTED REFERENCES 1. 2. 3. 4. 5.

Todd JL et al: Danger signals in regulating the immune response to solid organ transplantation. J Clin Invest. 127(7):2464-2472, 2017 Farrar CA et al: The innate immune system and transplantation. Cold Spring Harb Perspect Med. 3(10):a015479, 2013 Chong AS et al: The impact of infection and tissue damage in solid-organ transplantation. Nat Rev Immunol. 12(6):459-71, 2012 LaRosa DF et al: The innate immune system in allograft rejection and tolerance. J Immunol. 178(12):7503-9, 2007 Villard J: Immunity after organ transplantation. Swiss Med Wkly. 136(5-6):717, 2006

7

Immunology

Regulatory Immune Cells and Transplant Tolerance – Antigen-specific recognition essential for CD8(+) Treg function

TERMINOLOGY Synonyms

Innate Immune Components

• Immunological tolerance to allografts

Definitions • Survival of well-functioning allograft without need for maintenance immunosuppression in context of normal host immune response

• • • •

Dendritic cells (DCs) Monocytes Endothelial cells Complement

IMMUNOLOGICAL TOLERANCE INTRODUCTION

Mechanisms of Tolerance in Normal Immunity

Adaptive Immune Components • • • •

T cells B cells NK cells Regulatory T cells (Tregs) ○ a.k.a. suppressor cells and account for 10-15% of CD4 T cells ○ Highly heterogeneous and plastic population of cells ○ Usually CD4(+) T cells expressing CD25 (IL-2Rα) and transcription factor FOXP3 ○ Antigen-specific and require continuous antigen exposure to stay active ○ Anergic to T-cell receptor (TCR)-mediated activation – But suppresses activation of other T cells ○ Thymic-derived (natural) Tregs generated from precursors in thymus ○ Induced Tregs generated in periphery through costimulation blockade – Either TGF-β-secreting Th3 cells or IL-10-secreting Th1 cells ○ Major histocompatibility complex (MHC)-restricted antigen presentation of donor peptides by self- antigenpresenting cells induces antigen-specific Tregs capable of direct and indirect suppression of other alloreactive T cells ○ CD8(+) Tregs also described – Contain low levels of FOXP3 and represent < 1% of thymocytes

Lymphoid Aggregate (Left) A nodular inflammatory aggregate ﬈ is seen in a kidney allograft on surveillance biopsy. (Right) Double stain for FOXP3 (blue nuclear stain) and CD4 in aggregate on kidney protocol biopsy shows CD4(+) T cells, a few of which also express FOXP3 ﬈.

8

• Central tolerance (thymus dependent) ○ Occurs by deletion (negative selection) of self-reactive T cells in thymus – When T cells recognize endogenous peptides bound to self MHC molecules with either high or low affinity ○ Positive selection in thymus allows export and survival of only T cells with TCRs that recognize peptide and selfMHC with moderate affinity • Peripheral tolerance (non-thymus dependent) ○ Possible mechanisms of peripheral tolerance include – Immunological ignorance (sequestration of antigens) – Apoptosis of T cells through persistent activation – Clonal anergy: Results from lack of "2nd" signal (costimulation), which is required in addition to cognate (peptide + MHC) signal for T-cell activation

Organ Transplantation and Immunity • Allografts, single organs, or composite tissue can be transplanted into appropriate recipient and maintain normal function ○ However, lifelong immunosuppression required to maintain functional allograft, except in rare cases

TRANSPLANT TOLERANCE Operational Tolerance • Immunological unresponsiveness specifically directed to donor allograft without exogenous immunosuppression • Immunological tolerance can be either complete (full) or partial

FOXP3/CD4 Immunohistochemistry

Regulatory Immune Cells and Transplant Tolerance

Regulatory Immune Cells • Critical to maintenance of operational tolerance ○ Tregs: Play key role in tolerance induction through suppression of alloreactive T cells – Regulate peripheral tolerance through cytokines, such as IL-10 and TGF-β ○ Other mechanisms of Treg suppression include – Attenuation of DC maturation and function – Direct killing of effector T cells – Production of antiinflammatory cytokines • Regulatory B cells • Transitional B cells • Tolerogenic DCs • Tolerogenic macrophages • Plasmacytoid DCs (pDCs) • Myeloid-derived suppressor cells (MDSCs), all subsets • Mesenchymal stem cells

Rejection vs. Graft Acceptance/Tolerance • Participation of immune cells in determining allograft outcome depends on ○ Origin of immune cells and activation state ○ Status of organ prior to transplantation (tissue damage and ischemia-reperfusion injury) ○ Conditions in recipient (macro- and microenvironment) • Regulatory immune cells interact with each other in induction of immunological tolerance • Use of exogenous immunosuppression can lead to alteration in homeostasis (secondary to leukodepletion) and favor preferential reconstitution of regulatory immune cells • IL-10 is key cytokine common to several regulatory leukocytes and facilitates microenvironment for immune regulation, including generation and enhancement of regulatory cells post transplant

Assessment of Transplant Tolerance • Few small studies suggest mechanism of operational tolerance related to active induction of immunological tolerance as opposed to immune incompetence • Biomarkers of transplant tolerance ○ Reliable biomarkers or assays needed to determine immunological tolerance, whether achieved through full or partial approach ○ Invasive biopsies not suitable for regular (daily/weekly) monitoring of graft status ○ Ideal biomarkers measured by non- or minimally invasive methods (blood or urine) ○ Biomarkers for operational tolerance described for liver and renal transplantation; however, very different between 2 organs – To determine tolerance induction in liver transplantation (LT), NK-cell proportions and gene transcripts as well as γδ T-cell levels and transcripts suggested as biomarkers in blood – Other biomarkers in LT include □ Regulatory T-cell numbers [FOXP3(+)] or only CD4(+)25(+)(high) numbers □ Plasmacytoid:myeloid DC ratio (pDC:mDC) □ HLA-G expression on mDC □ Soluble HLA-G □ Proportion of peripheral B cells – Increased CD8(+) Tregs shown to correlate with reduced immunosuppressive dosage and decreased acute and chronic liver allograft rejection – In renal transplantation, biomarkers include □ Peripheral NK-cell and T-cell numbers □ T-cell alloreactivity □ FOXP3:α-1, -2 mannosidase ratio □ Absent levels of antidonor HLA antibodies – B cells play important role in maintenance of operational tolerance in kidney transplantation – B-cell biomarkers include number of peripheral B cells and their phenotype, B-cell cytokine expression (TGF-β and IL-10), B-cell-specific gene expression in blood, CD20 transcripts in urine – CD8 T cells may be relevant in operational tolerance with increase in central memory CD8 T cells and decreased effector cells with skewed TCR-Vβ usage ○ Caveats to clinical use of biomarkers in transplant patients – Additional data required for widespread use of these biomarkers in assessment of operational tolerance in liver and renal transplantation – Identification of markers for operational tolerance also requires markers for early identification of graft rejection – Biomarkers need validation for identification of patients with operational tolerance in whom immunosuppression can be minimized or withdrawn – From clinical perspective, immunosuppressive drugweaning or withdrawal approaches should not be initiated without robust assessment of extant immune tolerance

Immunology

○ Full tolerance involves indefinite existence of donor allograft without any form of extrinsic immunosuppression and intact immune function, including pathogen response and immune surveillance – Typically achieved by induction of donor chimerism through myeloablative or nonmyeloablative hematopoietic cell transplantation (HCT) in conjunction with organ transplantation – Caveats of full-tolerance approach □ Associated complications of HCT □ Availability of donor organs relative to preparative regimen for HCT □ Requirement for hematopoietic source and solid organ to be from same donor (living donor only) ○ Partial tolerance requires maintenance of allograft through minimal use of immunosuppression – Significantly reduces risk of immunosuppressionassociated morbidity (infection, malignancy, and other adverse effects) – Partial tolerance approaches often involve lymphocyte depletion with biological agents, such as alemtuzumab followed by reduced doses of calcineurin inhibitor (CNI) monotherapy or steroids or mammalian target of rapamycin (mTOR) inhibitor (sirolimus) therapy

9

Immunology

Regulatory Immune Cells and Transplant Tolerance □ Preliminary results encouraging but need additional studies to ensure optimal outcomes while maximizing patient safety – MSCs, by definition, are adherent cells that express CD105, CD73, CD90 □ Lack expression of CD45, CD34, CD14, CD11b, CD79-α, MHC class II – Should be capable of pluripotency and differentiate under appropriate conditions to different cell line lineages – Since MSCs preferentially home to sites of vascular damage or inflammation, they may mitigate toxic effects of ischemia-reperfusion injury, which can trigger innate and adaptive immune response ○ HCT used successfully to achieve immunological tolerance in patients receiving living donor renal transplants – Stable complete or mixed chimerism can allow longterm maintenance (3 months to > 10 years) of allograft without need for exogenous immunosuppression □ However, mixed chimerism more favorable at reducing GVHD than complete chimerism □ Chimerism seems essential for induction of tolerance but not maintenance □ Chronic rejection can appear in patients with transient mixed chimerism indicating that, without stable chimerism, there will be loss of immunological tolerance with time

Induction of Transplant Tolerance in Clinical Practice • Weaning off immunosuppression or active tolerance induction protocols is not commonplace in routine clinical practice ○ However, ongoing clinical trials are studying adoptive cellular therapies as well as drug therapies for achieving transplant tolerance • Safety evaluation (phase I/II) of "tolerance-inducing" adoptive immunotherapy in renal transplantation (Treg subsets, tolerogenic macrophages, and tolerogenic recipient-derived DCs) has been initiated with multicenter "The ONE Study"

Use of Regulatory Cell Therapies in Clinical Practice • Cellular therapeutic options not routinely available to treat rejection or graft-vs.-host disease (GVHD); however, several in pipeline for evaluation ○ Potential regulatory cells to be used therapeutically in organ transplantation are Tregs, regulatory macrophages, tolerogenic DCs, MDSCs, and mesenchymal stem cells ○ Noncellular approaches to boost Treg function, using drug or immunomodulatory therapies, include – Delaying use of CNIs – Alternatively, other immunosuppression protocols avoiding use of CNIs – Use of immunomodulatory agents, which are primarily depleting in nature, e.g., alemtuzumab (anti-CD52 monoclonal antibody) or polyclonal antithymocyte globulin – Other biological agents, such as mTOR inhibitors (rapamycin, everolimus) or T-cell ostimulation/activation blockers (belatacept, abatacept) ○ Regulatory macrophages isolated from donor allografts administered to 2 living donor renal transplant recipients, showing no adverse effects on allograft function – CNI therapy reduced within 6 months of transplantation ○ Other regulatory macrophage therapy studies performed in small numbers of renal transplant patients, including those receiving deceased donor organs ○ Mesenchymal stem cells (MSCs), typically derived from adipose tissue or less frequently, bone marrow or umbilical cord, used with positive early outcomes in kidney transplantation – Including decreased incidence of acute rejection, reduction in opportunistic infections, and sustained allograft function at 1-year post transplant – Respond differently in various physiological contexts, e.g., □ In noninflammatory settings, MSCs promote tissue regeneration and repair and do not exacerbate immunogenicity □ In presence of inflammatory cytokines, MSCs are highly immunosuppressive and express MHC, allowing effective allograft tolerance – Several clinical trials of MSCs in solid organ transplantation □ Most in kidney but also single trials in lung, liver, and islet cell 10

Summary • Regulation of immune responses and processes is complex phenomenon, involving many different cellular subsets and mechanisms • Careful monitoring of transplant patients over time may benefit identification of type of cellular therapy to use and best time in transplant process for cellular therapy intervention

SELECTED REFERENCES 1.

Wortel CM et al: Regulatory B cells: phenotype, function and role in transplantation. Transpl Immunol. 41:1-9, 2017 2. Scalea JR et al: Transplantation tolerance induction: cell therapies and their mechanisms. Front Immunol. 7:87, 2016 3. Strober S: Use of hematopoietic cell transplants to achieve tolerance in patients with solid organ transplants. Blood. 127(12):1539-43, 2016 4. Pileggi A et al: Mesenchymal stromal (stem) cells to improve solid organ transplant outcome: lessons from the initial clinical trials. Curr Opin Organ Transplant. 18(6):672-81, 2013 5. Scandling JD et al: Tolerance and withdrawal of immunosuppressive drugs in patients given kidney and hematopoietic cell transplants. Am J Transplant. 12(5):1133-45, 2012 6. Schliesser U et al: Tregs: application for solid-organ transplantation. Curr Opin Organ Transplant. 17(1):34-41, 2012 7. Wood KJ et al: Regulatory immune cells in transplantation. Nat Rev Immunol. 12(6):417-30, 2012 8. Newell KA et al: Regulatory cells and cell signatures in clinical transplantation tolerance. Curr Opin Immunol. 23(5):655-9, 2011 9. Li XC et al: An update on regulatory T cells in transplant tolerance and rejection. Nat Rev Nephrol. 6(10):577-83, 2010 10. Donckier V et al: Induction of tolerance in solid organ transplantation: the rationale to develop clinical protocols in liver transplantation. Transplant Proc. 41(2):603-6, 2009 11. Seyfert-Margolis V et al: Marking a path to transplant tolerance. J Clin Invest. 2008 Aug;118(8):2684-6. Erratum in: J Clin Invest. 118(9):3240, 2008 12. Matthews JB et al: Clinical trials of transplant tolerance: slow but steady progress. Am J Transplant. 3(7):794-803, 2003

Regulatory Immune Cells and Transplant Tolerance

Types of Regulatory Immune Cells (Left) A kidney allograft protocol biopsy at 1 year post transplant shows complete absence of inflammation. (Right) Regulatory innate immune cells include myeloidderived suppressor cells (MDSCs), mesenchymal stem cells (MSCs), and regulatory macrophages. MDSCs and MSCs accumulate in allograft from tissue injury-induced inflammation. They provide cellular suppression or produce cytokines and growth factors that promote regulatory T cells (Tregs) and type 1 regulatory T cells (Tr1).

Peripheral Induction of Regulatory T Cells

Immunology

No Rejection

CD8(+) Regulatory T Cells (Left) Induced Tr1 cells do not express FOXP3 protein and are induced by IL-10 from Tregs, tolerogenic dendritic cells (DCs), and regulatory B cells (Bregs) in draining lymphoid tissue or the allograft. Tr1 cells suppress proinflammatory responses of antigen-presenting cells (APCs) and effector T cells. (Right) Naive CD8 T cells can become CD8(+) Treg cells in the presence of IL-10 and functionally act similarly to Tr1 cells. CD8(+), CD28(-) T cells have an immune regulatory function and inhibit APC function.

Double-Negative Regulatory T Cells

Role of Tregs in Organ Transplantation (Left) CD4(-), CD8(-) Tregs ſt downregulate expression of costimulatory molecules on DCs, block proinflammatory immune responses, and induce apoptosis of DCs. (Right) Thymic Tregs suppress ischemia-reperfusion injury and inhibit T-cell proliferation. Tregs promote tolerance through production of IL-10, TGF-β, inhibition of APC function, modulation of energy metabolism, and amino acid availability.

11

Immunology

Regulatory Immune Cells and Transplant Tolerance

Role of Regulatory T Cells

B-Cell Development

Immune Response in Allograft Rejection

B-Cell Depletion Therapies

Negative Effects of B-Cell Depletion

Function of Antithymocyte Globulin

(Left) Tregs suppress immune responses through many different mechanisms, as illustrated. (Right) B cells develop in the bone marrow and then complete the process of differentiation in secondary lymphoid organs. B cells have distinct stages of development and terminally differentiate into antibody-producing plasma cells. Plasma cells can be long lived, return to the bone marrow, and produce specific antibodies. Bregs produce IL-10.

(Left) Immune-activation events in allograft rejection are shown. B cells play a critical and multifunctional role in allograft rejection through their role as APCs, Tcell costimulators, producers of cytokines, and activators of cytotoxic/inflammatory T cells. (Right) B-cell depletion therapies can limit the harmful contribution of B cells in alloimmunity, including antigen presentation to T cells, cytokine production, and antibody production. Bregs may promote allograft tolerance through several mechanisms.

(Left) A potential adverse consequence of B-cell depletion includes depletion of Bregs. The timing of B-cell depletion post transplant is critical to whether B-cell depletion promotes or curtails the alloimmune response. (Right) Antithymocyte globulin, a polyclonal antibody used for induction and treatment of allograft rejection, is not just a T-celldepleting agent. Depending on its dose, it can deplete other lymphocytes and modulate immune responses, as depicted.

12

Regulatory Immune Cells and Transplant Tolerance

Acute Rejection (Left) Clinical applications of regulatory immune cell therapies have great potential in organ transplantation. These approaches can include in vivo expansion of regulatory cells using pharmacological agents or adoptive transfer of ex vivo-expanded cellular subsets. (Right) Immunoperoxidase staining for CD8 in a transplant kidney biopsy shows acute T-cellmediated rejection. Numerous CD8(+) T cells are present in the interstitial infiltrate ﬈. CD4(+) T cells were also present.

T-Cell-Mediated Rejection

Immunology

Therapeutic Uses of Regulatory Cells

CD68 Immunohistochemistry (Left) Immunoperoxidase staining for CD4 in a transplant biopsy shows acute T-cell-mediated rejection. Several CD4(+) T cells are present in the interstitial infiltrate ﬈. CD8(+) T cells were also present. (Right) Immunoperoxidase staining for CD68 shows numerous CD68(+) monocytes/macrophages in the interstitium ﬉ and marginated in glomerular capillaries ﬈ in a case of acute cellular rejection with glomerulitis.

Acute T-Cell-Mediated Rejection

CD4/FOXP3 Immunohistochemistry (Left) This biopsy demonstrates acute T-cellmediated rejection, with interstitial inflammation and tubulitis. A glomerulus also shows glomerulitis ﬈. FOXP3(+) cells are seen in acute T-cell-mediated rejection as well as in focal infiltrates on protocol biopsies in stable allografts. (Right) Double stain for FOXP3 (blue nuclear stain) and CD4 on a kidney protocol biopsy shows CD4(+) T cells, a few of which also express FOXP3 ﬈.

13

Immunology

NK Cells

NK-CELL BIOLOGY Introduction • Human natural killer (NK) cells comprise ~ 15% of circulating lymphocytes ○ Half-life of 7-10 days in blood • NK cells develop primarily in bone marrow (BM) and also secondary lymphoid tissue ○ Occurs in 5-7 stages ○ IL-15 is most critical cytokine for NK-cell development – Other cytokines, such as IL-2, play role in NK maturation • NK cells straddle both innate and adaptive immune responses ○ Early production of cytokines/chemokines and ability to "kill" (cell lysis) target cells without prior sensitization (innate function) ○ Contain complex array of activating and inhibiting receptors ○ Undergo "licensing" to acquire functional competence ○ Develop "memory" (adaptive component) • 2 main mature NK-cell populations ○ Cytotoxic NK cells [CD16(+)] CD56(dim) comprise ~ 90% of circulating NK cells – > 95% of CD56(dim) NK cells express CD16 (FcγRIII) at high levels □ Contributes to their functional relevance ○ Cytokine-producing CD56(+++), CD16(dim) or CD56(+++), CD16(-) NK cells comprise ~ 10% of NK cells in blood – Most (50-70%) CD56(+++) NK cells lack CD16 expression – Smaller proportion of NK cells have low expression of FcγRIII (CD16) ○ CD56 (NCAM) has unknown function on human NK cells – Other surface markers confer unique phenotypic and functional properties to CD56(+++) or CD56(dim) NK cells ○ Resting CD56(dim), CD16(+) NK cells are more cytotoxic than CD56(+++) NK cells

– After IL-2 or IL-12 stimulation, CD56(+++) NK cells can have same cytotoxicity – CD56(dim), CD16(+) NK cells are typically more granular than CD56(+++) NK cells with more cytolytic enzymes within intracellular granules – NK cells also mediate antibody-dependent cellular cytotoxicity (ADCC) when CD16 activated by antibodycoated target cells – CD56(dim) NK cells more terminally differentiated than CD56(+++) NK cells □ Exact development of CD56(dim), CD16(+++) NK cells is not understood – FcγRIII (CD16) acquisition is late NK differentiation event in blood □ Surface CD16 expression correlates with NK-cell maturation – CD56(dim) NK cells may derive from CD56(+++) NK cells, since latter population is 1st to appear post hematopoietic transplantation and cytokine-induced NK differentiation – CD56(dim) NK cell differentiation may be progressive, ultimately resulting in CD94(low), CD62L(-), CD56(dim) phenotype – CD57 expression on CD56(dim) NK cells correlates with functional divergence in this subset – CD57(+), CD56(dim) NK cells are more differentiated with lower cytokine-induced proliferative capacity compared to CD57(-), CD56(dim) NK cells – CD57(+), CD16(+), and CD56(dim) NK cells are important effector cells: Target cell cytolysis and rapid release of proinflammatory cytokines/chemokines on binding of activating receptors ○ CD56(+++) NK cells produce immunoregulatory cytokines, including – IFN-γ – TNF-β – IL-10 – IL-13 – GM-CSF ○ CD56(dim), CD16(+) NK cells produce minimal amounts of cytokines, even after stimulation in vitro

Two Arms of Immune Response (Left) This graphic illustrates the immune response: Innate and adaptive immunity. NK cells are key lymphocytes in innate immune function but also serve as a link between innate and adaptive immunity. (Right) NK cells develop from hematopoietic precursors in the bone marrow. However, NK precursor cells are also present in other tissues and differentiate into immature and mature NK cells, which may recirculate.

14

Development of NK Cells

NK Cells

NK-Cell Memory • Memory-like properties recently observed in human NK cells ○ "Memory" NK response with cytokine preactivation in vitro results in increased production of IFN-γ ○ Surface phenotype of "memory" NK response (enhanced IFN-γ) in one study included CD94(+), NKG2A(+), NKG2C(+), CD69(+), CD57(-), KIR(-) – Another study of CMV reactivation post hematopoietic transplantation showed acute infection resulting in expansion of NKG2C(+) NK cells producing IFN-γ ○ NKG2C(+) NK cells persist (> 1 year post transplant) – Lack NKG2A, express CD158b(+), acquire CD57, and produce IFN-γ – Differences in NK cell memory markers between studies may be due to timing of analysis relative to NK cell memory differentiation ○ CD57(+), NKG2C(+) NK cells expand in solid organ transplant (SOT) patients with active CMV infection ○ Memory-like NK cells appear antigen-specific in response – CD57 is possible marker of "memory" phenotype in NK cells in response to infection – Human memory-like NK cells can persist and contribute to control of infection or neoplasia post transplantation

NK-Cell Education (Tolerance) and Regulation • Rapid NK cell response to cellular targets without prior "exposure" necessitates regulatory mechanisms to prevent unbridled tissue damage • Like B and T cells, NK cells undergo self-tolerance process, but use nonrearranged receptors ○ NK cells undergo host MHC class I-dependent functional maturation process called "licensing" ○ Licensing of NK cells involves inhibitory receptors that recognize MHC class I on target cells and block NK cell activation – Maturation results in 2 types of "tolerant" NK cells: Licensed (functionally competent) and unlicensed (functionally incompetent) – Licensing pairs inhibitory receptor with cognate selfMHC class I ligand for functional development and provides means for tolerance to self-antigens – Each NK cell potentially licensed separately □ Hosts with heterozygous MHC alleles would have individual NK cells that are licensed and inhibited by different MHC molecules ○ Target cells with normal MHC class I expression are more resistant to NK-cell killing than those lacking MHC class I ("missing self" hypothesis) ○ Protection of MHC class I-expressing cells due to MHC class I-specific inhibitory receptors on individual NK cells (KIRs and CD94/NKG2 receptors) – KIRs are highly polymorphic and include multiple haplotypes encoding different numbers of receptors with different alleles for each receptor ○ Potential for breaking NK-cell tolerance, if proinflammatory cytokines are produced and unlicensed NK cells are activated – Most, if not all, NK cells are activated during infection by proinflammatory cytokines • NK cells regulate other components of immune system, e.g., dendritic cells (DCs), B and T cells, endothelial cells ○ NK cells regulate DC homeostasis by killing immature DCs ○ Immature DCs killed by NKp30 receptor-dependent mechanism – Results in attenuation of antigen-specific immunity due to inability of immature DCs to costimulate T cells or secrete cytokines ○ Killing target cells by NK cells results in antigen presentation of apoptotic NK cell targets by DC subsets or stimulation of antigen-specific adaptive immune response ○ NK cells promote DC maturation through IFN-γ and TNF – DC production of IL-12 can reciprocally activate NK cells ○ NK cells influence adaptive immune responses by acting on T and B cells – NK cells can prime CD4(+) cells through production of IFN-γ – NK cytolysis can kill activated T cells with inadequate expression of MHC class I molecules – In vitro NK cells can suppress autoreactive B cells ○ Immunological activation against endothelial cells can lead to vasculopathy and organ dysfunction, including allograft rejection

Immunology

○ Normal proportions of NK-cell subsets alter with aging – CD56(dim), CD16(+) NK cells expand in older individuals □ CD56(+++) NK-cell levels remain unchanged – NK cells in older individuals less responsive to IL-2induced proliferation, likely related to expansion of CD56(dim), CD16(+) NK cells ○ NK cells subclassified as NK1 or NK2 cells, based on cytokine profiles – NK1 cells primarily produce IFN-γ, TNF-β (type 1 cytokines) but can produce IL-10 (type 2 cytokines) – NK2 cells produce IL-15 and IL-13 – Ex vivo, CD56(+++) NK cells produce more type 1 and 2 cytokines compared to CD56(dim) NK cells • 3 major superfamilies of NK cell receptors with activating and inhibitory forms ○ Killer Ig-like receptors (KIRs) recognize primarily HLA-A, HLA-B, and HLA-C ○ C-type lectin superfamily (CD94 and NKG2) recognizes HLA-E ○ Natural cytotoxicity receptors with unknown ligands and other classes of NK receptors including Ig-like transcripts (ILTs) described ○ CD56(+++) NK cells have low to absent KIR and ILT-2 expression, high level of CD94/NKG2A inhibitory receptors – Converse is true of CD56(dim) NK cells ○ Both CD56(+++) and CD56(dim) NK cells express activating receptor, NKG2D, which recognizes MHC class I-related molecules, MICA, and MICB ○ NK-cell subsets differentially express other cell-surface receptors with known or unknown biological significance

15

Immunology

NK Cells – NK cells can bind endothelial cells through ≥ 3 receptors □ α4β1 integrin (VLA4) to VCAM-1 □ CD62L to addressins □ CX3CR1 to CX3CL1 (fractalkine) – CX3CL1 activates NK-cell killing of endothelial cells – NK cells can mediate vascular injury in several pathologic conditions, including CMV infection through initiation of inflammatory response

NK CELLS IN SOLID ORGAN TRANSPLANTATION Overview • NK cells play proinflammatory and regulatory role in SOT ○ Allograft survival may require balance of these functions • NK cells and allograft rejection ○ Activated NK-cell production of IFN-γ augments alloreactive T-cell responses and suppresses Treg function (based on mouse models) ○ NK-cell allograft infiltration often occurs prior to T-cell infiltration – Consistent with role as early innate effector cells responding to inflammatory stimuli ○ If costimulatory signals are blocked, NK cells play important role in mediating graft rejection ○ Recognition of NKG2D (NK-activating receptor) ligands ○ MICA and MICB, expressed on renal and pancreatic allografts, can result in NK activation and correlate with rejection ○ Shedding NKG2D ligands, e.g., soluble MICA, prevents NK-cell activation due to receptor internalization and degradation – NKG2D expressed on activated T cells and macrophages may result in allograft recognition through NKG2D ligands and cause rejection □ Suggests NKG2D ligand induction is relevant for allotransplantation ○ Antigen-presenting cells (APCs) and T cells may be potential targets of NK-cell regulation ○ NK-cell regulation of host DC immunogenicity can contribute to allograft-specific immune response – DCs can become resistant to NK-mediated cell lysis by MHC class I upregulation during DC maturation – Mature DCs can escape NK-mediated cytotoxicity through increased cell surface expression of HLA-E, which binds NKG2A/CD94 (inhibitory receptor) – Mature DCs can promote allogeneic Th1-specific responses ○ NK cells infiltrating renal allografts express cytolytic protein, granzyme A and B – Unclear how cytotoxic NK cells modulate or mediate allograft rejection, based on whether proinflammatory or regulatory role of NK cells is dominant ○ KIR receptor interactions on NK cells with corresponding HLA-C ligands may influence NK cell activation, which corresponds to allograft outcome ○ Activating KIR genotype can help control viral infection (e.g., CMV) in renal transplantation • NK cells and allograft tolerance 16

○ NK cells are essential for tolerance induction to allografts in certain mouse models of islet or renal transplantation ○ Transcriptome analysis of peripheral blood cells from liver transplant patients reveal operational tolerance associated with increased transcripts of genes from NK and γδ T cells ○ Tolerance induction by NK cells possibly related to elimination of allogeneic DCs ○ Mouse islet transplant model shows perforin expression is essential to NK cell role in allograft tolerance – Suggests role for NK cytotoxicity in mediation of tolerance ○ NK-mediated killing of donor-derived APCs likely to be important in allograft tolerance induction • NK cells and immunosuppression ○ Steroids and calcineurin inhibitors downmodulate function of IL-2-activated NK cells – Cyclosporine A induces selective inhibition of IL-2 and IL-15-mediated proliferation of CD56(dim) NK cells – Steroids inhibit NK-cell cytotoxicity by downregulation of surface expression and function of activating receptors and affect granule exocytosis ○ NK-cell lymphopenia reported in allotransplant recipients treated with rapamycin (mTOR inhibitor) ○ Daclizumab, anti-IL-2Rα monoclonal antibody (mAb), used for immunosuppression or induction in certain SOT protocols – Associated with expansion of CD56(+++) NK cells and survival of activated T cells ○ Use of rabbit antithymocyte globulin (rATG) for induction in SOT decreases circulating CD56(+) NK cells between day 1 and day 11 post transplant – rATG and alemtuzumab (anti-CD52 mAb) induce apoptosis of NK cells with induction of proinflammatory cytokines in vitro ○ Since NK cells play key role in innate and potentially adaptive immune responses in allotransplantation, effect of immunosuppressive therapies on NK cells should be monitored

NKT CELLS IN SOT Overview • Natural killer T cells (NKT cells) are CD1d-restricted, glycolipid antigen-reactive immunoregulatory T cells that also express certain NK cell markers ○ Promote immune responses to tumors and pathogens ○ Suppress cell-mediated immunity in autoimmunity and allograft rejection • NKT cells differ from conventional T cells in their expression of T-cell receptor (TCR) that recognizes glycolipids associated with nonpolymorphic CD1d molecule ○ 2 types of NKT cells – Type I express invariant TCR α-chain – Type II express diverse TCR α-chain • NKT cells have regulatory functions that promote transplant tolerance ○ Can interact with regulatory T cells (Tregs) and modulate Treg function (e.g., being through IL-10) ○ In recent model system, Treg and NKT-cell interactions promote tolerance in combined BM and organ transplants

NK Cells

Antigen Expression

CD56 Bright

CD56 Dim

CD56

(++)

(+)

CD16

(-/+)

(++)

KIR

(-/+)

(++)

CD94

(++)

(-/+)

NKG2A

(+)

(-/+)

ILT-2

(-)

(+)

IL-2Rαβγ

(+)

(-)

IL-2Rβγ

(+)

(+)

C-Kit

(+)

(-)

IL-1RAcP

(+)

(+)

IL-1RI

(+)

(-/+)

IL-18R

(+)

(-/+)

CCR7

(++)

(-)

CXCR3

(+)

(-/+)

CXCR1

(-)

(++)

CX3CR1

(-)

(++)

CD2

(++)

(+)

L-selection (CD62L)

(++)

(-/+)

PENS-PSGL-1

(-)

(+)

LFA-1

(+)

(++)

CD44

(++)

(+)

CD49e

(++)

(+)

Immunology

Expression Profile of Mature NK-Cell Populations

NK Receptors

Cytokine and Chemokine Receptors

Adhesion Molecules

Adapted from Cooper MA et al: The Biology of Human Natural Killer-Cell Subsets. Trends in Immunol. 22:633-40, 2001.

– Mediated in part through IL-4 production by host NKT cells, which regulate PD-1 expression on host Treg and conventional CD4(+) and CD8(+) T cells

8.

Freud AG et al: Human natural killer cell development. Immunol Rev. 214:5672, 2006

SUMMARY Role of NK Cells in SOT • T cells and B cells long recognized as key players in maintenance of allografts, but role of NK cells is emerging and important • NK cells play roles in infection control post SOT, allograft survival, and regulation of immune response to immunosuppression

SELECTED REFERENCES 1. 2. 3. 4. 5. 6. 7.

Chu X et al: Islet allograft tolerance in the absence of invariant natural killer T cells. Clin Immunol. 141(3):268-72, 2011 Paust S et al: Natural killer cell memory. Nat Immunol. 12(6):500-8, 2011 Godfrey DI et al: Raising the NKT cell family. Nat Immunol. 11(3):197-206, 2010 Morteau O et al: Renal transplant immunosuppression impairs natural killer cell function in vitro and in vivo. PLoS One. 5(10):e13294, 2010 van der Touw W et al: Natural killer cells and the immune response in solid organ transplantation. Am J Transplant. 10(6):1354-8, 2010 Pratschke J et al: Role of NK and NKT cells in solid organ transplantation. Transpl Int. 22(9):859-68, 2009 Vivier E et al: Functions of natural killer cells. Nat Immunol. 9(5):503-10, 2008

17

Immunology

NK Cells

Identification of Lymphocytes in Blood

Identification of CD3(+) and CD3(-) Lymphocytes

Identification of CD4(+) and CD8(+) T Cells

Identification of B and NK Cells by Flow

NK-Cell Subsets in Healthy Controls

Identification of NK-Cell Subsets in Patient

(Left) Quantitation of T-, B-, and NK-cell lymphocyte subsets is performed by 6color flow cytometry. CD45(+) lymphocytes, shown in a "gate" (defined population in red), are identified by a combination of side scatter (measuring cellular complexity) and CD45 expression intensity. (Right) Separation of CD3(+) and CD3(-) lymphocytes by flow cytometry allows additional cellular subset identification. CD3(+) T cells are shown on the right and CD3(-) B and NK cells are shown on the left.

(Left) CD3(+) T cells are further differentiated by flow cytometry into CD8(+) T cells (lower right quadrant) and CD4(+) T cells (top left quadrant). (Right) CD3(-) lymphocytes are further subdivided into total NK cells with CD16(+) and CD56(+) (lower right quadrant) and CD19(+) (top left quadrant) B cells.

(Left) Quantitative NK-cell subset immunophenotyping using CD56 and CD16 identifies the 2 major NK cell subsets: CD56(dim) and CD16(+) (middle right), and CD56(bright) and CD16(+/-) (middle left and top). (Right) Quantitative NK-cell subset immunophenotyping using CD56 and CD16 identifies the 2 major NK-cell subsets in a patient with NK-cell lymphocytosis: CD56(dim) and CD16(+) (middle right), and CD56(bright) and CD16(+/-) (middle left and top).

18

NK Cells

NK-Cell Subsets in Blood (Left) NK cells regulate immune responses by boosting (red arrows) maturation/activation of DCs, T cells, and macrophages through a combination of surface receptors and cytokines, or killing (green arrows) of immature DCs, activated CD4 T cells, and hyperactivated macrophages. (Right) CD56(+++), CD16(+/-) (top left) and CD16(+++), CD56(+) (bottom right) NK-cell subsets are cytokineproducing and cytotoxic NK cells, respectively.

Licensing of NK Cells

Immunology

NK-Cell Regulation of Immune Response

NK-Cell Functions and NK-Cell Memory (Left) NK cells undergo a MHC class I-dependent functional maturation process called "licensing." Licensing blocks activation of effector NK-cell responses, resulting in 2 types of tolerant NK cells: Functionally competent (licensed; top) and functionally incompetent (unlicensed; bottom) NK cells. (Right) Graphic shows the innate (top) and adaptive (bottom) nature of NK cells. A parallel is drawn between the latter and antigen-specific memory T cells (middle).

Cellular Degranulation

Antibody-Mediated Rejection via Antibody-Dependent Cellular Cytotoxicity (Left) NK cells contain cytotoxic granules that mediate NK-cell effector function. Cytotoxic granules contain cytotoxic proteins such as perforin, and granzymes A and B. The surface of cytotoxic granules express CD107a/b. During degranulation, the granule membrane merges with the cell membrane, exposing CD107a/b prior to granule release, which can be detected by flow cytometry. (Right) NK cells play a role in donorspecific antibody-induced chronic rejection via Fc receptors.

19

Immunology

Complement

ROLE OF COMPLEMENT IN ATYPICAL HEMOLYTIC UREMIC SYNDROME AND THROMBOTIC MICROANGIOPATHY Introduction • Complement system consists of ○ Several fluid-phase proteins ○ Cell membrane-bound proteins • Circulating complement proteins are synthesized in liver ○ Usually present in inactive state (proenzyme) • Cell-bound proteins are receptors and regulators of complement pathway • Complement cascade can be activated by 3 different mechanisms ○ Classic pathway ○ Alternate pathway ○ Mannose-binding lectin pathway • All complement activation pathways converge at generation of C3 and C5 convertase

• C5b deposition on target results in membrane attack complex (MAC) formation and target lysis ○ Generated by cleavage of C5 into C5a and C5b • This chapter focuses on role of complement alternate pathway defects in pathogenesis of atypical hemolytic uremic syndrome (aHUS) and thrombotic microangiopathy (TMA)

Atypical Hemolytic Uremic Syndrome • Hemolytic uremic syndrome characterized by triad of hemolytic anemia, thrombocytopenia, and renal impairment ○ Atypical HUS defined as non-Shiga-toxin HUS, caused by defects in alternate pathway of complement – 60-70% of aHUS cases associated with pathogenic genetic variants &/or single nucleotide polymorphisms (SNPs) in complement genes or due to autoantibodies to factor H – Pathogenic genetic variants in complement regulatory proteins result in loss-of-function (LOF)

Complement Alternate Pathway Defects and Transplantation

Atypical hemolytic uremic syndrome (aHUS) is multifactorial in etiology and associated with genetic defects in the alternate pathway of complement. There is a high risk of recurrence post transplantation, depending on the underlying genetic defect. De novo aHUS can develop in a small subset of patients post transplantation. Other factors, in addition to genetic predisposition, also modulate the risk of developing aHUS and its recurrence.

20

Complement

Regulation and Dysregulation of Alternate Complement Pathway • During regulated activation of alternate pathway, following events take place ○ Complement factor H (FH) competes with complement factor B (FB) to bind C3b – Results in inhibition of generation of C3 convertase ○ FH binds glycosaminoglycans on endothelial surfaces ○ Membrane cofactor protein (MCP or CD46) acts as cofactor for complement factor I (FI)-mediated cleavage of C3b to generate iC3b ○ Thrombomodulin binds to C3b and FH and accelerates FI-mediated inactivation of C3 • During dysregulated activation of alternate pathway ○ Uncontrolled activation of alternate pathway leads to generation of membrane attack complex (MAC, C5b-C9), due to any combination of – LOF or GOF genetic variants in complement regulatory genes – Autoantibodies to FH ○ Mediated by action of complement factor B and complement factor D and generation of C3 and C5 convertases ○ Resulting injury and endothelial cell activation initiates thrombotic microangiopathic process

Genetic Defects in Atypical Hemolytic Uremic Syndrome/Thrombotic Microangiopathy • Pathogenic variants in CFH, CFHR1, CFHR3, CFHR5, CFI, CFB, CFD, MCP (CD46), C3, THBD, PLG, DGKE associated with aHUS • Penetrance for variants in CFH, CFI, MCP, C3, CFB, and THBD is ~ 50% • ~ 10% of aHUS patients have combination of 2 pathogenic variants (digenic) ○ Suggests disease may result from additive effects of multiple genetic factors • In addition, SNPs on other complement genes, such as C4bp (C4 binding protein), CFHR1, MCP and CFH reported at higher frequency in aHUS patients compared to healthy controls ○ Such SNPs may constitute minor susceptibility to aHUS – Could influence development and severity of disease • Disease-affected family members with inherited variantrelated aHUS have other genetic susceptibility factors for aHUS ○ While healthy carriers with same variant do not have additional susceptibility factors • Challenging to assess influence of each SNP and gene-gene interactions in specific patients ○ Because penetrance and expression of aHUS influenced by epigenetic and environmental factors

• Genetic testing for several of complement regulatory genes complicated by extensive sequence homology between genes, e.g., CFH and CFHR genes ○ CFH gene variants are most common in aHUS ○ CFH encodes FH protein – Has 20 globular domains, called short consensus repeats ○ By alternative splicing, CFH also encodes smaller protein, called FH-like 1 (FHL-1) ○ Close to CFH gene locus, there are 5 genes encoding proteins structurally related to CFH, called CFH-related proteins (CFHR1-5) ○ CFHR1-5 region contains multiple genomic duplications – Render this area susceptible to genomic rearrangements, such as gene conversion or nonhomologous recombination ○ Some rearrangements associated with aHUS, e.g., CFHCFHR1 hybrid gene ○ Homozygous deletion of CFHR1-CFHR3 associated with presence of FH autoantibodies – While heterozygous deletions are frequently observed ○ Rearrangements cannot be identified by standard nextgeneration sequencing techniques and require other analytical methods that evaluate for deletion/duplications and copy number variations • Multicenter cohorts assessing 795 aHUS patients screened for CFH, CFI, MCP, CFB, and C3 gene variants ○ 3.4% of patients had combined pathogenic variants in > 1 gene (digenic or trigenic) ○ 40.6% of patients had monogenic (single gene) defect ○ MCP and CFI variants are frequently present only in context of other gene variants – Likely indicating low pathogenic potential individually ○ Monogenic defects in CFH, C3, or CFB are sufficient to induce aHUS compared to MCP or CFI variants ○ Penetrance increased significantly with pathogenic variants in 1, 2, or 3 genes ○ Disease-risk haplotypes in CFH (c.1-332C>T, c.2808G>T) and MCPggaac (c.897T>C) – When present in at least 1 copy, significantly increases risk of penetrance in digenic patients, though penetrance still remains incomplete • Familial occurrence of aHUS observed in 20% of pedigrees ○ Inheritance can be either autosomal dominant or autosomal recessive • Age at onset in adults is similar, regardless of genetic defect, while in children, it varies depending on complement anomaly, with CFH, CFI and DGKE variants typically observed in young infants

Immunology

– Pathogenic genetic variants in complement activation proteins result in gain-of-function (GOF) ○ Additionally, pathogenic genetic variants in other proteins in coagulation pathway related to aHUS phenotype, such as – Thrombomodulin (THBD) – Plasminogen (PLG) – Diacylglycerol kinase epsilon (DGKE)

Atypical Hemolytic Uremic Syndrome and Transplant • Pathogenic variants in CFH, CFI, and MCP identified in 30% of de novo posttransplant HUS ○ Likely represents undiagnosed aHUS as original cause or contributing factor of end-stage renal disease (ESRD) • Risk for aHUS recurrence postrenal transplantation is low for MCP variants (~ 15%), yet high (~ 80%) for circulating complement factor genetic variants ○ MCP present in allograft and may supplement deficiency ○ Most circulating complement regulatory factors synthesized by liver 21

Immunology

Complement

•

•

•

•

•

•

•

•

– Successful reports of simultaneous liver and kidney transplantation Other contributory or modulatory factors, including ○ Posttransplant immunosuppression, particularly calcineurin inhibitors (CNI) ○ Infections ○ Ischemia-reperfusion injury ○ Allograft rejection Etiology of disparity in aHUS recurrence after transplantation related to underlying genetic defect is unknown Other risk factors associated with recurrent aHUS in renal allograft include ○ Duration of disease prior to transplant or renal failure ○ Living-related donor renal transplants ○ Use of CNI (particularly cyclosporine) De novo aHUS can affect 3-14% of renal transplant recipients ○ Modulated by immunosuppression, infections, and antibody-mediated rejection ○ Probably represents previously undiagnosed aHUS in recipient ○ MCP mutations in allograft cannot be excluded Recurrent aHUS in transplanted organ can be minimized by ○ Screening for genetic defects in complement regulatory genes ○ Appropriate selection of donor (avoiding living-related donors) ○ Prophylactic therapy Most complement regulatory proteins primarily produced in liver ○ Kidney transplantation alone to correct end-stage renal disease does not prevent recurrence ○ Liver transplantation as potential option to correct underlying genetic complement abnormality and prevent disease recurrence – Recommendations that in patients with CFH or CFI genetic variants, combined liver-kidney (if there is also ESRD) or isolated liver transplant (if renal function is normal or has recovered) should be considered No clear guidelines on whether combined liver-kidney or liver alone transplants should be considered for patients with genetic variants in other complement regulatory genes Liver transplant, combined with renal, or in isolation, is not widespread procedure for long-term correction and management of aHUS

Eculizumab and Atypical Hemolytic Uremic Syndrome • Eculizumab (Soliris, Alexion) is humanized monoclonal antibody targeting terminal complement pathway ○ Binds with high affinity to complement protein, C5, and prevents generation of anaphylotoxin, C5a, and MAC (C5b-C9) • Several studies have demonstrated its efficacy in controlling complement activation in patients with aHUS pre- and post transplantation • In patients with CFH or CFI variants and aHUS, eculizumab used pre transplantation and post transplantation ○ Duration of administration in latter setting is unknown 22

• Eculizumab is very expensive ○ Alternate approaches to wean off have been studied, though long-term, large cohort data are still lacking • One alternate strategy to adopt combined liver-kidney transplant in patients with ESRD, but this has substantial risk ○ Often need intensive plasmapheresis in pre- and perioperative period to ensure optimal complement levels • Standard dosing of eculizumab for recurrent aHUS is 900 mg, IV, every week for 4 weeks, followed by 1200 mg biweekly • Duration of eculizumab treatment has not been optimized ○ Many centers continue it indefinitely, but determination should be made on case-by-case basis • Patients with persistent symptoms on eculizumab may also be treated with either plasmapheresis or plasma infusion • Eculizumab treatment carries risk for life-threatening meningococcal infections, due to blockade of terminal pathway of complement activation ○ Patients considered for eculizumab therapy should receive meningococcal vaccine at least 2 weeks prior to initiation of therapy, and additionally, antimicrobial prophylaxis can be considered ○ If urgent eculizumab therapy indicated, antimicrobial prophylaxis can be administered simultaneously

Laboratory Testing for Diagnosis of Atypical Hemolytic Uremic Syndrome and Monitoring of Eculizumab • Patients who manifest features of HUS, should be evaluated to eliminate possibility of Shiga toxin-associated HUS • ADAMTS13 level and activity should also be measured to assess for thrombotic thrombocytopenic purpura (TTP) • If ADAMTS13 results are > 5-10%, then investigation for aHUS should include ○ Serological (serum or plasma) measurement of AH50 (functional assessment of alternate complement pathway) ○ C3 and C4 levels ○ Measurement of FH, FI, and FB levels, factor H autoantibodies, and sMAC levels – Provide measure of terminal activation of complement) ○ MCP (CD46) expression can be measured on surface of leukocytes by flow cytometry ○ Genetic testing for variants in complement regulatory genes – Should include deletion/duplication analysis and Sanger sequencing for homologous regions ○ Essential for complement serological analysis to collect and transport blood specimens appropriately for further processing and analysis – Due to likelihood of in vitro complement activation and consumption by sample handling ○ Additionally, split products, such as C3Bb and C4d can also be assessed as markers of complement activation • Normal range for plasma FH is large and influenced by genetic and environmental factors ○ Challenging to detect heterozygous FH deficiency by plasma measurements

Complement 6.

7. 8.

9.

Kavanagh D et al: Atypical hemolytic uremic syndrome, genetic basis, and clinical manifestations. Hematology Am Soc Hematol Educ Program. 2011:15-20, 2011 Loirat C et al: Atypical hemolytic uremic syndrome. Orphanet J Rare Dis. 6:60, 2011 Hakobyan S et al: Variant-specific quantification of factor H in plasma identifies null alleles associated with atypical hemolytic uremic syndrome. Kidney Int. 78(8):782-8, 2010 Saland JM et al: Liver-kidney transplantation to cure atypical hemolytic uremic syndrome. J Am Soc Nephrol. 20(5):940-9, 2009

Immunology

• Specific reagents (monoclonal antibodies) available that detect 2 allotypes of FH - Y402 and H402 utilizing common CFH SNP, Y402H (Tyr402His) in SCR7 domain of CFH and CFHL1 ○ These monoclonal antibodies allow determination of contribution of each CFH allele, in heterozygous Y402H individuals, to total plasma FH and FHL-1 protein pool ○ This method allows rapid and simple identification of patients carrying CFH null alleles (who may be missed by standard FH plasma quantitation methods) ○ Also allows quick assessment of impact of specific CFH variants on protein expression • Eculizumab treatment can be monitored by measuring C5 protein level and C5 function ○ In addition, there is new mass spectrometry assay that can quantify eculizumab drug, which in conjunction with C5 level and functional monitoring can provide insight into dosage and potential duration of administration ○ May indirectly provide information on presence of antidrug antibodies in individuals who may appear refractory to therapy after prolonged use, though there is no direct way of determining this

Summary • Atypical HUS represents multifactorial etiology, which is initiated by trigger events, including infections, drugs, malignancy, transplantation, pregnancy, and autoimmune diseases ○ Results in endothelial cell injury and thrombi in small vessels ○ Genetic variants in coagulation genes, such as PLG, ADAMTS13, and DKGE can contribute to pathology at this stage ○ Direct lysis by MAC and thrombus formation can cause mechanical damage to erythrocytes, which in presence of genetic variants in complement regulatory genes, e.g. CFH, CFI, C3, CFB, MCP, or autoantibodies to FH, result in overactivation of complement • aHUS can lead to ESRD, which requires treatment with either eculizumab, &/or renal transplantation • Duration of eculizumab treatment in individuals with genetic aHUS can be lifelong, unless combined liver-kidney or liver transplant is performed in suitable patients under appropriate conditions • Decision to discontinue eculizumab treatment pre or post renal transplantation has to be made on case-by-case basis, depending on clinical context, and genetic defect • Laboratory studies can help with diagnosis of aHUS, as well as monitoring of eculizumab therapy

SELECTED REFERENCES 1. 2.

3. 4. 5.

Coppo R et al: Liver transplantation for aHUS: still needed in the eculizumab era? Pediatr Nephrol. 31(5):759-68, 2016 Go RS et al: Thrombotic microangiopathy care pathway: a consensus statement for the Mayo Clinic complement alternative pathway-thrombotic microangiopathy (CAP-TMA) disease-oriented group. Mayo Clin Proc. 91(9):1189-211, 2016 Kolev M et al: Complement--tapping into new sites and effector systems. Nat Rev Immunol. 14(12):811-20, 2014 Saland J: Liver-kidney transplantation to cure atypical HUS: still an option post-eculizumab? Pediatr Nephrol. 29(3):329-32, 2014 Le Quintrec M et al: Complement genes strongly predict recurrence and graft outcome in adult renal transplant recipients with atypical hemolytic and uremic syndrome. Am J Transplant. 13(3):663-75, 2013

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Immunology

Laboratory-Based Immune Monitoring in Organ Transplantation

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INTRODUCTION Immune Monitoring in Organ Transplantation • Immunosuppression (IS) for allograft maintenance remains fundamental concept in solid organ transplantation (SOT) • Advances in IS make SOT viable option for end-stage disease or in contexts with significant morbidity/poor quality of life (e.g., face transplant, hand transplant) • Struggle to find correct balance between over- and underIS, each with its consequences ○ Maintenance of allograft with over-IS leads to significant infections and malignancy with increased morbidity and mortality ○ Conversely, decreased IS in absence of active immune tolerance results in allograft rejection • Personalized IS remains "Holy Grail" of transplantation • Immunosuppressive regimens in SOT divided into ○ Induction IS ○ Maintenance IS • Pharmacokinetic &/or pharmacodynamic monitoring of therapeutic IS ○ Widely accepted, yet incomplete approach of conventional immune monitoring • More robust and comprehensive approach to IS in SOT involves systematic and thoughtful use of immune monitoring ○ Immune monitoring is complete qualitative and quantitative assessment of immunological function and phenotype following use of immune-modifying agents (IS) ○ Balance needs to be achieved in IS – Long-term maintenance of allograft without significant compromise to immune function – Can result in high rates of infection &/or neoplasia ○ Assessment of immunological function and immunophenotype involves use of several laboratory assays typically offered in high-complexity clinical immunology laboratories • Monitoring based on pharmacokinetics (drug trough levels) does not necessarily correlate with immune function ○ Infection, rejection, and neoplasia remain significant risks, especially in multiple transplant recipients (same organ or different organs) • Future approach to effective management of IS is immunological titration of IS instead of pharmacological titration • Multiple biomarkers available to measure immune monitoring, and these reflect donor-specific and nondonor-specific immune activation states in transplant recipients, pre - and postinitiation of IS ○ Biomarkers can be broadly differentiated into – Biomarkers of rejection (immune activation) – Tolerance ○ Also, specific drug target molecules that are immunologically relevant (i.e., expressed on cells of immune system) may be useful in assessing response to specific IS drug

LABORATORY APPROACHES TO IMMUNE MONITORING Global Immunophenotyping Analysis by Flow Cytometry to Monitor Induction Therapies in SOT • Majority of induction approaches involve T-cell manipulation via direct depletion &/or functional suppression ○ Evaluation of T-cell counts to adjust induction therapy for each patient is critical • Antithymocyte globulin (ATG), rabbit or horse ○ Polyclonal antibody used frequently for T-cell depletion in induction strategies • Quantitation of lymphocyte subsets ○ T cells (CD3, CD4, and CD8), B cells [CD19(+)], and NK cells [CD16(+), CD56(+)] by multicolor flow cytometry ○ Allows accurate measurement of specific subset counts following induction therapy used • Absolute lymphocyte count (ALC) from CBC/differential often used to determine overall immune status in transplant patients ○ However, can have normal ALC with significant numerical "alterations" in lymphocyte subsets because lymphopenia in 1 subset associated with homeostatic expansion of other related subset(s) ○ Quantitative skewing of individual lymphocyte subsets (CD4 and CD8 T cells, B cells, and NK cells) only determined by flow cytometry • Lymphocyte subset analysis to titrate dosage prevents other consequences of over-IS, e.g., hypogammaglobulinemia due to B-cell depletion with ATG ○ Hypogammaglobulinemia in SOT associated with increased risk of opportunistic infections (< 6 months post transplant) ○ Hypogammaglobulinemia follows increased IS for rejection episodes ○ Hypogammaglobulinemia assessed post induction and during maintenance IS by immunoglobulin quantitation (IgG, IgA, and IgM isotypes) – Evaluation can enable use of replacement immunoglobulin therapy to manage severe hypogammaglobulinemia and ameliorate consequences • Other immunophenotyping assays include quantitation of regulatory T cells (Tregs) and conventional T-cell subsets ○ Naive vs. memory and activated T cells to predict risk of rejection and infection ○ Interpretation of quantitative immunophenotyping data, e.g., Treg or T-cell subsets, during serial monitoring must account for changes in parent T-cell populations [CD4(+) &/or CD8(+)] due to alteration in immunosuppressive therapy over time ○ If IS changed during time interval of immune monitoring, any change in total CD4(+) or CD8(+) T-cell counts can potentially affect both relative frequencies (%) and absolute counts of various T-cell or Treg subsets • Quantitation of NK-cell subsets

Laboratory-Based Immune Monitoring in Organ Transplantation Molecular Methods to Assess Allograft Status

• Immunophenotyping and quantitation of various lymphocyte subsets is critical for rapid adjustments in induction IS, but functional analysis also important for immune monitoring ○ Preservation of numerical integrity of lymphocyte subsets (whether by ALC analysis or flow cytometry) does not necessarily correlate with functional immune competence ○ SOT patients with normal ALC can have significantly impaired T-cell function, while patients with reduced ALC can have relatively more preserved function • Commonly used measurements for global T-cell function ○ Assessment of lymphocyte proliferation to mitogenic stimuli [typically phytohemagglutinin (PHA)] – Ideally performed by flow cytometry in select laboratories, though most labs use more traditional approach with tritiated thymidine (radioactive marker) – Measures T-cell proliferation to polyclonal mitogen; PHA broadly stimulates T cells to divide and proliferate regardless of T-cell antigenic specificity – Flow cytometric assay offers greater analytical sensitivity than radioactive method in patients with lymphopenia – Flow assay not limited by cellular dilution in lymphopenia and permits visualization of dividing cells and specific cellular markers to identify T cells ○ T-cell activation by analysis of cytokine production – Typically, when activated, CD4 and CD8 T cells produce various cytokines (IFN-γ, IL-2, TNF-α) in temporally regulated manner (polyfunctional T cells) ○ Assays measuring proliferation with PHA for shorter periods of time using purified T cells (CD3 or CD4 ± CD8) and surrogate markers for cellular DNA synthesis run risk of underestimating true T-cell proliferative response • Functional immune competence has many components; no single assay can be used for this purpose • Most robust hallmark of severe functional T-cell compromise is presence of opportunistic infections • T-cell proliferation may also be measured to other global stimuli, such as anti-CD3 with additional costimulation using anti-CD28 or exogenous IL-2 • Impairment in proliferative responses to mitogens, such as PHA (or anti-CD3) is diagnostically less sensitive but more specific test of abnormal T-cell function • Evaluation of antigen-specific T-cell responses considered diagnostically more sensitive but less specific indicator of impaired T-cell function • NK-cell function measured using assays for spontaneous NK cytotoxicity, cytokine-stimulated cytotoxicity, and antibodydependent cellular cytotoxicity • Cellular degranulation is another component used to measure both CD8(+) T-cell and NK-cell function ○ Degranulation assay involves flow cytometric measurement of CD107a/b (proteins expressed on surface of intracellular cytotoxic granules present in CD8 T cells and NK cells) • B-cell function typically measured by immunoglobulin isotype quantitation and assessment of antibody responses to vaccine antigens

• Push toward personalized IS in SOT demands greater investment in newer molecular techniques of mRNA and microRNA analysis, proteomics, and metabolomics for noninvasive allograft analysis • Recent studies use molecular approaches (gene expression methods) to develop scoring systems for predicting rejection, but robust clinical translation data still lacking ○ Transcript (mRNA) analysis of individual gene products in body fluids other than blood reported in specific SOT research protocols but not yet gained acceptance in clinical practice ○ In renal and cardiac transplants, molecular scores or classifiers based on limited gene expression sets reported to be useful for predicting allograft loss or rejection events ○ Donor-derived DNA in plasma of transplant patients also suggested as potential source for monitoring graft rejection, though like other molecular methods, active clinical use yet to come

Immunology

Global Immune Function Analysis

Other Immunophenotyping Tests to Predict Transplant Rejection Risk • Proprietary test (Pleximmune®) to measure inflammatory immune response of recipient T cells to donor cells in lymphocyte co-culture ○ Measures frequency of cytotoxic (CD8+) memory T cells [CD8(+)], expressing CD40L (CD154) in response to donor cell stimulation – Data expressed as immunoreactivity index, based on ratio of recipient CD8(+) CD40L(+) T cells to reference sample (non-HLA related to recipient and donor) expressing same markers ○ Broad applicability across multiple solid organs, and patient ages have yet to be established

IMMUNE RESPONSE BIOMARKERS Immune Activation Markers (Donor-Specific and Nondonor-Specific) • Soluble biomarkers ○ Antibodies to HLA and other molecules ○ Cytokines ○ Chemokines ○ Complement mediators and byproducts of complement activation ○ Other soluble proteins (e.g., membrane-bound proteins that are shed; inflammation-stimulating proteins) ○ Metabolic markers • Cellular biomarkers ○ Cell surface molecules on cells of adaptive and innate immune system

Immune Tolerance Markers • Soluble markers of transplant tolerance • Cellular markers of transplant tolerance ○ Expansion of regulatory Tregs (natural and induced) ○ Regulatory B cells (Bregs) ○ Other regulatory cells producing IL-10

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Immunology

Laboratory-Based Immune Monitoring in Organ Transplantation Other Parameters of Clinically Monitoring Immune Response to Immunosuppression in SOT • Monitoring biologically relevant molecules that are IS drugspecific [calcineurin inhibitors (CNI), Basiliximab, Daclizumab, Belatacept, etc.] • IL-2 production by activated T cells (CNI) • T-cell proliferation to anti-CD3(+), anti-CD28(+), or antiCD3(+) IL-2 (CNI) • Expression of CD25 on T cells (activated) (anti-CD25Rx) • Presence of CD57(+) CD4(+) T cells pretransplant (Belatacept)

INFECTIONS IN SOT Antigen-Specific Assessment of T-Cell Immune Competence (to CMV, EBV, and BKV) • Cytomegalovirus ○ CMV and herpesvirus pose major risk factors in SOT and are associated with significant morbidity and preventable mortality – Seroprevalence is 30-97% depending on population ○ Highest risk of CMV infection in donor seropositive [D(+)], recipient seronegative [R(-)] group ○ Lowest risk in D-negative/R-negative group of patients ○ Lung and small intestine organ recipients are at highest risk among SOT ○ Current laboratory methods focus on diagnosis of CMV infection: Histopathology, culture, serology, antigenemia, and molecular assays for viral nucleic acid quantitation ○ Immunological assays include CMV-specific CD8 T-cell quantitation and functional analysis using flow cytometry or cytokine production analysis – Flow cytometric assay currently available in clinical lab uses special reagents called MHC tetramers for antigen-specific CD8 T-cell quantitation – MHC tetramers: 4 soluble MHC class I molecules bound to specific antigenic peptide (e.g., CMV peptide) and linked by streptavidin molecule conjugated to fluorophore – MHC class II tetramers can be made for quantitation of antigen-specific CD4 T cells – Different tetramers (MHC class I) can be prepared for different antigenic peptides – 5 distinct MHC class I tetramers available for CMV – Peptides used in tetramer must bind specific MHC class I molecule with high affinity – Tetramer-based flow cytometric quantitation of antigen-specific CD8 T cells requires a priori knowledge of HLA class I alleles of patient – Also, functional status of antigen-specific (CMVpeptide-specific) T cells can be ascertained by flow cytometry – Involves stimulating peripheral blood mononuclear cells (including T cells) with specific CMV peptide(s) – Followed by measuring production of IFN-γ (marker of CD8 T-cell activation) and CD107a/b (marker of cellular degranulation)

26

– Both quantitation and functional analysis essential to assess CMV-specific T-cell immunity; antigen-specific T cells should be numerically present and functionally competent □ CMV-CD8(+) T-cell immune competence (CMV-TIC) score developed to incorporate functional components (CD107a/b expression, also referred to as cellular degranulation, and IFN-γ production), if CMV-specific CD8(+) T cells are ≥ 2 cells/μL. □ Global CD8(+) T-cell immune competence assessment performed by measuring production of IFN-γ and CD107a/b in CD8(+) T cells after PMA and ionomycin (nonspecific mitogenic stimulant) □ Retrospective review (unpublished observation, Meesing, Abraham and Razonable, 2018) of clinical utility of score was established in multiple SOT patients (130, of whom 72 had active CMV infection or disease; 64 lung and heart-lung recipients) □ Both CMV-TIC score and global immune competence assessment associated with clinical course of CMV infection and disease after SOT – CD8(+) T cells key effector cells in antiviral immune response, especially to CMV – Cytotoxic CD4(+) T cells and NK cells also important participants in antiviral immune response – Reliance on specific MHC tetramers restricts use of assay to patients with appropriate MHC class I or II alleles – MHC tetramer reagents limit epitope specificity of antigenic peptides used in assay to those peptides with high affinity for specific MHC allele – Broader peptide coverage necessitates more tetramer combinations leading to increased expense – Focuses solely on antiviral CD8 T cells, discounts role of CD4 cytotoxic T cells and NK cells in maintaining robust immune response to these pathogens – Future assays will focus on using more comprehensive strategy, non-MHC-restricted, broader array of antigenic peptides, and evaluating both CD4 and CD8 cytotoxic T cells – Alternative to flow cytometry/tetramer analysis is measurement of cytokine production by antigenspecific activated CD8(+) T cells after stimulation with CMV peptides – Cytokine production may be measured by ELISPOT (quantifies specific cytokine-producing CMV-specific CD8 T cells) or direct quantitation using ELISA techniques/immunoassays – QuantiFERON-CMV assay: Commercially available assay, detects IFN-γ secreted by CMV-specific CD8 T cells after whole blood stimulation with specific antigenic peptides – Both QuantiFERON-CMV assay and ELISPOT technique share some of same limitations as tetramer flow cytometry assay – Limited repertoire of CMV peptides used for stimulation (restricts assessment of breadth of CMV immune response) and focuses only on CD8 T-cell response (contribution of CMV-specific CD4 cytotoxic T cells not determined) • EBV and posttransplant lymphoproliferative disorder (PTLD)

Laboratory-Based Immune Monitoring in Organ Transplantation ○ Laboratory methods used to diagnose and guide IS reduction based on screening for BKV nucleic acid in urine and blood and immunohistopathology ○ No clinical diagnostic laboratory tests currently available to assess BKV-specific cellular immunity ○ Future assays for BKV-specific T-cell responses require broader and more comprehensive approach that is nonMHC-restricted and covers peptide epitopes for major protein components of virus

Immunology

○ PTLD: One of most devastating complications of SOT and EBV infection; found in majority of B-cell PTLD occurring within 1st year post transplant – EBV-PTLD characterized by lymphoproliferation post transplant; may or may not be symptomatic – PTLD develops early (< 12 months post transplant) or late (> 12 months post transplant) – Risk factors for early PTLD include primary EBV infection, type of organ transplant, use of T-cell or lymphocyte-depleting agents, age of recipient (especially young), CMV mismatch or disease – Risk factors for late PTLD include duration of IS, type of organ transplanted, and older recipient age ○ Similar to CMV, standard laboratory tests focus on pathogen and not host immune response ○ Laboratory tests for diagnosis of EBV infection include – CBC and differential – EBV viral load (PCR-based) – If EBV infection is present, EBV serology to determine past exposure – Histopathology and imaging ○ Assessment of host immune response just as relevant as with CMV infection – EBV-specific MHC class I tetramers commercially available for research purposes, not yet in routine clinical diagnostic use – Same limitations of tetramer approaches apply, as with CMV ○ Future clinical assay development focused on developing non-MHC-restricted flow cytometric assays assessing functional status of EBV-specific CD4(+) and CD8(+) cytotoxic T cells post stimulation with broad array of EBV peptides ○ NK cells are essential to control of EBV infection, as with CMV – Diagnostic test development to assess antigenspecific (CMV or EBV) NK-cell function is much more challenging to develop in clinical laboratory – Possible to use immunophenotyping flow cytometry approaches to quantitatively assess NK cell subsets (cytotoxic vs. cytokine-producing) and NKT cells in patients with active CMV or EBV infection or viremia • BK virus ○ Major cause of polyomavirus nephropathy (PVN) in renal transplant patients, affecting other SOTs less commonly – ~ 1-9% of renal transplant patients lose renal allograft to PVN ○ Risk factors for BKV infection include donor-related issues (HLA mismatches, deceased-donor organ, high BKV-specific antibodies, and female gender) – Recipient-related risk factors include older age, male gender, low to absent BKV-specific antibody titers – Other factors: IS with lymphocyte-depleting agents, higher IS drug trough levels, acute rejection, antirejection treatment, steroid exposure, type of maintenance IS combinations, and low to absent BKV T-cell responses ○ Main management strategy for controlling BKV infection is to reduce IS and allow BKV-specific cellular immune recovery, in addition to use of antiviral therapy

SUMMARY Relevance and Application of Immune Monitoring in SOT • Concept of immunological monitoring in SOT has gained significant traction in research setting and contributed to literature ○ Builds on premise of individualized medicine, "1 size approach does not fit all" • Substantial gap between research and clinical application remains • Complex immunological manipulation like SOT requires more thorough understanding and routine evaluation of immune responses for improving outcomes ○ Translational (practical) discordance remains • Complexity of immunological analysis in SOT restricts diagnostic testing to clinical laboratories capable of complex and high-throughput analysis involving cellular and molecular techniques • Robust laboratory-based immune monitoring will pave way for personalized IS regimens reducing comorbidities associated with IS • Will improve correlation between drug trough level (pharmacokinetics) and immune function on individual patient basis • Challenge of implementing immune monitoring in practice, obtaining consensus on relevant testing, and selection of appropriate patients remains major obstacle

SELECTED REFERENCES 1. 2. 3. 4.

Millán O et al: Cytokine-based immune monitoring. Clin Biochem. 49(45):338-46, 2016 Shipkova M et al: Editorial: Immune monitoring in solid organ transplantation. Clin Biochem. 49(4-5):317-9, 2016 Schröppel B et al: Gazing into a crystal ball to predict kidney transplant outcome. J Clin Invest. 120(6):1803-6, 2010 Dinavahi R et al: T-cell immune monitoring in organ transplantation. Transplantation. 88(10):1157-8, 2009

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Immunology

Laboratory-Based Immune Monitoring in Organ Transplantation Schematic Representation of CMV-Specific T-Cell Function Using MHC Class I Tetramers

Use of MHC Class I Tetramers for CMVCD8(+) T Cells

CMV-Tetramer-Stimulated CD8(+) T-Cell Degranulation (CD107a/b)

CMV-Tetramer-Stimulated CD8(+) T-Cell IFN-Gamma Production

Histogram of CD45(+) Lymphocyte Proliferation

Histogram of CD43(+) T-Cell Proliferation

(Left) Graphic shows quantitation of antigenspecific CD8 T cells by flow cytometry using MHC class I tetramers. Functional analysis of these T cells can be performed by measuring cytokines and degranulation. (Right) If the CMV CD8 T cell count in blood exceeds 2 cells/uL, functional flow analysis can be performed.

(Left) Flow cytometric assessment shows CD107a/b expression by CMV-specific CD8 T cells after peptide-MHC (tetramer) stimulation (right upper quadrant, Q2). (Right) Flow cytometric assessment shows intracellular IFNgamma production by CMVspecific CD8 T cells after peptide-MHC (tetramer) stimulation (right upper quadrant, Q2-1).

(Left) Quantitation of proliferating total lymphocytes [CD45(+)] is shown by histogram analysis of Edu(+) cells. (Right) Quantitation of proliferating total T cells [CD3(+)] is shown by histogram analysis of Edu(+) cells.

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Laboratory-Based Immune Monitoring in Organ Transplantation

IL-2 Production by CD4(+) T Cells (Left) Polyfunctional CD4 and CD8 T cells produce multiple cytokines at various stages of T-cell activation. The typical cytokines produced and temporally regulated are IFNgamma, IL-2, and TNF-alpha. The peak on the right depicts CD4 T cells producing TNFalpha after T-cell activation with mitogen. (Right) The peak on the right depicts CD4 T cells producing IL-2 after Tcell activation with mitogen.

IFN-Gamma Production by CD4(+) T Cells

Immunology

TNF-Alpha Production by CD4(+) T Cells

TNF-Alpha Production by CD8(+) T Cells (Left) Polyfunctional CD4 and CD8 T cells produce multiple cytokines at various stages of T-cell activation. The typical cytokines produced and temporally regulated are IFNgamma, IL-2, and TNF-alpha. The peak on the right depicts CD4 T cells producing IFNgamma after T-cell activation with mitogen. (Right) The peak on the right depicts CD8 T cells producing TNF-alpha after T-cell activation with mitogen.

IL-2 Production by CD8(+) T Cells

IFN-Gamma Production by CD8(+) T Cells (Left) Polyfunctional CD8 T cells produce multiple cytokines at various stages of T-cell activation. The typical cytokines produced and temporally regulated are IFNgamma, IL-2, and TNF-alpha. The peak on the right depicts CD8 T cells producing IL-2 after T-cell activation with mitogen. (Right) The peak on the right depicts CD8 T cells producing IFN-gamma after Tcell activation with mitogen.

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SECTION 2

HLA Testing

Human Leukocyte Antigen System Histocompatibility Testing Transplantation and HLA ABO Blood Group Antigens and Transplantation Apheresis and Transplantation HLA and Transfusion

32 34 36 40 42 44

HLA Testing

Human Leukocyte Antigen System

TERMINOLOGY

HUMAN LEUKOCYTE ANTIGEN PROPERTIES

Abbreviations

General Characteristics

• Human leukocyte antigen (HLA)

• Most polymorphic human antigens • Codominant expression • Expression on cell surface varies with certain cytokines increasing expression • Demonstrate linkage disequilibrium ○ Certain HLA gene combinations occur more frequently – As compared to that predicted by principles of gene inheritance

INTRODUCTION Human Leukocyte Antigens • Products of genes located in major histocompatibility complex (MHC)

Major Histocompatibility Complex • Located on short arm of chromosome 6 • Spans > 4 million base pair regions

Human Leukocyte Antigen Function • HLAs are key components of immune system • Presents bound peptide fragments on cell surface for recognition by appropriate T lymphocytes

GENETICS Major Histocompatibility Complex

Class I Human Leukocyte Antigen Molecules • Expressed on all nucleated cells with few exceptions • Present peptides to CD8(+) T lymphocytes

Class II Human Leukocyte Antigen Molecules • Expressed on antigen-presenting cells ○ e.g., B lymphocyte, dendritic cells, etc. • Present peptides to CD4(+) T lymphocytes

STRUCTURE

• Consists of > 200 genes ○ Genes grouped into 3 regions: Classes I, II, and III – Classes I and II regions encode genes for HLA – Class I region encodes α chain of HLA class I antigens – Class II region encodes α and β chains of HLA class II antigens

Class I and Class II

Classic Human Leukocyte Antigens

• Consists of α (heavy) chain and 1 β-2 microglobulin (light) chain ○ α chain has 3 distinct domains – α-1 – α-2 – α-3 ○ Peptide-binding groove formed between α-2 and α-3 domains ○ Majority of polymorphisms occur in α-2 and α-3 region ○ α-2 and α-3 regions are encoded by exons 2 and 3 of gene-encoding α chain • β-2-microglobulin (light) chain

• Class I ○ HLA-A, -B, and -C • Class II ○ HLA-DR, -DQ, and -DP

Haplotype • All MHC genes on 1 chromosome are inherited en bloc • Not independent of each other

• Glycoproteins composed of 2 dissimilar protein chains • Belong to immunoglobulin superfamily with similar 3D structure

Class I

Structure of HLA Class I and II Molecules (Left) This illustrations shows the structure of HLA class I and II molecules on the cell surface, which includes a small intracytoplasmic tail, a trans membrane region, and the extracellular structure. (Right) Illustration shows the gene map of the 4 million base pair regions of human major histocompatibility complex region located on the short arm of chromosome 6.

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Gene Map of Human Major Histocompatibility Region

Human Leukocyte Antigen System □ Alleles that differ by silent or noncoding substitutions identified by using 3rd set of digits (3rd field, e.g., HLA-A*02:01:01 and HLAA*02:01:02) □ Alleles that differ by intronic and 5' or 3' untranslated region (UTR) substitutions distinguished by using 4th set of digits (4th field, e.g., HLA-A*02:01:01:01 and HLA-A*02:01:01:03) – Typing for 1st field also called low-resolution typing – Typing for anything more than 1st field considered high-resolution typing □ First 2 field typing currently used for selecting donors for peripheral blood stem cell transplantation □ HLA typing by next generation sequencing allows high-resolution typing up to 4 fields; clinical significance under study

Class II • Consist of α and β chains ○ Polypeptides folded into 2 domains per chain – α chain: α-1 and α-2 – β chain: β-1 and β-2 ○ Peptide-binding groove formed between α-1 and β-1 domains ○ Majority of polymorphisms occur in α-1 and β-1 region ○ α-1 region and β-1 region encoded by exon 2 of geneencoding α and β regions, respectively • Presents peptides derived from exogenous sources • Polymorphisms can derive from both α and β chains depending on class II isoform ○ Both α and β chains are polymorphic for HLA-DQ and -DP ○ HLA-DR on α chain is monomorphic ○ All polymorphisms derived from β chain

NOMENCLATURE Naming System for Human Leukocyte Antigen Genes and Allele Sequences • Responsibility of WHO Nomenclature Committee for Factors of HLA System ○ 1st convened in 1968 ○ Meets every 4 years • 2 major forms of HLA nomenclature ○ Serological – Based on gene product specificities (epitopes) expressed on cell surface – Defined by serological or cellular techniques – a.k.a. antigen level typing – Reported as "HLA" followed by hyphen, capital letters (e.g., A, B, Cw, DR, DQ, DP) designating antigen and number designating specificity (e.g., HLA-A2, HLA-B7) ○ Molecular – Based on identifying polymorphisms in nucleotide sequences – Reported as "HLA" followed by hyphen, capital letters (A, B, C, DRB1, DQB1, DQA1, DPB1, DPA1) designating loci, asterisk, and minimum 2 digits indicating allele group, i.e., antigen (e.g., HLA-A*02) – Each HLA allele name has unique number corresponding to up to 4 sets of digits separated by colons (e.g., HLA-A*02:01:01:01) □ All alleles receive at least 4 digit name, which corresponds to first 2 sets of digits □ Longer names are only assigned when necessary □ Digits before 1st colon (1st field) correspond to serologic antigen (e.g., HLA-A*02 correspond to serologic HLA-A2) □ Next set of digits represent subtypes □ Alleles whose numbers differ in next 2 sets of digits (2nd field) have nucleotide substitutions that result in amino acid changes of protein (e.g., HLA-A*02:01 and HLA-A*02:05)

HLA Testing

○ Noncovalently associated with α chain ○ Encoded by β-2 microglobulin gene on chromosome 15 • Presents peptides to T-cell receptor derived from cellular components that have undergone routine turnover and degradation in cytoplasm

ANTIHUMAN LEUKOCYTE ANTIGEN ANTIBODIES Characteristics • Not naturally occurring • Form as result of alloimmunization ○ Common causes include – Pregnancy – Transfusions – Transplantation • Usually of IgG type • Fix complement • Normally do not develop antibodies to self ○ However, allele-specific antibodies can form e.g., HLAB*44:02 individual develops antibodies to HLA-B*44:03 • Preformed anti-HLA antibodies can result in graft rejection ○ De novo development of anti-HLA antibodies is associated with rejection and accelerated graft loss • Major immune cause of refractoriness to platelet transfusions • Implicated in transfusion-related acute lung injury

SELECTED REFERENCES 1. 2. 3.

4.

5.

6.

7. 8.

Milius RP et al: Genotype list string: a grammar for describing HLA and KIR genotyping results in a text string. Tissue Antigens. 82(2):106-12, 2013 Robinson J et al: The IMGT/HLA database. Nucleic Acids Res. 41(Database issue):D1222-7, 2013 Tait BD: The ever-expanding list of HLA alleles: changing HLA nomenclature and its relevance to clinical transplantation. Transplant Rev (Orlando). 25(1):1-8, 2011 AK Abbas et al: The major histocompatibility complex. In Cellular and Molecular Immunology. 6th ed. Philadelphia: Saunders-Elsevier. 97-1121, 2007 GE Rodney et al: History and nomenclature. HLA beyond tears. In Introduction to Human Histocompatibility. 2nd ed. Durango: De Novo. 1-12, 2000 GE Rodney et al: Structure and function of the HLA complex. HLA beyond tears. In Introduction to Human Histocompatibility. 2nd ed. Durango: De Novo, 2000 Maenaka K et al: MHC superfamily structure and the immune system. Curr Opin Struct Biol. 9(6):745-53, 1999 Natarajan K et al: MHC class I molecules, structure and function. Rev Immunogenet. 1(1):32-46, 1999

33

HLA Testing

Histocompatibility Testing

INTRODUCTION Major Tests Performed in Histocompatibility Laboratory • Human leukocyte antigen (HLA) typing • Anti-HLA antibody (HLA-Ab) screening and identification • Crossmatch testing

HLA TYPING Tests Performed to Identify HLA Class I and II Polymorphisms • Class I includes typing for ○ HLA-A, HLA-B, and HLA-C (using serologic or molecular method) • Class II includes typing for ○ HLA-DR, HLA-DRw, and HLA-DQ (using serologic method) ○ HLA-DRB1, HLA-DRB3/4/5, HLA-DQB1, HLA-DQA1, HLADPB1, and HLA-DPA1 (using molecular methods)

2 Major Methods • Serological ○ Earliest method to define HLA polymorphisms ○ Defines antigens (proteins) expressed on cell surface ○ Performed by complement-dependent cytotoxicity (CDC) method – Lymphocytes and polyclonal sera mixed in multi-well plate (Terasaki tray) for 30 minutes – Rabbit serum (source of complement) added and incubated for additional 60 minutes – Presence of antibody directed to antigen on cell surface leads to complement activation and cell death – Pattern of wells with live and dead cells defines HLA typing • Molecular ○ Defines HLA polymorphisms by genetic analysis ○ Early methods based on Southern blot identification of patterns of genomic DNA digested by restriction enzymes ○ Current methods based on PCR to produce multiple copies of HLA genes ○ Uses extracted DNA from peripheral blood cells, usually lymphocytes ○ Methods identifying polymorphisms in and around region encoding peptide binding groove (exons 2, 3, and 4 for class I and exons 2 and 3 for class II) include – Sequence-specific primer (SSP) method □ Includes multiple PCR reactions □ Primers facilitate amplification only when specific polymorphism present – Sequence-specific oligonucleotide (SSO) 2-step method □ 1st step: Amplification of HLA gene with primers annealing in conserved region of gene □ 2nd step: Amplified product from above identified based on hybridization pattern to multiple DNA probes – Sequence-based typing (SBT) □ Sanger sequencing: Region with majority of polymorphism sequenced 34

○ Methods that identify polymorphisms in entire gene – May include all or most of exons and 5' and 3' untranslated region (UTR) □ Some methods also interrogate intronic variations – Usually done by massive parallel sequencing (a.k.a. next-generation sequencing) □ Commercial kits available □ Bioinformatics support and software to analyse and interpret data needed

ANTI-HLA ANTIBODY TESTING Performed to Detect &/or Identify Anti-HLA-Ab • 2 major methods ○ Serological method ○ Solid phase assay (SpA) • Tests also classified as ○ Nonspecific – Reported as presence or absence of HLA-Ab – May be reported as percent panel reactive antibody (PRA) – Performed by both serological and SpA methods ○ Specific – Identifies both presence and specificity of HLA-Ab □ e.g., HLA-Ab is present, specifically directed to HLAA*02 antigen, i.e., anti-HLA*02 – Usually performed by SpA • Serological method ○ Usually performed by CDC method, similar to typing ○ Differs from typing in that patient serum is screened for HLA-Ab to multiple donors ○ Multi-well plate with lymphocytes from one donor in each well ○ Patient serum added to all wells, followed by addition of complement ○ Number of wells with cytotoxicity reported as percent PRA – e.g., if 34/50 wells demonstrate cytotoxicity, report HLA-Ab present with 68% PRA • SpA method ○ HLA antigen or peptide bound to solid phase plate (enzyme immunoassay) or beads (flow cytometer, Luminex) ○ HLA-Ab in patient serum binds to antigen attached to solid phase support ○ Presence of HLA-Ab detected by secondary anti-human antibody with chemiluminescent or fluorescence tag ○ SpA fluorescence reported in many formats, such as – Mean fluorescence intensity (MFI) – Normalized background ratio (NBG) ○ Fluorescence intensity may correlate with strength &/or avidity of HLA-Ab ○ 3 types of SpA – Screening SpA □ Cell membrane bound on solid phase plate □ Identify HLA-Ab directed to ≥ 1 HLA on cell membrane □ Identify IgG HLA-Ab □ Report HLA-Ab present or absent □ Possible to deduce HLA-Ab specificity – Specific SpA-like single antigen bead (SAB) assay

Histocompatibility Testing

CROSSMATCH TESTING Performed to Identify Presence of Donor-Specific Antibodies • Major methods are cell-based and SpA • Cell based: Recipient serum is reacted with donor cells to identify donor-specific antibodies (DSA) ○ Donor T lymphocytes identify class I HLA-Ab and B lymphocytes identify classes I and II HLA-Ab – 2 major methods: Serologic and flow cytometer based (FXM) – Serologic □ 2 major methods: CDC and AHG-CDC □ Positive result: Donor cell death when reacted with patient serum in presence of complement or CDC method □ CDC crossmatch: Least sensitive method □ AHG-CDC method: Anti-human globulin (AHG) used to enhance cytotoxicity in presence of lower levels of DSA; 10x more sensitive than CDC – FXM □ DSA identified using fluorescent-tagged antihuman secondary antibody □ Increased fluorescence, compared to negative control, is positive test □ FXM 10x more sensitive than AHG-CDC □ Does not involve cytotoxicity • SpA ○ Virtual crossmatch – Utilizes single antigen bead assay to identify specific HLA-Ab and donor HLA type – Positive result: Presence of HLA-Ab directed to donor HLA (identified by donor typing) – Good correlation with actual crossmatch □ Not 100% due to technical issues, non-HLA-Ab, etc.

○ Lysate crossmatch – Donor cells lysed and captured on beads – Beads mixed with recipient serum to identify DSA – Beads can be preserved for longer period – Not widely used due to technical issues

HLA Testing

□ Each solid phase (bead) coated with single HLA antigen or peptide □ Identify all IgG HLA-Ab □ Report: HLA-Ab present with specificity (e.g., antiHLA-A2 present) – SAB assay to specifically detect complement-binding HLA-Ab □ Identify antibodies that bind complement component 1q (C1q) or complement component 3d (C3d) □ Complement-binding donor specific HLA-Ab associated with antibody-mediated rejection and allograft loss □ Considered to be more specific to predict rejection and graft loss □ Identifies high-titer HLA-Ab detected by regular SAB assays that detect total IgG HLA-Ab ○ HLA-Ab specificity utilized to report "calculated" PRA (cPRA) ○ cPRA based on calculated frequency of antigen in given population to which HLA-Ab specificity is present – e.g., of 1,000 Caucasians, HLA-A2 is present in 270 individuals (27%) □ If individual demonstrates only anti-HLA-A2 antibody, cPRA will be 27%

OTHER TESTING Additional Testing • May be performed in some histocompatibility laboratories ○ Immune function monitoring – Assess global immune function in immunocompromised patients – Measures T-cell responses in whole blood □ Commercial assay available that measures T-cell response in presence of immunosuppressive drugs in blood ○ Chimerism studies – Monitor engraftment after allogeneic bone marrow transplantation – Detect relapse after allogeneic bone marrow transplantation – Quantifies amplified short tandem repeat (STR) &/or variable number tandem repeat (VNTR) nucleotide sequences ○ Testing other histocompatibility antigens – MHC class I related chain A (MICA) antigens – H-Y antigen – Killer immunoglobulin receptor (KIR) typing ○ Allograft gene expression profiling – Based on studies that analyzed gene expression and clinical phenotypes in transplant biopsy samples □ Microarray data provides new dimension in biopsy assessment for diagnosis of rejection – Based on studies that analyzed gene expression in peripheral blood and development of acute rejection □ Noninvasive genomic blood test has high correlation with rejection diagnosis at biopsy

SELECTED REFERENCES 1.

2.

3.

4. 5.

6. 7. 8.

Gandhi MJ et al: Targeted next-generation sequencing for human leukocyte antigen typing in a clinical laboratory: Metrics of relevance and considerations for its successful implementation. Arch Pathol Lab Med. 141(6):806-812, 2017 Moreno Gonzales MA et al: Comparison between total IgG, C1q, and C3d single antigen bead assays in detecting class I complement-binding anti-HLA antibodies. Transplant Proc. 49(9):2031-2035, 2017 Reeve J et al: Assessing rejection-related disease in kidney transplant biopsies based on archetypal analysis of molecular phenotypes. JCI Insight. 2(12), 2017 El-Awar N et al: HLA antibody identification with single antigen beads compared to conventional methods. Hum Immunol. 66(9):989-97, 2005 Cao K et al: High and intermediate resolution DNA typing systems for class I HLA-A, B, C genes by hybridization with sequence-specific oligonucleotide probes (SSOP). Rev Immunogenet. 1(2):177-208, 1999 Bray RA et al: The flow cytometric crossmatch. Dual-color analysis of T cell and B cell reactivities. Transplantation. 48(5):834-40, 1989 Patel R et al: Significance of the positive crossmatch test in kidney transplantation. N Engl J Med. 280(14):735-9, 1969 Terasaki PI et al: Microdroplet assay of human serum cytotoxins. Nature. 204:998-1000, 1964

35

HLA Testing

Transplantation and HLA

INTRODUCTION Transplant • Process of taking cells, tissues, or organs (called "graft") from one individual and placing into (usually) another individual

Types of Transplants • Solid organ transplants ○ Includes – Kidney – Liver – Pancreas – Heart – Lung – Small intestine – Composite vascular grafts ○ Usually allografts – Intraspecies but from genetically different individual ○ Sometimes isograft – Graft from genetically similar individual, like syngeneic twin ○ Source of organ – Kidney □ Living or deceased donor – Liver □ Usually deceased donor □ Partial living liver donors and living donors in domino transplant – Other organs □ Deceased donor – Xenograft □ Organ from animals □ Rare experimental attempts • Stem cell transplants ○ Can be allograft or autograft ○ Allograft stem cell sources – Peripheral blood (PB) – Bone marrow (BM) – Umbilical cord (UC)

IMMUNE RESPONSES TO ALLOGRAFTS

36

Rejection • Immune response instigated by recipient's immune system ○ Response to recognition of nonself HLA disparity of graft • Results in both cellular and humoral response • Mechanism ○ Direct pathway – Unique to transplantation – Instigated by passenger antigen presenting cells (APCs) of donor origin that persist in graft post transplantation – Recipient T cell recognizes unprocessed allogeneic MHC molecule on graft APCs – Due to cross reaction of MHC-peptide complex □ Allogeneic MHC-peptide complex create domain that will be recognized by recipient T cells ○ Indirect pathway – Activated by recipient APCs – Recipient T cell recognizes processed peptide of allogeneic MHC molecule bound to self MHC molecule – Similar to host's natural response to infection

Types of Rejection • Classified on basis of histopathologic features or time course of rejection after transplantation • Patterns of rejections based on renal transplantation ○ Hyperacute – Activation of complement and coagulation cascade – Thrombotic occlusion of graft vasculature – Within minutes to hours after anastomosing host and graft blood vessels – Mediated by preexisting donor-specific antibodies that bind to donor endothelial antigen ○ Acute – Graft vascular and parenchymal injury – Mediated by recipient T cells and antibodies – Usually begins 1 week post transplantation ○ Graft vasculopathy and chronic rejection – Slow development of graft arterial occlusion – Secondary to proliferation of vascular intimal smooth muscle cells – Graft parenchyma replaced with fibrous tissue

HLA Antigens

Prevention and Treatment of Rejection

• Expressed on surface of virtually all nucleated cells • Play pivotal role in immune system recognition of self from nonself • Present peptides to T cells • Nonself determinants presented by HLA can result in immune response ○ HLA-peptide interaction with T-cell receptor • High polymorphism ○ Due to evolutionary pressure generated by ability of pathogens to constantly mutate ○ Helps immune system deal with nonself antigens ○ Also major hindrance for transplantation – Allograft is nonself antigen – Results in immune responses that cause graft "rejection"

• Immunosuppression ○ Drugs that inhibit or kill T lymphocytes – Calcineurin inhibitors □ Cyclosporine □ FK-506 – Antiproliferatives □ Azathioprine □ Mycophenolate mofetil – mTOR inhibitor □ Rapamycin – Corticosteroids □ Prednisone □ Hydrocortisone – Antibody based □ Anti-IL-2 receptor (CD25) antibody □ Anti-CD3 antibody

Transplantation and HLA

Graft-vs.-Host Disease • Passenger APCs of donor origin persist, expand and circulate in host post transplantation • Host incapable of mounting effective immunologic reaction against graft • Passenger APCs recognize host tissue as nonself and mount immune response ○ In other words, graft rejects host tissue ○ Can be fatal • Mechanism is unclear ○ HLA mismatching associated with increased risk of graftvs.-host disease (GVHD) in stem cell transplantation • Most common in stem cell transplants ○ Higher incidence in – Mismatched unrelated donors – Haploidentical related donors without posttransplant cyclophosphamide • Amongst solid organ transplants, most common after liver transplantation

Prevention and Treatment of Graft-vs.-Host Disease • GVHD may not respond to treatment • Emphasis on GVHD prophylaxis especially for stem cell transplantation ○ Pharmacologic – Common □ Combination of calcineurin inhibitors cyclosporine or tacrolimus with short course of methotrexate □ ± corticosteroids – Alternate drugs □ Sirolimus □ Mycophenolate mofetil □ Antibiotics ○ T-cell depletion – In case of stem cell transplant only – Associated with increased relapse □ Possibly due to loss of graft vs. leukemia effect • Treatment ○ Pharmacotherapy – Glucocorticoids – Second line □ Cyclosporine □ Tacrolimus □ Antithymocyte globulin □ Mycophenolate mofetil □ Pentostatin ○ Extracorporeal photopheresis

– T lymphocytes isolated from whole blood by pheresis instrument – T lymphocytes treated with psoralens – Exposed to ultraviolet-A light – Treated T lymphocytes reinfused – Mechanism of action unknown □ Possible immunomodulation effect

HLA Testing

□ Anti-CD40 ligand • Reduce graft immunogenicity • Induce donor-specific tolerance • Therapeutic apheresis ○ Therapeutic plasma exchange – To reduce or eliminate preexisting donor-specific antibodies – In conjunction with drugs to eliminate or reduce antibody-producing cells ○ Extracorporeal photopheresis – Role in lung allografts to treat chronic rejection – Evolving literature to treat cardiac allograft vasculopathy

HLA MATCHING Solid Organ Transplant • Depends on organ being considered for transplant • Matching is number of serological antigen specificities at HLA-A, -B, and -DR loci ○ Generally reported as degree of mismatch – e.g., completely matched donor reported as 0/6 mismatch – Mismatched donor reported as 6/6 mismatch • Degree of HLA matching considered suitable varies from each country and also within each center ○ Factors considered – Race – Cold ischemia time – Geography – Recipient and donor age • Kidney transplantation ○ Avoidance of mismatches associated with improved graft survival ○ Strongest impact due to matching at HLA-DR followed by HLA-B and lastly HLA-A ○ Benefit of kidney allocation on basis of HLA matching debated – Current multidrug immunosuppression can minimize effect of poorly matched graft – Matching may result in long-distance shipping of organs with increased cold ischemia time – Potential for longer wait times for patients with less common phenotypes (minorities) – Aggressive immunosuppression can lead to infection and cancer ○ To avoid hyperacute rejection due to presence of donorspecific antibodies (DSA) – Organs allocated to avoid antigens to which recipient has DSA – Such antigens are listed as unacceptable antigens ○ Policies for kidney allocation worldwide usually influenced by geography and patient pool – HLA matched organs shared □ In United States, 0/6 mismatch for highly alloimmunized recipients □ For Eurotransplant program, whenever possible to minimize effect of HLA mismatches • Heart and lung transplantation ○ Influence of HLA for allocation of donors overridden by other factors – Cold ischemia time – ABO matching – CMV matching ○ If 2 potentially similar recipients, matching HLA-DR with recipient preferred 37

HLA Testing

Transplantation and HLA

38

Stem Cell Transplant • Degree of acceptable matching dependent on stem cell source and posttransplant treatment ○ UC vs. PB or BM – Comparable outcomes between UC transplantation with 2 mismatches to fully matched PB or BM transplantation ○ For haploidentical PB or BM transplant, posttransplant, high-dose cyclophosphamide (CPM) outcomes similar to UC transplantation • HLA mismatching associated with ○ Graft failure ○ Increased risk of acute GVHD ○ Decreased overall survival (OS) – ~ 10% cumulative decrease in OS with each mismatch for PB transplantation • Degree of HLA matching ○ For PB transplantation – Matching at allele level or high resolution – Matching at minimum of HLA-A, -B, -C, and -DR loci – Match grade for donor matched at all loci at allele level reported as 8/8 – Usual practice to not use donors with < 7/8 match grade – Additional matching at HLA-DQ and -DP loci also considered ○ For UC transplantation – Matching at HLA-A, -B, and -DR loci □ Allele level for HLA-DR loci □ Antigen (serological) level for HLA-A and -B – Donor fully matched at all 3 loci (serological at HLA-A and -B, and allele level at HLA-DR) reported as 6/6 match – Usual practice to not use cords that are < 4/6 match – Significant interplay between cell dose and HLA matching □ Higher cell dose and higher degree of matching associated with better outcomes – Evolving role for matching at HLA-C □ Matching at HLA-C associated with better outcomes when considering > 5/6 match cord • Alternate protocol ○ Haploidentical transplant – Donor is related and matched at one haplotype with recipient □ In other words, it is half-matched transplant – Alternate source of stem cells to patients who do not have fully matched donor – Early protocol □ T-cell-replete grafts using conventional preparative regimens including low-dose CPM □ Associated with unacceptable GVHD and graft rejection – Later protocol to overcome this □ Ex vivo T-cell depletion of graft often combined with intense preparative regimens □ Associated with decreased GVHD but increased relapse – 3 major current protocols □ 1. T-cell depletion with mega dose CD34(+) cells

□ 2. GCSF-stimulation of donor; intensified posttransplant immunosuppression; antithymoglobulin and combination of PB and BM allografts (GIAC) □ 3. T-cell-replete PB or BM grafts in combination with high dose of posttransplantation cyclophosphamide □ Acceptable GVHD and graft rejection □ Outcomes similar to UC transplant – Advantages of modern haploidentical protocols □ Easy availability of donors (e.g., siblings, parents, children) that are highly motivated □ Rapid availability of adequate stem cell dose □ Lower cost □ Stronger graft vs. leukemia effect

ANTI-HLA ANTIBODIES Solid Organ Transplant • Anti-HLA antibodies directed to donor HLA can be present ○ Pretransplant ○ Develop de novo • Pretransplant DSA associated with ○ Increased incidence of antibody-mediated rejection post transplant ○ May result in increased incidence of graft failure • De novo DSA associated with ○ Antibody-mediated rejection ○ Accelerated graft dysfunction – Lungs: Bronchiolitis obliterans syndrome – Heart: Cardiac allograft vasculopathy – Kidney: Transplant glomerulopathy • Role of DSA in liver transplant debatable ○ Literature regarding liver as protective organ for DSA ○ Emerging literature that high titer DSA associated with graft injury • Detection of anti-HLA antibody dependent on method used • DSA and organ allocation ○ Organs allocated to avoid DSA and to prevent hyperacute rejection ○ Definition of DSA depends on method used – Varies from center to center – Varies by organ □ Low levels of DSA significant for pancreas, kidney transplants □ Liver considered protective to DSA injury and transplants done with high level of DSA ○ Higher alloimmunization may result in higher priority for organ allocation ○ Highly alloimmunized recipients (PRA > 99%) longer waiting period – Alternate options □ Desensitization protocols to decrease DSA at time of transplantation □ Paired donation for kidney transplants

Bone Marrow Transplant • Definition of DSA depends on detection method and center

Transplantation and HLA

Organ

HLA Antigens

Matching Resolution

Solid organs

HLA-A, -B, and -DR

Serological antigen level

Peripheral blood stem cells

HLA-A, -B, -C, and -DR; -DQ and -DP also considered

Allele level

Umbilical cord blood stem cells

HLA-A, -B, and -DR; evolving role for HLA-C

HLA-A and -B antigen level; HLA-DR allele level

HLA Testing

HLA Antigens and Matching Resolution Level Considered for Transplantation

Stem cell transplantation

Methods to Detect Anti-HLA Antibodies Method

Reportable Results

Specific

Cytotoxicity

% PRA

No

ELISA

% PRA

Semi

Flow cytometer

% PRA

Semi

Luminex screening beads

% PRA

Semi

Luminex single antigen

Anti-HLA antibody specificity and calculated % PRA

Yes

Solid Phase

Percent panel reactive antibodies = % PRA.

○ Even when using same method, cutoff for definition varies from one center to another • DSA associated with ○ Delayed engraftment ○ Failure to engraft ○ Usually an issue when – Donor is mismatched □ UC transplant □ Mismatched PB transplantation □ Haplotransplant (half-match transplant) – PB donor matched at HLA-A, -B, -C, -DRB1, and -DQB1 □ ~ 80% of unrelated donors mismatch at HLA-DPB1 □ Can have anti-HLA-DP DSA

SELECTED REFERENCES 1.

2. 3. 4.

5.

6.

7. 8. 9.

Moyer AM et al: Clinical outcomes of HLA-DPB1 mismatches in 10/10 HLAmatched unrelated donor-recipient pairs undergoing allogeneic stem cell transplant. Eur J Haematol. 99(3):275-282, 2017 Kanakry CG et al: Modern approaches to HLA-haploidentical blood or marrow transplantation. Nat Rev Clin Oncol. 13(1):10-24, 2016 Bentall A et al: Five-year outcomes in living donor kidney transplants with a positive crossmatch. Am J Transplant. 13(1):76-85, 2013 Dong M et al: Acute pancreas allograft rejection is associated with increased risk of graft failure in pancreas transplantation. Am J Transplant. 13(4):101925, 2013 Topilsky Y et al: Combined heart and liver transplant attenuates cardiac allograft vasculopathy compared with isolated heart transplantation. Transplantation. 95(6):859-65, 2013 Topilsky Y et al: Donor-specific antibodies to class II antigens are associated with accelerated cardiac allograft vasculopathy: a three-dimensional volumetric intravascular ultrasound study. Transplantation. 95(2):389-96, 2013 Montgomery RA et al: HLA incompatible renal transplantation. Curr Opin Organ Transplant. 17(4):386-92, 2012 Murphey CL et al: Histocompatibility considerations for kidney paired donor exchange programs. Curr Opin Organ Transplant. 17(4):427-32, 2012 Süsal C et al: Impact of HLA matching and HLA antibodies in organ transplantation: a collaborative transplant study view. Methods Mol Biol. 882:267-77, 2012

10. Taner T et al: Prevalence, course and impact of HLA donor-specific antibodies in liver transplantation in the first year. Am J Transplant. 12(6):1504-10, 2012 11. Ciurea SO et al: Donor-specific anti-HLA Abs and graft failure in matched unrelated donor hematopoietic stem cell transplantation. Blood. 118(22):5957-64, 2011 12. Cutler C et al: Donor-specific anti-HLA antibodies predict outcome in double umbilical cord blood transplantation. Blood. 118(25):6691-7, 2011 13. Eapen M et al: Effect of donor-recipient HLA matching at HLA A, B, C, and DRB1 on outcomes after umbilical-cord blood transplantation for leukaemia and myelodysplastic syndrome: a retrospective analysis. Lancet Oncol. 12(13):1214-21, 2011 14. Delaney M et al: The role of HLA in umbilical cord blood transplantation. Best Pract Res Clin Haematol. 23(2):179-87, 2010 15. Spellman S et al: Advances in the selection of HLA-compatible donors: refinements in HLA typing and matching over the first 20 years of the National Marrow Donor Program Registry. Biol Blood Marrow Transplant. 14(9 Suppl):37-44, 2008 16. Hornick P: Direct and indirect allorecognition. Methods Mol Biol. 333:145-56, 2006 17. Sheldon S et al: HLA typing and its influence on organ transplantation. Methods Mol Biol. 333:157-74, 2006

39

HLA Testing

ABO Blood Group Antigens and Transplantation

ABO BLOOD GROUP SYSTEM Introduction • 1900: Landsteiner describes ABO blood group system ○ Groups A, B, and O described ○ Group AB described by Landsteiner's associates in 1902 • > 35 blood group systems described • Importance of ABO system in solid organ transplantation ○ ABO(H) antigens expressed on almost all cells, including endothelial cells ○ Individuals have antibodies against those ABO(H) antigens they lack – Can induce hyperacute rejection of grafts expressing foreign A &/or B antigens

ABH Antigens • Carbohydrate epitopes on different core saccharide chains bound to lipids (glycolipids) or proteins (glycoproteins) • Result from interaction of ABO genes with several other blood group systems, especially H genes action • H gene ○ Now known as FUT1 gene ○ Does not code for H antigen ○ Expression of glycosyltransferase that adds L-fucose to precursor chain to produce H antigen ○ 99.9% of population has ≥ 1 H gene (HH or Hh genotype) ○ Rare individuals without H gene (hh genotype) cannot produce H antigen – Express Bombay phenotype • H antigen ○ Building block for producing A and B antigen ○ Sugars formed in response to inherited A &/or B genes are attached to H antigen to form A &/or B antigen • ABO genes ○ Do not code for ABO antigens ○ Expression of specific glycosyltransferases that add sugars to H antigen ○ A allele – Production of glycosyltransferase that adds N-acetylD-galactosamine (GalNAc) to H antigen – Elicits higher concentration of transferase than B gene ○ B allele – Production of glycosyltransferase that adds Dgalactose (Gal) to H antigen ○ O allele – Considered amorph – No expression of active glycosyltransferase – H antigen is unchanged • A antigen ○ GalNAc sugar attached to H antigen ○ 2 major subgroups: A₁ and A₂ – Have different amounts of A antigen – A₁ antigen □ 810,000-1,170,000 A₁ antigen sites per adult red blood cell (RBC) □ A₁ gene converts almost all H antigens to A antigens – A₂ antigen □ A₂ gene creates only 240,000-290,000 A₂ antigen sites per adult RBC 40

□ Available H antigen sites remain on surface • B antigen ○ Gal sugar attached to H antigen ○ Fewer B antigen than A₁ antigen sites per adult RBC

ABO Antibodies • Considered "naturally occurring" by Landsteiner (1945) • Anti-A & anti-B ABO antibodies (abs) are stimulated by substances present ubiquitously in nature ○ Production initiated at birth ○ Titers generally too low to detect until age 3-6 mos • Serum has antibodies to those antigens that are lacking on individual's RBC ○ Group A individuals have anti-B abs ○ Group B individuals have anti-A abs ○ Group O individuals have anti-A, anti-B, and anti-AB abs • Generally IgM but mixture with IgG, IgA ○ Predominant anti-A abs in group B individual and anti-B abs in group A individual are IgM ○ Group O individual – Predominantly anti-A and anti-B IgM abs – Anti-AB abs can be predominantly IgG or mixture of IgG and IgM • Wide variation in titers of ABO isoagglutinins ○ e.g., anti-A from group O < anti-A from group B < anti-B from group A • Anti-A &/or anti-B titers are required for transplantation across ABO barrier

ABO AND TRANSPLANTATION Expression of ABH Antigen • Kidney ○ Vascular endothelium, distal convoluted tubules, collecting tubules, glomeruli ○ B antigen expression in group B individuals weaker than A antigen expression in group A and AB individuals ○ Group A₂ individuals express small amount of A antigen compared to A₁ individual • Liver ○ Hepatic artery, portal vein, capillary, sinusoidal lining cells, and bile duct epithelium • Heart ○ Endothelium, mesothelial cells on surface of epicardium

Humoral Immune Response: ABO as Barrier to Transplant • Transplanting ABO incompatible organ may result in hyperacute or delayed hyperacute antibody-mediated rejection ○ Due to anti-A, anti-B, or anti-AB abs in serum of recipient to A &/or B antigen that are present on graft – Leads to activation of complement and coagulation cascade – Results in intravascular thrombosis and ischemic necrosis of graft • Can be avoided by removing preformed abs ○ Done by desensitization protocols ○ Measurement of isoagglutinin titers required – No standard method – Variable results within and between institutions

ABO Blood Group Antigens and Transplantation

Blood Type

Antigen

Antibodies

Frequency

A

A

Anti-B

40%

B

B

Anti-A

10%

AB

A and B

None

5%

O

None (H antigen)

Anti-A, anti-B, and anti-AB

45%

ABO Barrier to Transplantation • ABO incompatibility ○ Only group O can provide organs to all recipients, as graft lacks A or B antigen ○ Groups A and B can provide organs to only group A and B recipients, respectively • Shortage of organs • Long wait periods for individuals needing transplant ○ Average wait time for recipients listed for deceased donor kidney in 2015-2016 in USA – Group AB: 2 years – Group A: 3 years – Group O: 5 years – Group B: 6 years • Crossing ABO barrier may increase donor availability, especially for group O and B individuals

Rationale for Crossing ABO Barrier • Availability of better induction therapy and immunosuppression ○ Principles of induction therapy – Reduce preexisting recipient abs to prevent immediate antibody-mediated rejection □ Usually achieved by plasmapheresis or by immunoadsorption □ Typical goal to reduce isoagglutinin titers to 1:8 or less – Reduce continued antibody production by depleting B-cell populations □ Currently achieved by anti-CD20, IVIg, etc. □ Previously achieved by splenectomy • Graft acceptance and long-term survival due to accommodation and tolerance ○ Accommodation – Survival and functioning graft in presence of abs that can bind donor antigens – Postulated to be mediated by increased frequency of T-helper type 2 (Th2) response – ↑ expression of protective genes in graft ○ B-cell-mediated immunological tolerance – Absence of abs to donor AB antigens – Seen in children • Better posttransplantation and acute rejection treatment ○ Availability of immunosuppressive medications – Calcineurin inhibitors – mTOR inhibitors – Monoclonal anti-IL-2Rα receptor abs – Polyclonal anti-T-cell abs ○ Posttransplant monitoring of anti-A and anti-B titers and plasmapheresis

HLA Testing

ABO Blood Groups

Transplantation of A2/A2B Donor Kidneys Into Blood Group B Candidates • Blood group B more common in minority groups compared to Caucasians ○ Group B individuals wait longest to receive kidney ○ Group A2 individuals considered less immunogenic – Low to nonexistent proximal and distal tubule and glomerular staining for A antigens – Lower blood group A expression in vascular endothelium and renal cortex • A2/A2B donor kidneys can be successfully transplanted in carefully selected blood group B candidates ○ Blood group B recipients with low anti-A titer – Many different methods to determine titer – Need to define ideal titer; varies between programs ○ Anti-A titers are monitored pre and post transplant ○ Since December 2014 in USA, OPTN/UNOS allocated of A2/A2B deceased donor kidneys to group B candidates

Current Status • ABO-incompatible transplants occur most often in kidneys ○ Kidneys – Both living donor and deceased donor transplants □ Majority of deceased donor transplants limited to blood group A2/A2B donor to group B recipients – Outcomes similar to ABO-compatible transplants ○ Liver – Recent studies □ Improved transplant techniques and protocols demonstrate similar outcomes (from Japan) □ Being considered in pediatric population ○ Heart – Increasing in pediatric and neonatal population – More common with group A₂ to group O or group A2/A2B to group B ○ Lungs – Not done routinely

SELECTED REFERENCES 1.

2.

3. 4. 5.

Williams WW et al: First report on the OPTN national variance: allocation of A2 /A2 B deceased donor kidneys to blood group B increases minority transplantation. Am J Transplant. 15(12):3134-42, 2015 Irving C et al: Pushing the boundaries: the current status of ABOincompatible cardiac transplantation. J Heart Lung Transplant. 31(8):791-6, 2012 Tanabe M et al: Current progress in ABO-incompatible liver transplantation. Eur J Clin Invest. 40(10):943-9, 2010 Gloor JM et al: ABO incompatible kidney transplantation. Curr Opin Nephrol Hypertens. 16(6):529-34, 2007 Harmening D et al: The ABO blood group system. In Harmening D et al: Modern Blood Banking and Transfusion Practices. 5th ed. Philadelphia: F.A. Davis. 108-33, 2005

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HLA Testing

Apheresis and Transplantation

INTRODUCTION Therapeutic Apheresis • Whole blood circulated through external circuit • Whole blood components (plasma, red blood cells, white blood cells and platelets) separated by ○ Centrifugation ○ Capillary separator • Process ○ May alter composition of blood – Blood component exchanged □ Therapeutic plasma exchange (TPE): Patient plasma replaced with fresh frozen plasma (FFP) or albumin □ Therapeutic red cell exchange: Damaged/infected red blood cells replaced with normal donor red blood cells ○ May alter function of blood – Extracorporeal photopheresis (ECP) □ Buffy coat separated from blood □ Treated extracorporeally with photo active compound (e.g., psoralens) □ Exposed to ultraviolet-A (UVA) light to activate photo active compound □ Treated cells returned to patient in same procedure □ Treated cells have immunomodulatory effect • Technical considerations ○ Adequate venous access – Needles in both arms – Single needle procedures being performed more frequently – May need placement of central venous catheter ○ Blood volume treated – Usually 1-1.5x blood volume processed per session ○ Frequency and number of procedures – Depends on procedure type and clinical condition □ ECP protocols perform procedures 2 consecutive days at a time, over several months □ TPE protocols usually short, with 5-10 procedures on consecutive or every other day • Risks and side effects ○ Usually safe procedure ○ Common side effects – Fatigue – Nausea – Dizziness – Feeling cold – Tingling in fingers and around mouth – Allergic reaction – Lowered blood pressure

INDICATIONS FOR THERAPEUTIC APHERESIS IN TRANSPLANTATION Desensitization: ABO-Incompatible Transplantation • Anti-A and anti-B antibodies can be barrier to transplant ○ ABH blood group antigens expressed on vascular endothelium ○ Anti-A &/or anti-B antibodies directed toward donor ABO antigen may cause hyperacute/acute humoral rejection • Group O patients wait longer for organs 42

○ Demand significantly higher than supply ○ Group O donor can donate organ to any ABO blood group recipient – Group O patient can only receive organs from group O donor • Protocols to reduce pretransplant anti-A and anti-B antibody titers to prevent hyperacute rejection ○ Immunosuppression – Typically consists of □ Tacrolimus □ Mycophenolate □ Steroids □ Induction therapy with IL-2-receptor blocking agent ○ Elimination or reduction of antibody-producing cells – B lymphocytes – Plasma cells ○ Reduction of existing antibodies by TPE or immunoadsorption – Number of procedures needed based on baseline IgG (not IgM) ABO antibody titer □ e.g., total TPE = 1 TPE per ABO antibody tube dilution + 2 □ IgG antibody titers performed in tubes using 2-fold dilutions: 1:2, 1:4, 1:16, etc. □ Thus, when anti-A IgG titer 1:32 = 5 tubes, so total TPE = 5 +2 – Frequency: Every day or every other day – 1-1.5x blood volumes processed ○ Posttransplant TPE or immunoadsorption to promote accommodation – Usually performed for 1st 2 weeks post transplantation □ Majority of acute rejections occur during this period – Frequency: Every day or every other day ○ Therapeutic apheresis (TA) protocols may be supplemented with IVIg ○ Organs transplanted using protocol – Kidney, liver, heart

Desensitization: Donor-Specific Anti-HLA Antibodies • Anti-HLA antibodies (HLA-Ab) can be barrier to transplant ○ HLA antigens expressed on vascular endothelium ○ HLA-Ab formed due to exposure to foreign HLA antigens ○ High titer HLA-Ab directed to HLA antigens on donor endothelium in presence of complement may cause hyperacute rejection • Recipients highly alloimmunized to HLA antigens wait longer to get donor organs ○ Shortage of organs ○ HLA polymorphism – Difficult to find HLA compatible donor • Protocols to reduce pretransplant HLA-Ab titers to prevent hyperacute rejection ○ Similar to ABO desensitization protocol – Immunosuppression followed by – Step 1: Use agents to eliminate (reduce) antibodyproducing cells – Step 2: Use TA modalities to reduce existing HLA-Ab □ No specific immunoadsorption columns available □ Only TPE protocols as described for ABO desensitization

Apheresis and Transplantation

Posttransplant Treatment of Rejection • Posttransplant rejection ○ Hyperacute ○ Acute – Antibody-mediated rejection (AMR) □ Predominantly antibody mediated □ Due to persistent or de novo development of donor-specific antibodies (DSA) – Acute cellular rejection (ACR) □ Predominantly mediated by T lymphocytes ○ Chronic • Role of TA ○ AMR – TPE reduces DSA titer □ 1-1.5x blood volume processed □ Frequency: Every day or every other day □ Duration: Based on clinical improvement – Treated in conjunction with other modalities to eliminate/reduce antibody-producing cells – Transplanted organs where treatment used □ Kidney □ Heart ○ ACR and chronic rejection – ECP □ Early studies treated refractory bronchiolitis obliterans syndrome (BOS) or chronic rejection in lung allografts □ Currently also used to treat ACR and early BOS □ 2 blood volumes processed □ Usually 24 procedures over 6-month period □ Limited data for treatment of cardiac allograft rejection

Transplant-Associated Thrombotic Microangiopathy • Rare • Associated with ○ Calcineurin inhibitors, mTOR inhibitors ○ AMR ○ Atypical hemolytic uremic syndrome ○ Infectious diseases such as HIV, CMV, Parvovirus B19 • Treatment ○ Switching immunosuppression or treating underlying infection ○ TPE used in refractory cases – 1-1.5x blood volume processed – Frequency: Every day; may taper to every other day – FFP used as replacement fluid

HLA Testing

○ Prophylactic – Reduced recurrence rates and improved graft survival – TPE on days 5, 3, and 1 prior to transplant ○ Therapeutic – Up to 80% remission of proteinuria with TPE and ACE inhibitor □ Started within 48 hours of recurrence – TPE protocol □ Post recurrence daily for 3-7 days □ Followed by 3x weekly for 4-12 weeks □ Anti-CD20 antibody may be administered

– Step 3: Posttransplant TPE to prevent acute rejections ○ Protocols supplemented by IVIg ○ Limited role of TPE in protocols using eculizumab (antiC5 antibody) ○ Organs transplanted using protocol – Kidney, heart

ECP and Transplantation • Possible mechanism of action ○ Ex vivo treated lymphocytes after return to patient may undergo apoptosis and modulate in vivo immune responses – Alter T helper sets (decrease effector T cells while expanding T regulatory cells) – Increased dendritic cell differentiation, downregulation of auto-reactive B cells • Indications ○ Solid organ transplantation – Lung transplant □ Stabilization of lung function in patients with persistent acute rejection and early BOS □ Stabilization or improved of lung function in transplant patients with refractory BOS (stages II-III) – Cardiac transplant □ Improve outcome after recalcitrant/severe rejection ○ Treatment of graft-vs.-host disease (GVHD) after peripheral blood stem cell transplantation – Steroid-resistant acute GVHD (grade II-IV) □ 52-100% respond, with highest response rate in skin acute GVHD and least in liver acute GVHD – Steroid-resistant chronic GVHD □ Clinical consensus practice guidelines consider ECP 2nd-line therapy option for steroid refractory chronic GVHD □ 30-65% improve, most with partial responses □ Highest response rates for skin involvement followed by liver and oral mucosa

SELECTED REFERENCES 1.

2. 3. 4.

Schwartz J et al: Guidelines on the Use of Therapeutic Apheresis in Clinical Practice-Evidence-Based Approach from the Writing Committee of the American Society for Apheresis: The Seventh Special Issue. J Clin Apher. 31(3):149-62, 2016 Ward DM: Extracorporeal photopheresis: how, when, and why. J Clin Apher. 26(5):276-85, 2011 Gloor J et al: Sensitized renal transplant recipients: current protocols and future directions. Nat Rev Nephrol. 6(5):297-306, 2010 Gloor JM et al: ABO incompatible kidney transplantation. Curr Opin Nephrol Hypertens. 16(6):529-34, 2007

Recurrent Focal Segmental Glomerulosclerosis • Nephrotic syndrome seen in both adults and children ○ "Permeability factors" postulated as cause • Following renal transplant, 40% have recurrence of focal segmental glomerulosclerosis (FSGS) • Role of TPE 43

HLA Testing

HLA and Transfusion

INTRODUCTION Foreign (Nonself) Antigen Exposure • Antigens expressed on transplanted tissue or transfused cell surface ○ Recognized by recipient immune system – Develops humoral and cellular responses

Human Leukocyte Antigen System • Most immunogenic ○ Exposure to foreign human leukocyte antigen (HLA) results in strong immune response • Most polymorphic ○ Very high likelihood of transfusing HLA-incompatible blood products ○ May result in development of anti-HLA antibodies (HLAAb) • HLA antigens on blood products ○ HLA class I antigens present on following blood products – Platelets (mainly HLA-A and HLA-B antigens) – White cells □ Usually present as contaminant in cellular blood products ○ HLA class II antigens on antigen-presenting cells – B-lymphocytes, monocytes, etc. ○ Red cells not nucleated so no HLA expression – However, Bennett-Goodspeed antigens HLA derived

HUMAN LEUKOCYTE ANTIGEN-RELATED IMMUNOLOGICAL TRANSFUSION REACTIONS Recipient Human Leukocyte Antigen Related • Due to preformed HLA-Ab in recipient ○ HLA-Ab develop due to prior exposure to foreign HLA from 1 of following – Pregnancy – Transfusion – Transplantation ○ 2 major reactions – Febrile nonhemolytic transfusion reactions – Platelet refractoriness

Febrile Nonhemolytic Transfusion Reactions • Most common transfusion reaction • Occur during or 30-60 minutes following transfusion • Common cause is antibodies to 1 of following antigens ○ HLA ○ Human neutrophil antigens (HNA) ○ Human platelet antigens (HPA) • Symptoms ○ Fever, chills ○ Temperature ↑ by 1-2° C • Differential diagnosis ○ Hemolytic transfusion reaction • Treatment ○ Symptomatic treatment for fever • Mitigation ○ Leukoreduction ○ Fresh blood products 44

Platelet Refractoriness • Failure to gain adequate platelet increments 1 hour or up to 24 hours after platelet transfusion ○ Usually estimated by < 5,000/μL platelet increment post transfusion ○ Precisely measured by corrected count increment – Normalizes for body surface area • 2 major causes ○ Nonimmunologic – Nearly 80% of cases – Common factors □ Disseminated intravascular coagulation □ Sepsis □ Splenomegaly □ Medications, such as amphotericin B, ciprofloxacin, certain chemotherapeutic agents, etc. – Management □ Treat underlying cause ○ Immunologic – Less common – Usually antibody mediated – Common factors □ Most common is class I HLA-Ab □ Blood group ABO alloantibodies □ Antibodies to HPA – Diagnosis □ Failure to achieve adequate platelet increment 1 hour post transfusion after ≥ 2 consecutive ABOmatched platelet transfusions – Management approach: Crossmatch-compatible platelets □ Recipient serum crossmatched with donor platelets □ Useful if patient not highly alloimmunized – Management approach: Provide HLA-compatible platelets based on recipient HLA-Ab profile □ Recipient serum screened for presence of HLA-Ab □ Provide platelets from donors that lack antigens to which patient demonstrates HLA-Ab – Management approach: Provide HLA-compatible platelets that are HLA matched with recipient □ Only HLA class I expressed on platelets □ Donor HLA-A and HLA-B considered for matching with recipient HLA type □ Matching can be based on cross-reactive groups: CREG (serologically defined) or eplets (structurally defined) – Usually combination of above approaches utilized – Alternate approaches include □ IVIg □ Plasma exchange (rarely utilized and efficacy debatable) – If above approaches not available and patient is bleeding □ Provide freshest ABO-compatible platelet transfusion

Donor Human Leukocyte Antigen Related • Due to HLA-Ab or viable lymphocytes present in blood product • 2 main reactions

HLA and Transfusion

Transfusion-Related Acute Lung Injury • Rare but life-threatening complication of blood transfusion • Usually due to antibodies in donor blood reacting with recipient antigens ○ Rare cases due to recipient HLA-Ab interacting with transfused cellular products • Symptoms occur within 6 hours of transfusion ○ Fever/chills ○ Hypotension ○ Cyanosis ○ Nonproductive cough, dyspnea, and, sometimes, severe hypoxia • Clinically indistinguishable from adult respiratory distress syndrome • Chest x-ray ○ Severe bilateral pulmonary edema or perihilar and lung infiltration (white out) ○ No cardiac enlargement or involvement of vessels • Differential diagnosis ○ Transfusion-associated circulatory overload ○ Anaphylactic transfusion reaction ○ Bacterial contamination of transfused blood products • Etiology ○ 2-hit hypothesis – 1st hit usually presence of antibodies or soluble mediators of inflammation □ Most commonly implicated are HLA-Ab (classes I and II) in plasma of transfused products □ Also implicated are anti-HNA antibodies, especially anti-HNA3a □ Bioactive lipids that accumulate in stored blood are implicated when antibodies absent – 2nd hit is clinical condition of patient □ Most common factor is presence of lung injury ○ Usually combination of above factors needed ○ Recent published data on animal model – Requirement of recipient T cells to modulate severity of TRALI – Platelets as major contributors of inflammatory process by interplay between platelets and neutrophils ○ Small amount of plasma with implicated substance can cause TRALI ○ Most common with high plasma volume blood products – Fresh frozen plasma (FFP) – Apheresis platelets ○ Not uncommon with low plasma volume blood products – Packed red blood cells • Management ○ Intensive respiratory and circulatory support – Oxygen supplementation in all cases – Mechanical ventilation in some cases ○ Role of steroids unknown ○ Majority of patients improve within 2-3 days • Mitigation ○ Multiple strategies – Production of FFP from male donors only

HLA Testing

– Collecting platelets predominantly from males – Screening blood donors for presence of HLA-Ab &/or anti-HNA antibodies ○ Strategies have resulted in significant decrease in TRALI cases

○ Transfusion-related acute lung injury (TRALI) ○ Transfusion-associated graft-vs.-host disease (TAGVHD)

Transfusion-Associated Graft-vs.-Host Disease • Rare reaction that is almost fatal • Due to transfusion of viable lymphocytes, usually in immunocompromised patient ○ Reported in some nonimmunocompromised patients • Symptoms occur within 8-10 days post transfusion • Etiology is similar to acute graft-vs.-host disease after allogenic stem cell transplantation ○ TAGVHD usually fatal and more severe • Symptoms ○ Fever, diarrhea, abnormal liver function tests ○ Rash, usually on palms • Etiology ○ Presence of viable lymphocytes in transfused product ○ HLA haplotype shared between recipient and donor – Usually donor is homozygous for HLA haplotype shared with recipient ○ Host does not reject immunocompetent donor lymphocytes – Recipient immune system does not recognize donor T lymphocytes as foreign, as they share HLA haplotype ○ Donor T lymphocytes recognize host antigens as foreign and cause TAGVHD • Diagnosis ○ Find evidence of donor T lymphocytes in appropriate clinical setting – Chimerism testing multiple samples at different time points • Management ○ No good therapy ○ Immunosuppression may be beneficial • Mitigation ○ Gamma irradiation of cellular blood products, especially in immunocompromised recipients – Pathogen reduction techniques that intercalate DNA or damage DNA may be used as alternate to gamma irradiation ○ Avoiding directed donations from blood relatives – Especially from siblings and parents

SELECTED REFERENCES 1. 2. 3.

4.

Juskewitch JE et al: How do I … manage the platelet transfusion-refractory patient? Transfusion. 57(12):2828-2835, 2017 Fast LD: Developments in the prevention of transfusion-associated graftversus-host disease. Br J Haematol. 158(5):563-8, 2012 Sayah DM et al: Transfusion reactions: newer concepts on the pathophysiology, incidence, treatment, and prevention of transfusionrelated acute lung injury. Crit Care Clin. 28(3):363-72, v, 2012 Perrotta PL et al: Non-infectious complications of transfusion therapy. Blood Rev. 15(2):69-83, 2001

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SECTION 3

Immunosuppressive Drugs

Mechanism of Action of Immunosuppressive Drugs Therapeutic Drug Monitoring in Transplant Patients History of Immunosuppression Drugs in Transplantation

48 50 54

Immunosuppressive Drugs

Mechanism of Action of Immunosuppressive Drugs

MECHANISMS OF REJECTION Main Components • T lymphocytes are fundamental to transplant rejection ○ Upon antigen presentation, T-cell differentiation results in lymphokine secretion and self proliferation ○ Secreted lymphokines cause following events – Conversion of B cells into antibody-secreting plasma cells – Chemotaxis of macrophages and natural killer cells – Conversion of precytotoxic T lymphocytes (pre-CTLs) into CTLs – All these processes contribute to destruction of targeted tissue ○ To avoid cell-mediated rejection, prevention of T-cell activation is important pharmacological target • Donor-specific antibody (DSA) and complement ○ Alloantibody directed against donor human leukocyte antigen (HLA), blood group antigens, endothelial cell antigens, or other donor-specific antigens with associated complement activation – Causes transplant rejection [antibody-mediated rejection (AMR)] ○ Prevention and treatment of AMR is less developed than for T-cell-mediated rejection

IMMUNOSUPPRESSIVE DRUGS Corticosteroids • Antiinflammatory effect • Actions on T-cell proliferation ○ Induces transcription of inhibitor of NF-kappa B alpha (IκBα) – Binds NF-κB (nuclear factor kappa light chain enhancer of activated B cells) – Prevents its translocation to nucleus ○ Inhibits transcription of IL-1, IL-2, IL-6, interferon-γ, TNF-α

Calcineurin Inhibitors • Cyclosporine (CsA) ○ Cyclic peptide of fungal (Tolypocladium inflatum) origin

– Composed of 11 amino acids ○ Blocks T lymphocyte activity in multifaceted way ○ Forms pharmacologically active complex with intracellular immunophilin cyclophilin (CyP) – Inhibits Ca²(+)/calmodulin-activated serine/threonine phosphatase calcineurin (CN) ○ CsA-CyP-CN complex prevents dephosphorylation of NFATc, preventing initiation of gene transcription for – IL-2 and other cytokine genes (IL-3, IL-4, IL-12) – Inflammatory mediators (TNF-α) – Growth factors [granulocyte colony stimulating factor (G-CSF), macrophage/monocyte colony stimulating factor (M-CSF)] ○ Inhibition of immunocompetent lymphocytes in G0 phase or G1 phase of cell cycle • Tacrolimus (FK506) ○ Macrolide isolated from actinomycete Streptomyces tsukubaensis ○ Similar mechanism of action to CsA ○ Forms pharmacologically active complex with FK506 binding protein-12 (FKBP12) – Complex inhibits Ca2(+)/calmodulin-activated serine/threonine phosphatase CN

Mammalian Target of Rapamycin Inhibitors • Sirolimus (rapamycin) ○ Cyclic macrolide isolated from bacterium Streptomyces hygroscopicus ○ Complexes with FKBP12 ○ Sirolimus: FKBP12 complex inhibits activation of mTOR, a key regulatory kinase – Reduces activity of ribosomal protein S6 kinase beta-1 (S6K1) and eukaryotic elongation factor 4E-binding protein (4EBP1), resulting in decreased protein synthesis ○ Inhibits cytokine-dependent proliferation and differentiation and antibody production ○ Inhibits expression of hypoxia-inducible factor (e.g., HIF1) ○ Reduces expression of vascular endothelial growth factor (VEGF)

Calcineurin Inhibitor Mechanism of Action (Left) Mechanism of action for cyclosporine (CsA) and tacrolimus (FK506), mediated through calcineurin (CN) inhibition, prevents initiation of transcription of IL-2 as well as other cytokines and growth factors. (Right) Mechanism of action for sirolimus (rapamycin) and everolimus, mediated through mammalian target of rapamycin (mTOR) inhibition, ↓ ribosomal protein activity and results in ↓ mRNA translation, protein synthesis.

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mTOR Mechanism of Action

Mechanism of Action of Immunosuppressive Drugs

Antimetabolites • Azathioprine (AZA) ○ Pro-drug metabolized to 6-mercaptopurine (6-MP) ○ 6-MP activation via hypoxanthine-guanine phosphoribosyltransferase (HGPRT) and multienzymatic processes involving kinases – Forms 6-thioguanine nucleotides (6-TGNs) as major metabolites ○ 6-TGN acts as purine antagonist and incorporates into replicating DNA – Inhibits DNA synthesis – Inhibits progression to S phase ○ During breakdown of 6-MP, thioinosinic acid is produced – Blocks conversion of inosine monophosphate (IMP) to adenosine or guanosine monophosphate (AMP/GMP) • Mycophenolate mofetil (MMF) and mycophenolic acid (MPA) ○ MMF (RS-61443) is morpholinoethyl ester of MPA ○ Prodrug rapidly hydrolyzed to biologically active MPA – Potent, selective, noncompetitive, and reversible inhibitor of inosine monophosphate dehydrogenase (IMPDH) – Blocks de novo synthesis of guanosine nucleotides – Lymphocytes rely heavily on de novo purine synthesis pathways □ Antiproliferative effects more potent on lymphocytes than other cell types – Leads to cellular depletion of GMP and guanosine 5'triphosphate (GTP) □ Required for DNA synthesis and necessary for cell growth and replication – Inhibits progression to S phase – Inhibits proliferation of T and B cells and suppresses antibody formation by B cells – Prevents glycosylation of lymphocyte and monocyte glycoproteins involved in intercellular adhesion to endothelial cells – May inhibit recruitment of leukocytes into sites of inflammation and graft rejection

Anti-Interleukin-2 Receptor Antibodies • Monoclonal antibodies against α-chain of interleukin-2 receptor (IL-2Rα) ○ 2 preparations available – Basiliximab (chimeric antibody) – Daclizumab (humanized antibody) • Blocks high-affinity IL-2R on activated T lymphocytes • Inhibits IL-2-induced phosphorylation of Jak1, Jak3, and STAT5A/B components of IL-2R-dependent activation pathway

• Cellular lysis of antibody-coated cells via antibodydependent cell-mediated cytotoxicity &/or their elimination by long-term IL-2 deprivation could occur

Antithymocyte Globulin • Inactivates and depletes lymphocytes by complementdependent lysis or opsonization and phagocytosis

T-Cell Costimulation Blockade • Belatacept ○ Fusion protein composed of Fc fragment of human IgG1 linked to extracellular domain of CTLA-4 – Binds CD80 and CD86 receptors on antigenpresenting cell and prevents them from binding CD28 – Selective blocker of T-cell activation through costimulation blockade

Immunosuppressive Drugs

○ Inhibits kinase activity of cdk4/cyclin D and cdk2/cyclin E complexes that normally peak in middle to late G1 phase ○ Inhibits progression from G1 to S phase of cell cycle • Everolimus ○ 40-O-(2-hydroxyethyl) derivative of sirolimus ○ Similar mechanism of action to sirolimus ○ Complex with FKBP12 – Everolimus: FKBP12 complex forms inhibitory complex formation with mTOR

Experimental Drugs for Transplant Patients • Eculizumab ○ Humanized anti-C5 antibody ○ Inhibits membrane-attack complex formation after complement cascade activation by DSA – Blocks effect of DSA – Does not affect serum DSA levels ○ Salvage therapy for acute AMR ○ Prevents AMR in sensitized kidney transplant patients • Rituximab ○ Humanized anti-CD20 monoclonal antibody – CD20 expressed on B cells, but not plasma cells – Induces apoptosis and prevents B-cell differentiation into DSA-secreting plasma cells – Existing plasma cells not targeted • Intravenous immunoglobulin ○ Decreases serum DSA levels and blocks effect of DSA • Bortezomib ○ Reversible inhibitor of chymotrypsin-like activity of 26S proteasome – Disruption of normal homeostatic mechanisms can lead to cell death ○ Targets plasma cells to decrease production of DSA ○ Salvage therapy in AMR • IgG endopeptidase ○ IgG-degrading enzyme derived from Streptococcus pyogenes (IdeS) ○ Cleaves human IgG into F(ab') and Fc fragments

SELECTED REFERENCES 1. 2. 3.

4.

5. 6.

Jordan SC et al: IgG Endopeptidase in highly sensitized patients undergoing transplantation. N Engl J Med. 377(5):442-453, 2017 Stegall MD et al: The role of complement in antibody-mediated rejection in kidney transplantation. Nat Rev Nephrol. 8(11):670-8, 2012 Snozek C et al: Therapeutic drugs and their management. In Burtis CA et al: Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. 5th ed. St. Louis: Saunders.1057-1108, 2011 Stegall MD et al: Terminal complement inhibition decreases antibodymediated rejection in sensitized renal transplant recipients. Am J Transplant. 11(11):2405-13, 2011 Van Gelder T et al: Anti-interleukin-2 receptor antibodies in transplantation: what is the basis for choice? Drugs. 64(16):1737-41, 2004 Ringe B et al. Immunosuppressive drugs. DOI: 10.1038/ npg.els.0001243. John Wiley & Sons, 2001

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Immunosuppressive Drugs

Therapeutic Drug Monitoring in Transplant Patients

TERMINOLOGY Definitions • Therapeutic drug monitoring (TDM) ○ Monitoring drug concentrations and optimizing dosage – Traditionally based on maintaining steady state concentrations within target ranges ○ Assists patient management through evaluation of – Rejection – Toxicity – Dose adjustments – Compliance • Pharmacodynamic (PD) monitoring ○ Measures activity of target enzymes or altered cellular functioning to evaluate efficacy of drug • Pharmacokinetics (PK) ○ Study of mechanisms of chemicals in body – Absorption – Distribution – Metabolism – Elimination • Pharmacogenomics (PGx) ○ Study of genetic alterations that determine variation in drug response or toxicity

CLINICAL ISSUES Epidemiology • Current drug regimens vary widely according to transplanted organ(s), time since transplant, and other clinical scenarios • Target therapeutic ranges also vary according to transplanted organ(s), time since transplant, and other clinical scenarios ○ Most immunosuppressant drugs have narrow therapeutic indices – Ranges established through experience for specific clinical scenario or population – Do not necessarily ensure adequate immunosuppression nor prevention of toxicity in individual patient ○ Therapeutic ranges generally higher immediately post transplant and lower during maintenance therapy ○ PK are highly variable – Particularly in early posttransplant period ○ Target ranges frequently lower for combination therapies • TDM should not be sole basis for adjusting therapy ○ Careful attention should be paid to – Clinical signs/symptoms – Tissue biopsy findings – Other laboratory and clinical parameters

ETIOLOGY/PATHOGENESIS Analytical Issues • Therapeutic ranges vary with analytical technique ○ Immunoassay – Variable metabolite cross-reactivity of detection antibody in commercially available immunoassays alter therapeutic ranges 50

– Values using this type of analytical technique tend to be higher than chromatographic techniques ○ Liquid chromatography-tandem mass spectrometry (LCMS/MS) – LC-MS/MS more commonly used due to its increased sensitivity and specificity and lack of cross-reactivity of metabolites ○ High-performance liquid chromatography (HPLC) – HPLC methods generally not used due to poor spectral characteristics of some drugs and long analytical times • Trough concentrations generally less effective than interval area under curve (AUC) monitoring ○ Trough samples should be collected immediately prior to dose ○ Impracticality of measuring AUC prevents its general use • Samples collected via venous, arterial, or central access lines should be properly flushed to prevent contamination

PHARMACOLOGIC AGENTS Cyclosporine (CsA) • Pharmacokinetic parameters ○ Adsorption – Bioavailability: 5-60% (mean: 30%) ○ Distribution – Volume of distribution: 3-5 L/kg – 41-58% in erythrocytes (concentration dependent) – Ratio of whole blood concentration to plasma concentration: 4.5 ± 1.5 ○ Metabolism – Extensively metabolized by cytochrome P450 3A (CYP3A4) enzyme system in liver – > 25 metabolites identified – Biological activity of metabolites and their contributions to toxicity considerably less than those of parent compound – Interactions with other drugs also metabolized by CYP3A4 &/or substrates of adenosine triphosphatebinding cassette protein transporters □ e.g., P170 glycoprotein or multidrug resistance protein (MRP)2 ○ Elimination – Half-life: 10-27 hours (mean: 19 hours) – Excretion is biphasic and primarily biliary • Monitoring ○ Sample type = whole blood ○ Trough blood level-adjusted dosage is recommended ○ Drug concentration correlates to immunosuppressive efficacy and toxicity – Poor correlation between dosage and concentration ○ High inter- and intravariability in PK ○ Nephrotoxicity associated with trough plasma levels > 250 ng/mL when measured by radioimmunoassay (RIA) – Nephrotoxicity may be potentiated by other nephrotoxic drugs (e.g., amphotericin B) ○ Hepatotoxicity with levels > 1,000 ng/mL when measured by RIA • Major side effects ○ Nephrotoxicity ○ Hepatotoxicity

Therapeutic Drug Monitoring in Transplant Patients Neurotoxicity Hypertension Hyperlipidemia Hirsutism Tremor Gingival hyperplasia

Tacrolimus (FK506) • Pharmacokinetic parameters ○ Absorption – Bioavailability: 5-65% (mean: 27%) ○ Distribution – Volume of distribution: 5-65 L/kg – Ratio of whole blood to plasma concentration: 35 (1267) ○ Metabolism – Extensively metabolized by CYP3A4 – 31-demethyl metabolite reportedly has same activity as tacrolimus □ Does not contribute significantly to immunosuppressive effect due to its low concentration ○ Elimination – Half-life: 3.5-40.5 hours (mean: 11.3 hours) – Primarily biliary excretion • Monitoring ○ Sample type = whole blood ○ Trough blood level-adjusted dosage recommended ○ Drug concentration correlates with immunosuppressive efficacy and toxicity ○ High inter- and intravariability in PK ○ Interaction with other drugs also metabolized by CYP3A4 &/or substrates of adenosine triphosphatebinding cassette protein transporters (e.g., P170 glycoprotein or MRP2) • Major side effects ○ Nephrotoxicity ○ Neurotoxicity ○ Hypertension ○ Hyperlipidemia ○ Hyperglycemia

Sirolimus (Rapamycin) • Pharmacokinetic parameters ○ Absorption – Bioavailability: 14% – Food decreases absorption ○ Distribution – Volume of distribution: 12 ± 8 L/kg – Distributes primarily in red blood cells; 2-3% in plasma – Ratio of whole blood to plasma concentration: 36 ± 18 ○ Metabolism – Extensively metabolized by CYP3A4 – Pharmacologically active metabolites include □ 41-O- and 7-O-demethyl sirolimus □ Hydroxy sirolimus □ Hydroxy-dimethyl sirolimus ○ Elimination – Half-life: 62 ± 16 hours – Elimination primarily through biliary tract

• Monitoring ○ Sample type = whole blood ○ Trough blood level-adjusted dosage recommended ○ Interaction with other drugs also metabolized by CYP3A &/or substrates of adenosine triphosphate-binding cassette protein transporters • Major side effects ○ Hyperlipidemia ○ Myelosuppression ○ Synergistic effect with cyclosporine in vitro and in vivo

Immunosuppressive Drugs

○ ○ ○ ○ ○ ○

Everolimus • Pharmacokinetic parameters ○ Absorption – Bioavailability: 16% ○ Distribution – Volume of distribution: 107-342 L – Ratio of whole blood to plasma concentration: 17-73 (concentration dependent) ○ Metabolism – Extensively metabolized by CYP3A4 – Metabolites include □ Hydroxy-everolimus □ Dimethyl-everolimus □ Dihydroxy-everolimus □ Ring-opened everolimus – ~ 100x less activity than everolimus ○ Elimination – Half-life: 18-35 hours – 98% of everolimus excreted in bile ○ Major side effects – Hyperlipidemia – Thrombocytopenia – Nephrotoxicity • Monitoring ○ Sample type = blood ○ Trough blood level-adjusted dosage recommended ○ Interaction with other drugs also metabolized by CYP3A &/or substrates of adenosine triphosphate-binding cassette protein transporters

Mycophenolate Mofetil (MMF, RS-61443) • Rapidly absorbed following oral administration • Undergoes rapid and complete metabolism to mycophenolic acid (MPA) ○ Active metabolite • Pharmacokinetic parameters (based on MPA) ○ Absorption – Bioavailability: 94% ○ Distribution – Volume of distribution: 3.6 ± 1.5 L/kg – Ratio of whole blood to plasma concentration: 0.6 – 97% bound to plasma albumin ○ Metabolism – MPA metabolized principally by glucuronyl transferase to form mycophenolic acid glucuronide (MPAG) □ Not pharmacologically active – MPAG converted to MPA via enterohepatic recirculation ○ Elimination 51

Immunosuppressive Drugs

Therapeutic Drug Monitoring in Transplant Patients – Half-life: 17.9 ± 6.5 hours – 93% of MPA excreted in urine • Monitoring ○ Sample type = serum or plasma ○ Pharmacodynamic assay for monitoring of inosine monophosphate dehydrogenase (IMPDH) catalytic activity ○ Utility of monitoring free fraction not clear but may have utility in patients with – Poor kidney function – Hypoalbuminemia – Hyperbilirubinemia • Major side effects ○ Leukopenia ○ Gastrointestinal symptoms

Azathioprine • Prodrug is metabolized to 6-mercaptopurine (6-MP) ○ 6-MP activation occurs via series of multienzymatic processes to form 6-thioguanine nucleotides (6-TGNs) • Pharmacokinetic parameters ○ Absorption – Bioavailability: 88% ○ Distribution – Volume of distribution: 0.808 L/kg – Protein binding: 30% (azathioprine and mercaptopurine) ○ Metabolism – Azathioprine (AZA) rapidly converted to 6-MP by glutathione S-transferase – 6-MP then rapidly enters cells where it is subject to 3 competing metabolic pathways – Activation □ Formation of 6-TGN by sequential actions of hypoxanthine guanine phosphoribosyltransferase (HGPRT), inosine 5'-monophosphate dehydrogenase, and guanidine 5'-monophosphate synthetase – Inactivation □ Oxidation of 6-MP to inactive metabolite 6-thiouric acid by xanthine oxidase (XO) to 6-thiouric acid (6TU) □ Metabolism by thiopurine methyl-transferase (TPMT) to 6-methylmercaptopurine (6-MMP) – Relative activities of XO, HGPRT, and TPMT determine net concentration of active metabolite, 6-TGN ○ Elimination – Half-life: 5 hours (metabolites) • Monitoring ○ AZA blood levels are of little predictive value for therapy – Clinical effects correlate with 6-TGN levels ○ 6-TGN and 6-MMP concentrations are associated with efficacy and may predict toxicity ○ TPMT activity can be associated with genetic polymorphisms – Patients with decreased TPMT activity may be at increased risk of myelotoxicity – TPMT activity correlates inversely with 6-TGN levels in erythrocytes

– TPMT genotyping or phenotyping (red blood cell TPMT activity) can help identify patients at increased risk for developing AZA toxicity • Major side effects ○ Myelosuppression ○ Hepatotoxicity

Monoclonal Antibodies • Most antibodies demonstrate nonlinear, dose-dependent PK • Often exhibit PK/PD properties that are much more complex than those typically associated with smallmolecule drugs • Subject-to-subject variability in PK of these drugs and in development of neutralizing anti-drug antibodies (ADA) • Absorption ○ Absorption following intramuscular or subcutaneous administration is slow and frequently dose dependent ○ Bioavailability is incomplete – Varies from 50-100% due to presystemic extracellular degradation and endocytosis • Distribution ○ Volume of distribution generally close to total blood volume – Cannot be estimated properly due to slow distribution to and elimination from peripheral tissue ○ Distribution to tissue likely to occur through convection and possibly through endothelial endocytosis • Metabolism/elimination ○ Renal or hepatic pathways not involved in metabolism or elimination ○ Elimination occurs via intracellular lysosomal degradation – Following fluid-phase or receptor-mediated endocytosis dose-dependent PK • Monitoring ○ Monitoring of drug serum trough concentrations and of ADA may contribute to improved personalized dosing – Infrequently performed in routine clinical practice because of limited assay availability ○ Antigen-specific biologics have high specificity – Therapeutic range is relatively wide – Correlation of serum drug concentrations or ADA with toxicity is less clear • Basiliximab ○ Volume of distribution: 8.6 ± 4.1 L ○ Half-life: 7.2 ± 3.2 days ○ Complete suppression of interleukin-2 receptor α-chain on peripheral T lymphocytes reported with concentrations of ≥ 0.2 μg/mL • Daclizumab ○ Volume of distribution: 0.074 L/kg ○ Half-life: 11-38 days ○ Saturation of Tac subunit on interleukin-2 receptors reported with concentrations of 5-10 μg/mL

SELECTED REFERENCES 1. 2. 3.

52

PROGRAF (tacrolimus) [package insert]. Northbrook: Astellas Pharmaceuticals, 2012 AFINITOR (everolimus) [package insert]. East Hanover: Novartis Pharmaceuticals, 2011 IMMURAN (azathiopurine) [package insert]. San Diego: Prometheus Laboratories Inc, 2011

Therapeutic Drug Monitoring in Transplant Patients

Transplant Type

Mycophenolic Acid

Cyclosporine

Tacrolimus

Sirolimus

Everolimus

Generic

1.0-3.5 µg/mL

100-400 ng/mL

5-15 ng/mL

4-20 ng/mL

3-8 ng/mL 6-10 ng/mL (monotherapy)

100-200 ng/mL

5-15 ng/mL

10-15 ng/mL

12-20 ng/mL

12-30 ng/mL

Adult Pediatric Kidney

1.5-3.5 µg/mL

150-300 ng/mL, immediately post 8-10 ng/mL < 1 month transplant

Heart Liver

Lung 4.

5.

6. 7.

8.

9. 10. 11.

12. 13. 14. 15. 16. 17.

18.

Immunosuppressive Drugs

Examples of Therapeutic Ranges for Trough Concentrations

9-12 ng/mL 150-250 ng/mL, < 1 year post transplant

4-12 ng/mL, < 60 days post transplant

50-150 ng/mL, > 1 year post transplant

4-10 ng/mL, > 60 days post transplant

175-225 ng/mL

10-15 ng/mL

Snozek C et al: Therapeutic drugs and their management. In Burtis CA et al: Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. 5th ed. St. Louis: Saunders. 1057-1108, 2011 van Rossum HH et al: Pharmacodynamic monitoring of calcineurin inhibition therapy: principles, performance, and perspectives. Ther Drug Monit. 32(1):3-10, 2010 ZORTRESS oral tablets, everolimus oral tablets [package insert]. Stein, Switzerland: Novartis Pharmaceuticals, 2010 CELLCEPT oral capsules, tablets, suspension, IV injection, mycophenolate mofetil oral capsules, tablets, suspension, mycophenolate mofetil HCl IV injection [package insert]. Nutley: Roche Laboratories, 2009 Wallemacq P et al: Opportunities to optimize tacrolimus therapy in solid organ transplantation: report of the European consensus conference. Ther Drug Monit. 31(2):139-52, 2009 Sahasranaman S et al: Clinical pharmacology and pharmacogenetics of thiopurines. Eur J Clin Pharmacol. 64(8):753-67, 2008 Wang W et al: Monoclonal antibody pharmacokinetics and pharmacodynamics. Clin Pharmacol Ther. 84(5):548-58, 2008 SANDIMMUNE oral soft gelatin capsules, oral solution, injection for infusion, cyclosporine oral soft gelatin capsules, oral solution, injection for infusion [package insert]. East Hanover, NJ: Novartis Pharmaceuticals, 2005 SIMULECT (basiliximab) [package insert]. East Hanover, NJ: Novartis Pharmaceuticals, 2005 Kirchner GI et al: Clinical pharmacokinetics of everolimus. Clin Pharmacokinet. 43(2):83-95, 2004 ZENAPAX (daclizumab) [package insert]. Nutley, NJ: Hoffmann-La Roche, 2002 Ringe B et al: Immunosuppressive Drugs. In: eLS. Chichester: John Wiley & Sons Ltd, 2001 RAPAMUNE (sirolimus) [package insert]. Philadelphia, PA: Wyeth-Ayerst Laboratories, 1999 Langman LJ et al: Pharmacodynamic assessment of mycophenolic acidinduced immunosuppression by measuring IMP dehydrogenase activity. Clin Chem. 41(2):295-9, 1995 Venkataramanan R et al: Pharmacokinetics of FK 506 in transplant patients. Transplant Proc. 23(6):2736-40, 1991

53

Immunosuppressive Drugs

History of Immunosuppression Drugs in Transplantation

HISTORICAL OVERVIEW OF IMMUNOSUPPRESSION Beginnings • Immunosuppression has changed significantly over years ○ Greatly increased number of agents available for induction therapy and maintenance immunosuppression • Short-term efficacy of immunosuppressive agents substantially improved ○ As assessed by 1-year graft survival and early acute cellular rejection episodes • Focus of immunosuppression management now shifted to ○ Chronic rejection/long-term graft survival ○ Minimization of overall immunosuppression ○ Investigation of drugs targeting antibody response to graft

Irradiation • Total body and total lymphoid irradiation ○ 1908 – Benjamin et al found that total body irradiation of rabbits impaired synthesis of antibodies toward bovine serum ○ 1914 – Murphy showed that irradiation slowed development of immunity toward tumor allografts ○ Total lymphoid irradiation did improve transplant outcomes – Drawback was tendency toward opportunistic infections and unpredictable, potentially fatal outcomes

Emergence of "Chemical" Immunosuppression • 1914-1916 ○ Murphy and Hekton independently documented immunosuppressive effects of simple compounds, benzene and toluene • 1952 ○ Combination of methyl-bis-(β-chloroethyl)-amine hydrochloride (nitrogen mustard), corticosteroids, and splenectomy prolonged graft survival of mongrel canine allografts • 1959 ○ Schwartz and Dameshek demonstrated that antileukemic drug 6-mercaptopurine (6-MP) prevented rabbits from producing antibodies to many antigenic stimuli

DRUGS IN CLINICAL USE (1962-1994)

54

□ Rates and severity of acute cellular rejection remained high (~ 65%) □ 1-year graft survival: ~ 40% • T-cell-depleting agents ○ Antithymocyte globulin (Thymoglobulin) and muromonab-CD3 (OKT3) – Added broad degree of T-cell inactivation – Conferred heightened clinical efficacy

Cyclosporine Era (1983-1994) • Cyclosporine A ○ Cyclic peptide ○ Possesses immunosuppressive properties ○ Derived from fermentation product of Tolypocladium inflatum fungus ○ Addition of cyclosporine to existing drug regimens decreased acute rejection and improved graft survival – During this era, therapeutic drug monitoring became standard of practice ○ Calcineurin inhibitor ○ Still in clinical use as alternative to tacrolimus

MODERN ERA (1995-PRESENT) Tacrolimus • Carboxy-cyclic macrolide isolated from actinomycete Streptomyces tsukubaensis ○ 1st approved by Food and Drug Administration for use in liver transplantation – Extended to include kidney and other solid organ transplants • Calcineurin inhibitor

Sirolimus • Structurally related to tacrolimus • Produced by Streptomyces hygroscopicus

Everolimus • Structural analog of sirolimus • Also approved treatment for several types of cancer

Mycophenolate Mofetil and Mycophenolic Acid • Mycophenolate mofetil (RS-61443) ○ Morpholinoethyl ester of mycophenolic acid (MPA) • Prodrug rapidly hydrolyzed by cytosolic esterase following absorption to yield active compound MPA • Originally approved for renal transplants, but now, approval includes cardiac and liver transplants

ANTIBODY THERAPIES

Azathioprine Era (1962-1983)

Antibodies Approved for Use in Transplantation

• Corticosteroids ○ Late 1950s to early 1960s: High-dose steroids were combined with irradiation – Associated with significant morbidity ○ Currently used in combinations ± azathioprine and T-celldepleting agents • Azathioprine ○ Used in clinical transplantation – Not very effective in humans until corticosteroids added

• Antithymocyte globulin (Thymoglobulin and Atgam) ○ Polyclonal rabbit (rATG, Thymoglobulin) or equine (eATG, Atgam) antibodies ○ Mechanism of immune response suppression is not fully known • Muromonab-CD3 (Orthoclone OKT3) ○ Mouse monoclonal antibody directed against CD3 resulting in depletion of T cells ○ Used for immunosuppression induction and treatment of rejection – Decreased usage with declining rejection rates

History of Immunosuppression Drugs in Transplantation

Year

Drug

1949

Cortisol

1959

6-mercaptopurine (6-MP)

1959

Cyclophosphamide

1961

Methotrexate

1963

AZA (prodrug to 6-MP)

1975

Mizoribine

1976

Cyclosporine

1977

Sirolimus

1978

Leflunomide

1981

Muromonab-CD3

1983

Antiinterleukin-2 receptor (anti-CD25)

1984

Thymoglobulin

1991

Mycophenolate mofetil/mycophenolic acid

1987

Tacrolimus

1992

Brequinar

1997

Daclizumab

1998

Basiliximab

2003

Everolimus

2011

Belatacept

Immunosuppressive Drugs

Chronology of Discovery of Immunosuppressive Activity

Allison AC: Immunosuppressive drugs: the first 50 years and a glance forward. Immunopharmacology. 47(2-3):63-83, 2000.

• Daclizumab and basiliximab ○ Antiinterleukin-2 receptor (anti-CD25) antibodies ○ Indicated in patients with low to moderate risk of rejection • Belatacept (Nulojix) ○ Selective T-cell costimulation blocker ○ Modified extracellular domain of CTLA-4 fused to portion of Fc domain of IgG1

Antibodies Approved for Other Indications • Alemtuzumab (anti-CD52) ○ Chronic lymphocytic leukemia • Rituximab (anti-CD20) ○ Non-Hodgkin lymphoma ○ Rheumatoid arthritis (RA) • Infliximab [antitumor necrosis factor (TNF)-α] ○ RA ○ Moderate to severe Crohn disease • Adalimumab (anti-TNF) ○ Moderate to severe RA ○ Polyarticular juvenile RA ○ Psoriatic arthritis ○ Ankylosing spondylitis ○ Moderate to severe Crohn disease ○ Moderate to severe plaque psoriasis

3.

Halloran PF: Immunosuppressive drugs for kidney transplantation. N Engl J Med. 2004 Dec 23;351(26):2715-29. Review. Erratum in: N Engl J Med. 352(10):1056, 2005 4. Zand MS: Immunosuppression and immune monitoring after renal transplantation. Semin Dial. 18(6):511-9, 2005 5. Ringe B et al: Immunosuppressive Drugs. In: eLS. Chichester: John Wiley & Sons Ltd, 2001 6. Allison AC: Immunosuppressive drugs: the first 50 years and a glance forward. Immunopharmacology. 47(2-3):63-83, 2000 7. Kahan BD: Timeline of immunosuppression. Transplant Proc. 25(4 Suppl 3):14, 1993 8. Caralps A: History of immunosuppression in kidney transplantation. Transplant Proc. 20(5 Suppl 6):3-4, 1988 9. Levin B et al: Treatment of cadaveric renal transplant recipients with total lymphoid irradiation, antithymocyte globulin, and low-dose prednisone. Lancet. 2(8468):1321-5, 1985 10. Murphy JB: Heteroplastic tissue grafting effected through roentgen-ray lymphoid destruction. JAMA. LXII(19):1459, 1914

SELECTED REFERENCES 1.

2.

Snozek C et al: Therapeutic drugs and their management. In Burtis CA et al: Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. 5th ed. St Louis: WB Saunders. 1057-1108, 2011 Mukherjee S et al: A comprehensive review of immunosuppression used for liver transplantation. J Transplant. 2009:701464, 2009

55

SECTION 4

Kidney Transplantation

History of Kidney Transplantation Pathologic Classification of Renal Allograft Diseases Evaluation of the End-Stage Kidney Evaluation of Allograft Kidney Evaluation of the Donor Kidney Evaluation of Transplant Nephrectomy Evaluation of Fibrosis Protocol Biopsies Accommodation Tolerance

58 60 62 68 74 80 86 90 94 96

Surgical Complications Acute Allograft Ischemia Urine Leak Lymphocele Renal Artery or Vein Thrombosis Transplant Renal Artery Stenosis

100 104 106 108 110

Allograft Rejection Hyperacute Rejection Acute T-Cell-Mediated Rejection Chronic T-Cell-Mediated Rejection Acute Antibody-Mediated Rejection Chronic Antibody-Mediated Rejection

112 116 128 132 140

Recurrent Diseases Recurrent Diseases in the Allograft

152

De Novo Diseases De Novo Focal Segmental Glomerulosclerosis De Novo Membranous Glomerulonephritis Anti-GBM Disease in Alport Syndrome Hyperperfusion Injury Engraftment Syndrome Kidney Diseases in Nonrenal Transplant Recipients Graft-vs.-Host Glomerulopathies

160 162 166 168 170 174 178

Drug Toxicities 180 186

Calcineurin Inhibitor Toxicity mTOR Inhibitor Toxicity

Infections Acute Pyelonephritis Polyomavirus Nephritis Adenovirus, Kidney Cytomegalovirus Infection Histoplasmosis Candidiasis Cryptococcosis Mucormycosis Aspergillosis Coccidioidomycosis Paracoccidioidomycosis Microsporidiosis Tuberculosis Malakoplakia Nocardiosis

188 192 200 204 208 210 212 214 216 218 220 222 224 226 230

Kidney Transplantation

History of Kidney Transplantation

CHRONOLOGY AND EVOLUTION 1st Successful Human Living-Donor Kidney Transplants • 1954 ○ Peter Bent Brigham Hospital, Boston, Massachusetts – Transplant donor and recipient were identical twin brothers, one had end-stage renal disease – No immunosuppression necessary in identical twin donor-recipient pairs – Skin autograft and isograft performed on recipient prior to kidney transplant; no evidence of skin rejection – Kidney transplant functioned for 8 years until recipient died of cardiovascular disease • 1956 ○ Peter Bent Brigham Hospital, Boston, Massachusetts – 1st living-donor kidney transplant performed in woman □ Donor was twin sister – Recipient lived 54 years after kidney transplantation □ Longest surviving kidney transplant recipient; died of causes unrelated to transplant – 1st transplant recipient to become pregnant and give birth

Progress in Immunosuppressive Therapy • Early transplants done in recipients after total body irradiation ○ Irradiation allowed transplantation from nonidentical twin relatives or unrelated donors ○ Some allografts functioned for years, others failed early post transplant • Late 1950s to early 1960s ○ High-dose corticosteroids, total body irradiation, graft irradiation ○ Graft loss due to rejection was common • 1963, Murray et al ○ Prolonged allograft survival with azathioprine (Imuran) immunosuppressive therapy ○ Acute T-cell-mediated rejection (TCMR) still common but improved graft survival • 1963, Starzl et al ○ Combined use of azathioprine and prednisone in human kidney transplant recipients reduced rejection • 1970 to mid 1980s ○ Introduction of T-cell-depleting agents ○ Improved prevention and treatment of acute TCMR rejection ○ Minnesota antilymphocyte globulin – Production shut down in 1992 ○ Antithymocyte globulin (Thymoglobulin, Atgam) ○ OKT3 • 1979, Calne et al ○ Use of cyclosporine A (CSA) monotherapy for immunosuppression in kidney allografts • 1980, Starzl et al ○ Combined CSA and prednisone used in deceased-donor kidney transplant recipients • 1980 to mid 1990s 58

○ Significant improvements in acute rejection rates and long-term graft survival with addition of CSA as maintenance immunosuppression • Mid 1990s to early 2000s ○ Introduction of oral agents tacrolimus (Prograf) and mycophenolate mofetil (CellCept) as part of maintenance immunosuppression ○ Intravenous nondepleting T-cell antibodies as induction therapy: Basiliximab (Simulect), daclizumab (Zenapax) – Directed against IL-2 receptor α chain (CD25), present on cell surface of activated T cells • Current greatest obstacle in immunosuppression is overcoming antibody response in sensitized patients ○ Early acute antibody-mediated rejection (AMR) in positive crossmatch (XM) transplants prevented by inhibition of complement component C5 ○ No effective drugs for chronic AMR

Renal Allograft Pathology • 1991 ○ 1st Banff conference on renal allograft pathology – Consensus classification for acute rejection – Grading scheme for use in multicenter clinical trials • 1997 ○ Banff and Cooperative Clinical Trials in Transplantation classification systems merged ○ "Banff '97" basis for current schema • 1999 ○ Utility of C4d staining for diagnosis of acute AMR and correlation with serum donor-specific antibody (DSA) (Collins et al) • 2001 ○ Addition of AMR criteria to Banff classification (published 2003) ○ Introduction of term "chronic allograft nephropathy" (CAN) • 2005 ○ Removal of term CAN due to lack of specificity (published 2007) ○ Specific disease process (e.g., chronic AMR or BK nephropathy) should be sought as cause of interstitial fibrosis and tubular atrophy (IFTA) – If no specific identifiable disease, IFTA may be used • 2009 ○ Creation of working groups to study other specific disease processes and histologic features in renal allografts • 2013 ○ Recognition of C4d-negative AMR • 2017 ○ Incorporation of IFTA with inflammation ("i-IFTA") as form of chronic TCMR

LABORATORY TESTING Tissue Typing in Early Kidney Transplants • 1965 ○ Most important determinant in long-term graft outcome is degree of histocompatibility between donor and recipient [human leukocyte antigen (HLA) matching] • Testing for donor and recipient HLA types

History of Kidney Transplantation

Serologic Studies for Alloantibodies • 1969, Patel and Terasaki ○ Hyperacute rejection due to preformed antibodies directed against donor antigens causing graft injury ○ Avoidance of transplants with preformed DSA decreases risk of hyperacute rejection • Complement-dependent cytotoxicity (CDC) XM test ○ Early test for DSA ○ Recipient serum incubated with donor lymphocytes in presence of complement – Cell lysis is indication of specific alloantibody activity ○ High rate (80%) of immediate graft failure with positive cytotoxic XM with donor lymphocytes • Flow XM (T and B cell) ○ Donor lymphocytes incubated with recipient sera, exposed to antihuman globulin labeled with fluorochrome, and analyzed by flow cytometry ○ More sensitive than CDC XM test • Solid-phase assays ○ Purified HLA antigens bound to solid surfaces (flow beads or wells of microtiter plates) incubated with recipient serum – Analyzed using flow cytometry or enzyme-linked immunosorbent assay ○ More sensitive than CDC XM test

ALLOANTIBODY TESTING

• Chimerism protocols in human kidney transplantation ○ 2006, HLA-identical sibling donors – Patients with renal failure due to multiple myeloma underwent simultaneous allogeneic kidney and bone marrow transplantation ○ 2008, HLA-mismatched donors and recipients (Kawai et al) – Nonmyeloablative allogeneic bone marrow transplantation and kidney transplantation from same donor □ Induction of mixed chimerism – 4 of 5 patients off immunosuppression with stable functioning grafts at 2-5 years post transplant – Longer term follow-up showed continued functioning grafts in absence of immunosuppression in several patients □ Subset of patients developed DSA and chronic rejection, recurrent glomerular disease

Kidney Transplantation

○ HLA matching less important now with improved T-cell immunosuppressive agents

Xenotransplantation • Currently only performed experimentally • Many immunologic obstacles, including AMR ○ Use of α-1,3-galactosyltransferase gene-knockout pigs as kidney donors prevents hyperacute rejection ○ CRISPR technology should accelerate genetic engineering progress

Ex Vivo Perfusion • Current promising clinical trials in lung, heart, kidney, and liver allografts ○ Recover marginal organs, extend ex vivo time

Relevance to Clinical Kidney Transplantation • +XM and blood group-incompatible kidney transplantation ○ Patients with ABO-incompatible donor or with elevated serum anti-HLA DSA levels considered for pretransplant desensitization – Plasmapheresis, high-dose IVIG ○ Lower level DSA still predisposes patients to early acute AMR &/or chronic AMR • Kidney paired donation (KPD)/paired donor exchange ○ Method of matching willing living donors to compatible recipients ("kidney swap"); 1st proposed by F. Rapaport in 1986 ○ Alternative to +XM or ABO-incompatible transplantation in patients with available living donor ○ Avoidance of unacceptable antigens in presensitized patients ○ Some patients (broadly sensitized) cannot find acceptable donor in KPD programs – +XM transplantation confers greater patient survival than remaining on dialysis awaiting compatible donor

OTHER ENDEAVORS Allograft Tolerance • "Tolerance" refers to immunologic acceptance of functioning graft in setting of immune competence ○ Major mechanisms – Clonal deletion or anergy of alloreactive cells – Regulation or suppression of alloreactive cells • Allogeneic bone marrow transplantation allows for specific acceptance of solid organ allografts from same donor

SELECTED REFERENCES 1.

Haas M et al: The Banff 2017 Kidney Meeting report: revised diagnostic criteria for chronic active T cell-mediated rejection, antibody-mediated rejection, and prospects for integrative endpoints for next-generation clinical trials. Am J Transplant. ePub, 2017 2. Hosgood SA et al: Successful transplantation of human kidneys deemed untransplantable but resuscitated by ex vivo normothermic machine perfusion. Am J Transplant. 16(11):3282-3285, 2016 3. Solez K et al: The Banff classification revisited. Kidney Int. 83(2):201-6, 2013 4. Sharif A et al: Incompatible kidney transplantation: a brief overview of the past, present and future. QJM. 105(12):1141-50, 2012 5. Shimizu A et al: Pathologic characteristics of transplanted kidney xenografts. J Am Soc Nephrol. 23(2):225-35, 2012 6. Kawai T et al: HLA-mismatched renal transplantation without maintenance immunosuppression. N Engl J Med. 358(4):353-61, 2008 7. Zand MS: Immunosuppression and immune monitoring after renal transplantation. Semin Dial. 18(6):511-9, 2005 8. Groth CG et al: Historic landmarks in clinical transplantation: conclusions from the consensus conference at the University of California, Los Angeles. World J Surg. 24(7):834-43, 2000 9. Murray JE: The Nobel lectures in immunology. the Nobel prize for physiology or medicine, 1990. the first successful organ transplants in man. Scand J Immunol. 39(1):1-11, 1994 10. Medawar PB: The Nobel lectures in immunology. the Nobel prize for physiology or medicine, 1960. Immunological tolerance. Scand J Immunol. 33(4):337-44, 1991 11. Patel R et al: Significance of the positive crossmatch test in kidney transplantation. N Engl J Med. 280(14):735-9, 1969 12. Starzl TE et al: Chronic survival after human renal homotransplantation. lymphocyte-antigen matching, pathology and influence of thymectomy. Ann Surg. 162(4):749-87, 1965

59

Kidney Transplantation

Pathologic Classification of Renal Allograft Diseases

TERMINOLOGY Pathogenetic Classification • Based on pathogenesis, divided into broad categories of alloimmune, drug-related, nonalloimmune, and donorderived diseases

Abbreviations • • • • • • •

T-cell-mediated rejection (TCMR) Antibody-mediated rejection (AMR) Peritubular capillaries (PTC) Focal segmental glomerulosclerosis (FSGS) Membranous glomerulonephritis (MGN) Membranoproliferative glomerulonephritis (MPGN) Interstitial fibrosis and tubular atrophy (IFTA), not otherwise specified (NOS) • Thrombotic microangiopathy (TMA) • Mammalian target of rapamycin (mTOR)

DEFINITIONS T-Cell-Mediated Rejection • Target antigens expressed on cell surfaces of endothelial and parenchymal cells • Accessory cells, such as macrophages, participate • Alloimmune reaction of T cells to donor alloantigens, predominantly those of major histocompatibility complex (MHC), in humans termed human leukocyte antigen (HLA) • Non-HLA antigens are also target (e.g., in HLA-identical siblings)

Antibody-Mediated Rejection • Alloantibody-mediated injury to cells expressing donor alloantigens, predominantly HLA class I and II antigens • Accessory mechanisms include complement activation and cells with Fc receptors including macrophages, NK cells, and neutrophils

HLA Molecules • Polymorphic antigens encoded in MHC locus on chromosome 6 ○ Class I molecules (A, B, C) widely expressed on all nucleated cells ○ Class II molecules (DR, DQ, DP), limited expression, B cells, dendritic cells, endothelial cells, macrophages, and activated T cells • Increased expression by interferon-γ

Minor Histocompatibility Antigens • Non-MHC antigens that are polymorphic • Able to trigger allograft rejection in MHC identical recipients

C4d • Fragment of C4 produced by complement activation that binds covalently to nearby molecules • Marker of antibody interaction with endothelium

Multilamination • Multilayered basement membranes • Generally applies to PTC

60

Transplant Glomerulopathy • Glomerular disease seen in transplants defined by duplication of glomerular basement membrane (GBM) • Has several causes, including chronic AMR, TMA, and recurrent MPGN

Transplant Glomerulitis • Mononuclear or polymorphonuclear cells in glomerular capillary loops • Glomerular endothelial swelling and mesangiolysis in more severe cases

Capillaritis • Accumulation of mononuclear or polymorphonuclear cells in PTC

Endarteritis/Endothelialitis • Mononuclear cell infiltration under arterial endothelium

Tubulitis • Mononuclear infiltration in tubules

CATEGORY 1: ALLOIMMUNE RESPONSES T-Cell-Mediated Rejection • Acute TCMR (acute cellular rejection) ○ Tubulointerstitial (Banff type I) ○ Endarteritis/endothelialitis (Banff type II) ○ Arterial transmural inflammation or fibrinoid necrosis (Banff type III) ○ Transplant glomerulitis (no Banff type) • Chronic TCMR ○ Tubulointerstitial (Banff type I + fibrosis and tubular atrophy) ○ Transplant arteriopathy (Banff type II + intimal fibrosis/foam cells)

Antibody-Mediated Rejection • Hyperacute rejection ○ Usually C4d(+) PTC (early samples may be negative) ○ Pathology similar to AHR but occurring < 24 hours post transplant • Acute AMR (acute humoral rejection) ○ Tubular injury (Banff type I) ○ Capillaritis with neutrophils (Banff type II) ○ Arterial fibrinoid necrosis (Banff type III) ○ C4d(+) PTC ○ May be manifested by endarteritis or TMA • Chronic AMR (chronic humoral rejection) ○ Transplant glomerulopathy and glomerulitis ○ PTC multilamination and capillaritis ○ Transplant arteriopathy ○ Usually C4d(+) PTC, may be C4d(-) • Variants ○ Smoldering/indolent (mononuclear capillaritis) ○ C4d(-) (mostly chronic or smoldering) ○ C4d(+) without evidence of active rejection (accommodation)

Pathologic Classification of Renal Allograft Diseases

Calcineurin Inhibitor Toxicity • Cyclosporine and tacrolimus • Chronic (hyaline arteriolopathy, fibrosis, tubular atrophy, FSGS, TMA) • Functional (vasospasm) • Acute (tubulopathy, TMA)

mTOR Inhibitor Toxicity • • • •

Rapamycin (sirolimus) and related drugs Acute tubular injury FSGS TMA

Antiviral Tubular Toxicity • Crystal formation (foscarnet) • Mitochondrial toxicity (adefovir, tenofovir)

Drug-Associated Acute Interstitial Nephritis • Antibiotics and others

CATEGORY 3: INFECTION Viral Infection • • • •

Polyomavirus Cytomegalovirus Adenovirus Herpes simplex virus

Bacterial and Fungal Infection (Partial List) • • • • •

Acute pyelonephritis Chronic pyelonephritis Tuberculosis Malakoplakia Microsporidiosis

CATEGORY 4: ANATOMIC COMPLICATIONS Major Vessel Disease • • • •

Arterial thrombosis Venous thrombosis Arterial dissection Arterial stenosis

Pelvis/Ureter • Urine leak • Obstruction

CATEGORY 5: RECURRENT AND DE NOVO DISEASES Recurrent Primary Disease (Partial List) • • • • • •

FSGS (idiopathic or primary) Atypical hemolytic-uremic syndrome C3 glomerulopathy IgA nephropathy MGN Diabetic nephropathy

• FSGS, adaptive • Diabetic nephropathy • Anti-angiotensin II type 1 receptor autoantibody syndrome

Recipient-Specific De Novo Diseases • Anti-GBM disease in Alport syndrome • Nephrotic syndrome in congenital nephrosis, Finnish type • Antitubular basement membrane disease in tubular basement membrane antigen-deficient recipients

Kidney Transplantation

CATEGORY 2: DRUG TOXICITY AND HYPERSENSITIVITY

Donor-Specific De Novo Diseases • Collapsing FSGS (APOL1 risk alleles in donor)

CATEGORY 6: DONOR DISEASE Present at Time of Transplant (Partial List) • • • • • •

Acute tubular injury (ischemia) Arteriosclerosis TMA Rhabdomyolysis (trauma) Pyelonephritis Glomerular disease ○ Global glomerulosclerosis due to hypertension/aging ○ IgA nephropathy, MGN, and others • Neoplasia ○ Primary (renal cell carcinoma) ○ Metastatic tumor

CATEGORY 7: OTHER DISEASES Neoplasia (Often Viral Related) • Posttransplant lymphoproliferative disease

Idiopathic • IFTA, NOS

NOTES Comments • Rejection may have clinical signs or be subclinical (normal renal function) • Biopsy specimens often show combinations of diseases • Knowledge of original native kidney disease, drug regimen, and time post transplant are essential for diagnostic interpretation

SELECTED REFERENCES 1.

2.

3.

4. 5.

Becker JU et al: Banff borderline changes suspicious for acute t cellmediated rejection: where do we stand? Am J Transplant. 16(9):2654-60, 2016 Haas M: The revised (2013) Banff classification for antibody-mediated rejection of renal allografts: update, difficulties, and future considerations. Am J Transplant. 16(5):1352-7, 2016 Haas M et al: Banff 2013 meeting report: inclusion of C4d-negative antibodymediated rejection and antibody-associated arterial lesions. Am J Transplant. 14(2):272-83, 2014 Mengel M et al: Banff 2011 Meeting report: new concepts in antibodymediated rejection. Am J Transplant. 12(3):563-70, 2012 Williams WW et al: Clinical role of the renal transplant biopsy. Nat Rev Nephrol. 8(2):110-21, 2012

De Novo Disease (Partial List) • MGN (probably alloimmune) 61

Kidney Transplantation

Evaluation of the End-Stage Kidney

TERMINOLOGY Synonyms • End-stage renal disease (ESRD) • End-stage kidney disease (ESKD)

•

Definitions • ESRD defined as renal failure lasting ○ ≥ 3 months ○ Glomerular filtration rate (GFR) ≤ 15 mL/min/1.73 m² ○ Require renal replacement therapy • Pathologic correlates include diffuse glomerular sclerosis, interstitial fibrosis, tubular atrophy, and vascular sclerosis ± cyst formation

•

•

EPIDEMIOLOGY Prevalence • ~ 0.2% in United States with ESRD

Age Range

•

• Median age: 64.2 years • May occur in either children or adults

Ethnicity Relationship • Cumulative lifetime risk of ESRD ○ 7.3% for black men; 2.5% for white men ○ 7.8% for black women; 1.5% for white women

ETIOLOGY/PATHOGENESIS Causes of End-Stage Renal Disease in Adults • Diabetic nephropathy • Hypertensive nephrosclerosis • Glomerular diseases ○ Focal segmental glomerulosclerosis (FSGS) ○ Glomerulonephritis (GN) – IgA nephropathy – Membranoproliferative GN – Crescentic GN

•

□ Pauciimmune [antineutrophil cytoplasmic autoantigen (ANCA)-associated] GN □ Antiglomerular basement membrane GN ○ Membranous nephropathy Hereditary renal diseases ○ Autosomal dominant polycystic kidney disease (ADPKD) ○ Alport nephropathy and others Tubulointerstitial diseases ○ Chronic pyelonephritis ± reflux ○ Obstructive nephropathy ○ Acute tubular necrosis/injury ○ Chronic interstitial nephritis Secondary GN, vasculitis, and thrombotic microangiopathies ○ Lupus nephritis ○ Systemic ANCA vasculitides, including microscopic polyangiitis and granulomatosis with polyangiitis ○ Hemolytic uremic syndrome/thrombotic thrombocytopenic purpura Neoplasia ○ Light chain (myeloma) cast nephropathy ○ Primary renal cell carcinoma (RCC) or urothelial carcinoma Other disorders

Causes of End-Stage Renal Disease in Children Aged < 18 Years • Glomerular diseases ○ GN – Membranoproliferative GN – IgA nephropathy – Crescentic GN, mainly ANCA associated ○ FSGS • Congenital and hereditary diseases ○ Congenital anomalies of kidney and urinary tract – Renal hypoplasia or dysplasia – Congenital obstructive uropathy ○ Cystic diseases – Autosomal recessive polycystic kidney disease (infantile)

Diffuse Granular Cortical Surface (Left) The outer surface of an end-stage kidney due to chronic glomerulonephritis (GN) or hypertensive nephrosclerosis has a diffusely fine granular appearance after removal of the capsule ſt. The depressed areas are due to fibrosis and tubular atrophy. (Right) Ischemic glomerular obsolescence has a shriveled hypocellular capillary tuft ﬈ and collagen deposition in Bowman space ﬊. Bowman capsule is wrinkled ﬉ and sometimes frayed.

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Ischemic Glomerular Obsolescence

Evaluation of the End-Stage Kidney

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• • •

•

Pathogenesis of End-Stage Renal Disease • Initial disease causes progressive damage of nephrons and chronic kidney dysfunction • When GFR decreases to ~ 30% of normal, subsequent renal function declines along final common pathway ○ Independent of original injury ○ Loss of functioning nephrons results in hyperperfusion of remnant glomeruli – Increased individual glomerular filtration (hyperfiltration) ○ Hyperfiltration accelerates podocyte senescence and loss – Leads to segmental glomerular sclerosis and proteinuria – Progressive glomerular sclerosis, tubular atrophy, and interstitial fibrosis • Long-term nonselective proteinuria has toxic effects on tubules ○ Plasma proteins injure epithelium, leading to local inflammation, tubular atrophy, and interstitial fibrosis • Hypertension, hyperfiltration, and proteinuria-induced tubular injury lead to recruitment of pericytes • Pericytes thought to transform to myofibroblasts ○ Myofibroblasts are main source of collagen for interstitial fibrosis

MACROSCOPIC Kidney Size • Very small (< 50 g) kidneys ○ Congenital hypoplasia, renal artery stenosis, and advanced hypertensive nephrosclerosis ○ Few pyramids (≤ 5) in congenital hypoplasia ○ Renal artery stenosis has normal numbers of pyramids • Small (< 125 g) kidneys ○ Surface findings after stripping capsule may be helpful – Discrete and wedge-shaped cortical depressions □ Prior infarction □ Thromboembolism or medium-sized vessel vasculitis (e.g., polyarteritis nodosa)

– Irregular broad depressions at poles typical of chronic pyelonephritis – Diffuse and finely granular subcapsular surface □ Chronic glomerular or tubulointerstitial diseases or hypertensive nephrosclerosis ○ Unilateral small kidney typical of renal artery stenosis • Small or large kidneys with dilated pelvicalyceal system characteristic of hydronephrosis ○ Common causes of obstruction – Ureteral stenosis: Pelviureteric junction or bladder wall – Urolithiasis – Prostatic enlargement: Hyperplasia or carcinoma – Retroperitoneal tumors or fibrosis – Urothelial carcinoma • Large kidneys may or may not have parenchymal cyst formation ○ Large kidneys without cysts suggest diabetic nephropathy or amyloidosis ○ Very large kidneys (500-2,000 g) – Multiple cysts (0.5-3.0 cm) replacing parenchyma are characteristic of ADPKD • Large or small kidneys with irregularly distributed, predominantly cortical cysts seen in acquired cystic disease

Kidney Transplantation

•

– Medullary cystic disease – Nephronophthisis ○ Other hereditary disorders, including podocytopathies and cystinosis Tubulointerstitial diseases ○ Chronic pyelonephritis ± reflux ○ Obstructive nephropathy Secondary GN or vasculitis ○ Lupus nephritis ○ Henoch-Schönlein GN ○ Hemolytic uremic syndrome Hypertensive nephrosclerosis Diabetic nephropathy Neoplasms ○ Primary renal sarcomas ○ Wilms tumor ○ Others Miscellaneous causes ○ Sickle cell nephropathy ○ HIV-associated nephropathy

Pathologic Features of Renal Medulla • Medullary papillae ○ Effaced in hydronephrosis ○ Necrotic, ragged, or calcified in papillary necrosis – Due to diabetic nephropathy ± acute obstructive pyelonephritis □ Also fungal infections □ Sickle cell nephropathy □ Nonsteroidal antiinflammatory agents □ Vasculitis

Secondary Changes in End-Stage Kidney Disease • Secondary changes of acquired cystic disease and hypertensive renal disease common to ESRD from any cause ○ Increased frequency of RCC • Calcium oxalate or phosphate deposition may be nonspecific feature of ESRD ○ Exclude hyperoxaluria and hyperparathyroidism • Gross findings must be correlated with clinical history

MICROSCOPIC General Features • Glomerular sclerosis may have glomerulopathic or ischemic patterns • Tubular atrophy has 4 main morphologic types ○ Classic: Luminal shrinkage with thickening and lamination of tubular basement membrane ○ Thyroidization: Microcystic changes with attenuation of epithelium and inspissation of luminal casts ○ Endocrine: Small, solid tubules with thin basement membranes ○ Super tubules: Enlarged tubules with increased angularity of tubular profiles and epithelial apical snouts

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Kidney Transplantation

Evaluation of the End-Stage Kidney Diabetic Nephropathy • Nodular mesangial sclerosis (Kimmelstiel-Wilson nodules) ○ Thickened capillary basement membranes on PAS or silver stains • Hyaline accumulation in glomerular tufts ("fibrin caps") ○ Between parietal epithelial cells and Bowman capsule ("capsular drops") ○ Hyalinosis of afferent and efferent arterioles

Hypertensive Nephrosclerosis • Arterial and arteriolar sclerosis are relatively nonspecific ○ Can be primary or secondary • Ischemic glomerular obsolescence, interstitial fibrosis, and tubular atrophy ○ Most prominent in outer cortex • Prominent juxtaglomerular apparatuses in hyperreninemic states

Glomerular Diseases • Global glomerular sclerosis ○ End-stage FSGS: Primary or secondary ○ End-stage GN • Nonsclerosed glomeruli may have diagnostic features of original glomerular disease • Immune complex GN may be detectable by immunofluorescence and electron microscopy ○ Useful for lupus nephritis, IgA nephropathy, or other forms of GN ○ Nonspecific coarse granular IgM and C3 ± C1q – In globally sclerotic glomeruli (so-called scar pattern) • Fibrous crescents with synechiae and destruction of Bowman capsules ○ May indicate crescentic GN

○ Calcium phosphate or oxalate in tubules and interstitium • Acquired cystic kidney disease ○ Occurs in patients on peritoneal or hemodialysis or in nondialyzed patients with chronic uremia ○ Frequency ~ 20% at 3 years increasing to ~ 90% at 10 years of dialysis ○ 3-5 cysts per kidney or replacement of 10% of parenchyma are acceptable minimal criteria ○ Cysts may be cortical, medullary, or both – Lined by flattened to low cuboidal epithelium ○ Areas of epithelial crowding or proliferation, taller epithelium, and nuclear atypia – May be associated with development of RCC ○ Intervening parenchyma typically has ESKD changes • RCC ○ Risk of RCC in ESRD markedly increases with duration of dialysis and persists after transplantation ○ RCC in ~ 17% of end-stage kidneys and multifocal in ~ 10% – 41% clear cell, chromophobe, or papillary RCC in largest series – 36% acquired cystic disease-associated RCC □ Acini or solid tubules with high-grade nuclear features □ Abundant calcium oxalate crystals – 23% clear cell papillary RCC ○ 85% CD57(+) – Suggests thin ascending limb of Henle origin ○ Rarely present with metastasis (< 5%)

SELECTED REFERENCES 1.

Tubulointerstitial Diseases • Extensive interstitial fibrosis and tubular atrophy ○ Relative sparing of glomeruli • Medullary effacement and transparenchymal scars with thinning/atrophy of cortex in obstructive nephropathy • Thyroidization of cortical tubules, lymphoid aggregates, and lymphocytic pyelitis in chronic pyelonephritis • Multiple thin-walled cysts lined by flat or cuboidal epithelium in ADPKD

Vascular Diseases • Vessels of all calibers may be affected in thromboembolism ○ Fibrin-platelet thrombi or cholesterol atheroemboli • Wedge-shaped remote cortical infarcts ○ Condensed globally sclerotic glomeruli with interstitial fibrosis and tubular loss

Secondary Changes of End-Stage Kidney Disease • Hypertensive vascular disease ○ Advanced renal scarring complicated by hypertension ○ Arterial and arteriolar sclerosis may be severe and progressive ○ Progressive vascular narrowing enhances renal ischemia • Secondary FSGS ○ Characterized by enlarged glomeruli with perihilar segmental sclerosis ○ Tubules are hypertrophic (super tubules) • Calcium salt deposits 64

2.

3.

4.

5.

Bhatnagar R et al: Renal-cell carcinomas in end-stage kidneys: a clinicopathological study with emphasis on clear-cell papillary renal-cell carcinoma and acquired cystic kidney disease-associated carcinoma. Int J Surg Pathol. 20(1):19-28, 2012 Enoki Y et al: Clinicopathological features and CD57 expression in renal cell carcinoma in acquired cystic disease of the kidneys: with special emphasis on a relation to the duration of haemodialysis, the degree of calcium oxalate deposition, histological type, and possible tumorigenesis. Histopathology. 56(3):384-94, 2010 Tickoo SK et al: Spectrum of epithelial neoplasms in end-stage renal disease: an experience from 66 tumor-bearing kidneys with emphasis on histologic patterns distinct from those in sporadic adult renal neoplasia. Am J Surg Pathol. 30(2):141-53, 2006 Dunnill MS et al: Acquired cystic disease of the kidneys: a hazard of longterm intermittent maintenance haemodialysis. J Clin Pathol. 30(9):868-77, 1977 Schwartz MM et al: Primary renal disease in transplant recipients. Hum Pathol. 7(4):455-9, 1976

Evaluation of the End-Stage Kidney End-Stage Membranoproliferative Glomerulonephritis (Left) End-stage diabetic glomerulosclerosis has solidified glomeruli and broad adhesions to Bowman capsule ﬊. Kimmelstiel-Wilson nodules ﬈ and fibrin caps ﬉ are also apparent. (Right) Chronic membranoproliferative GN shows segmental sclerosis and some capillary double contours ﬈. The glomerulus on the left has active GN with endocapillary hypercellularity and double contours ﬉. Also note marked arteriolar hyalinosis ﬇.

End-Stage Crescentic Glomerulonephritis

Kidney Transplantation

Diabetic Global Glomerular Sclerosis

End-Stage Kidney Disease (Left) Fibrous crescents ﬈ and fragmentation of Bowman capsule are evident in this example of pauciimmune crescentic GN. One glomerulus has extensive sclerosis ﬉. Classic tubular atrophy and interstitial fibrosis are also evident ﬊. (Right) Thyroidization ﬈ with tubular casts, calcium phosphate ﬉ and oxalate st deposition, global glomerular ﬊ sclerosis, and severe arteriosclerosis ſt are evident in this example of end-stage kidney disease (ESKD), which is difficult to classify further.

Tubular Atrophy

Super Tubules (Left) Endocrine-type tubular atrophy ﬈ has small, solid tubules with uniform rounded nuclei and thin basement membranes. "Thyroidization" ﬉ is evident beside these atrophic tubules, and there is severe arteriolosclerosis st. (Right) Super tubules have complex profiles, hypercellularity, voluminous cytoplasm, and apical snouts ﬈. These are features of probable compensatory hypertrophy and hyperplasia. There is extensive interstitial fibrosis and focal mononuclear inflammatory infiltrates ﬉.

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Kidney Transplantation

Evaluation of the End-Stage Kidney

Arteriosclerosis

Arteriolar Hyalinosis

End-Stage Hydronephrosis

Renal Cortical Fibrosis in Hydronephrosis

Autosomal Dominant Polycystic Kidney Disease

Polycystic Kidney Disease

(Left) Severe arteriosclerosis is characterized by marked intimal thickening, myofibroblastic proliferation ﬈, focal fragmentation of the internal elastic lamina ﬉, and medial thickening. (Right) Severe hyaline arteriolosclerosis with intimal hyaline and luminal occlusion ﬈ is evident in this example of ESKD. Hypertension or diabetes may be responsible for these changes.

(Left) A nonfunctioning kidney removed from a 44-year-old woman shows lower ureteral stenosis, obstruction, and marked hydronephrosis. There is marked dilation of the collecting system and parenchymal atrophy. (Right) A kidney with obstructive hydronephrosis shows diffuse interstitial fibrosis and tubular atrophy affecting pars convoluta ﬇ and medullary rays ﬊. Dense outer cortical mononuclear inflammation is also present. Glomeruli are generally spared.

(Left) Autosomal dominant polycystic kidney disease typically has diffuse enlargement (2,310 g in this example), and both the cortex and medulla are entirely replaced by thin-walled unilocular cysts. (Right) Characteristic thin-walled cysts are lined by flat ﬈ or cuboidal ﬉ epithelium in ADPKD. Small papillary proliferations ﬊ may also be evident. The intervening renal parenchyma ﬈ has shrunken or atubular glomeruli with interstitial fibrosis, tubular atrophy, and mononuclear infiltrates ﬊.

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Evaluation of the End-Stage Kidney

Acquired Cystic Kidney Disease (Left) Secondary focal segmental glomerular sclerosis may be evident when there is advanced nephron loss from any primary renal disease. Typically, the glomeruli are enlarged and have perihilar hyalinization ﬈. Adjacent is a globally sclerotic glomerulus ﬈. (Right) A bisected kidney with acquired cystic disease has multiple cysts of varying size ﬈ and a circumscribed mass in the upper pole ﬊. Adipose tissue is prominent in the renal sinus, which is a common finding in ESKD.

Acquired Cystic Kidney Disease

Kidney Transplantation

Secondary (Adaptive) Focal Segmental Glomerulosclerosis

Acquired Cystic Disease in Medulla (Left) Multiple irregularly shaped cysts of variable size and with flattened epithelial lining are present in the cortex of this example of acquired cystic kidney disease. The surrounding tissue has tubular atrophy, interstitial fibrosis, and calcium deposits. (Right) Multiple medullary cysts of variable size with a flat epithelial lining ﬈ are also features of acquired cystic kidney disease. The medullary papilla is effaced ﬊.

Acquired Cystic Disease-Associated Renal Cell Carcinoma

Clear Cell Papillary Renal Cell Carcinoma in End-Stage Kidney Disease (Left) Acquired cystic kidney disease-associated renal cell carcinoma is composed of acini with high-grade nuclei and abundant calcium oxalate crystal deposition ﬈ within the tumor. Basophilic calcium phosphate is also evident. (Right) Clear cell papillary renal cell carcinoma is also observed with high frequency in ESKD. The tumors tend to be multicystic. Papillary fronds lined by clear cells with lowgrade nuclei are characteristic.

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Kidney Transplantation

Evaluation of Allograft Kidney ○ Processed and presented by recipient dendritic cells (indirect pathway)

TERMINOLOGY Abbreviations • • • • •

Antibody-mediated rejection (ABMR) T-cell-mediated rejection (TCMR) Human leukocyte antigen (HLA) Major histocompatibility complex (MHC) Donor-specific antibody (DSA)

Definitions • MHC ○ Locus on chromosome 6 – Encodes class I and class II HLA – Primary target of graft rejection • Minor histocompatibility antigens ○ Non-MHC antigens – Polymorphic and may elicit rejection ○ Mostly unknown structure • Class I MHC antigens ○ Heterodimeric protein ○ Expressed by nucleated cells universally ○ Consist of polymorphic α chain and monomorphic β2microglobulin ○ Recognized by CD8(+) T cells and NK cells ○ Encoded by 3 loci (A, B, C) • Class II MHC antigens ○ Heterodimeric protein expressed constitutively by dendritic cells, B cells, monocytes, and endothelial cells ○ Consist of polymorphic α chain and β chain ○ Recognized by CD4(+) T cells ○ Encoded by 3 loci (DR, DP, DQ) ○ Increased by γ-interferon on many cell types

ETIOLOGY/PATHOGENESIS Afferent Phase • Donor MHC antigens elicit immune response by recipient T cells and B cells ○ On donor cells (direct pathway)

Efferent Phase • Donor-reactive T cells infiltrate graft ○ Direct injury to donor cells (cytotoxicity) ○ Indirect injury (via secreted cytokines and other mediators) ○ Other cells recruited to participate – Macrophages, granulocytes, NK cells ○ Targets include arterial and capillary endothelium, tubular epithelium, and others • Antibodies to donor HLA antigens bind to endothelium ○ Complement fixation – Release of chemotactic and vasoconstrictive mediators – Lysis and loss of endothelial cells – Activation of endothelial cells – Likely critical in early acute ABMR patients with preformed DSA ○ Noncomplement-dependent mechanisms possible – Activation of endothelial cells – Induction of proliferation ○ Cellular participation via Fc receptors – Monocytes, NK cells, granulocytes

Effects Determined by Many Variables • Cellular target (endothelium of arteries, tubular epithelium, glomeruli) • Type of immune response (antibodies vs. T cells) • Intensity of immune response • Resistance of graft to injury • Immunosuppressive drug therapy

Consequences • Ischemia • Loss of nephron integrity • Promotion of fibrosis ○ Includes "i-IFTA": Inflammation in areas of interstitial fibrosis and tubular atrophy • Repair and recovery

Disease Timelines in Renal Transplant (Left) Timeline of major potential diseases in the transplanted kidney begins with donor disease and progresses through rejection (above) and nonrejection categories (below). (Courtesy J. Chapman, MD.) (Right) Diagram represents the 2 major mechanisms of acute renal allograft rejection: Tcell- and antibody-mediated rejection (TCMR and ABMR) [T-cell- and antibody-mediated (humoral) rejection, respectively]. These overlap in a substantial number of cases.

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Mechanisms of Acute Rejection

Evaluation of Allograft Kidney

Clinical Risk Factors • HLA mismatch ○ HLA identical sibling graft half-life: 29 years ○ HLA haploidentical living donor graft half-life: 19 years • Deceased vs. living donor kidneys ○ Living unrelated – Graft half-life: 18 years – 1-year graft survival: 95% ○ Deceased – Graft half-life: 10 years – 1-year graft survival: 89% ○ If deceased donor, expanded criteria donor vs. standard criteria donor • Older vs. younger donor • Presensitization (preexisting DSA) • Delayed graft function • Certain recipient diseases that recur

Current Drug Therapy • Induction ○ Anti-T-cell antibody (antithymocyte globulin, alemtuzumab) ○ Anti-CD25 antibody (basiliximab, daclizumab) • Maintenance ○ Calcineurin inhibitors (cyclosporine, tacrolimus) ○ Corticosteroids (prednisone) ○ Antiproliferative agent (azathioprine, mycophenolate mofetil) ○ Alternatives – Inhibitors of mTOR (rapamycin, everolimus) – Steroid-free regimens – CTLA4-Ig-based therapy (belatacept) • Acute rejection treatment ○ Pulse steroids ○ Anti-T-cell antibody (e.g., antithymocyte globulin) ○ Plasmapheresis for acute ABMR ○ Terminal complement inhibition for acute ABMR ○ Intravenous immunoglobulin (IVIg) for ABMR

BIOPSY PROCESSING Sample • 16-gauge needle yields better sample than 18 gauge ○ 47% of 18-gauge single cores are inadequate (< 7 glomeruli and 1 artery) vs. 24% for 16 gauge ○ No greater complication rate • 2 cores needed for adequacy ○ Single core has ~ 90% sensitivity for acute rejection ○ 2 cores approach 99% sensitivity

Adequacy • Minimal sample of 7 glomeruli, 1 artery (Banff) • Adequate sample of 10 glomeruli, 2 arteries (Banff) • Depends on disease to be identified ○ Some diagnoses can be made in medulla – C4d(+) ABMR – Polyomavirus infection

Routine Pathology Techniques • Light microscopy (LM): Multiple levels (2-3 microns) ○ Stained with H&E, PAS, trichrome, and others ○ IHC for viruses, cell phenotype as needed ○ IHC for C4d on paraffin if no frozen tissue • Immunofluorescence (IF) microscopy on frozen tissue: C4d, full panel if glomerular disease suspected • Electron microscopy (EM), especially if glomerular disease or chronic ABMR suspected

Kidney Transplantation

CLINICAL IMPLICATIONS

EVALUATION OF BIOPSY Careful Examination of All 4 Components • Glomeruli, tubules, interstitium, and vessels • Multiple levels important, since most processes are focal (e.g., endothelialitis)

Assess and Report Extent of Changes • Give number of glomeruli, arteries, tissue cores • Quantitate pathologic features (percentage affected) ○ Cortex – Note interstitial fibrosis, inflammatory infiltrate, peritubular capillaritis ○ Glomeruli – Note global or segmental sclerosis, glomerulitis, glomerular basement membrane (GBM) duplication, mesangial sclerosis, glomerulomegaly, any features of recurrent or de novo glomerular disease ○ Tubules – Note tubular atrophy, tubulitis, viral inclusions ○ Arteries – Note endothelialitis (endarteritis), fibrinoid necrosis, intimal fibrosis, inflammatory cells in thickened intima • Compare with previous biopsy if any ○ Acute TCMR may persist on later biopsies and manifest as i-IFTA (chronic TCMR) ○ Determine cause of i-IFTA pattern – e.g., chronic pyelonephritis ○ Early posttransplant arteriosclerosis may represent donor disease – Can compare to time-zero biopsy if performed • Report diagnostic findings according to current consensus definitions ○ Banff classification widely used • Interpretation of findings needs to be made in conjunction with clinical information

SPECIAL CONSIDERATIONS IN TRANSPLANT BIOPSIES Important Clinical Information • • • • • • • •

Donor source Time post transplant Whether kidney had good initial function Immunosuppressive drug therapy Original disease Renal function Antidonor HLA antibodies (DSA) Proteinuria

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Kidney Transplantation

Evaluation of Allograft Kidney Multiple Diseases May Be Present

Banff Categories (2017)

• Rejection, drug toxicity, viral infection, donor disease

• (1) Normal biopsy or nonspecific changes • (2) Antibody-mediated changes ○ Hyperacute rejection ○ Active AMR (all 3 features must be present for diagnosis) – Histologic evidence of acute tissue injury, including ≥ 1 of following □ Microvascular inflammation (g > 0 or ptc > 0) □ Intimal or transmural arteritis (v > 0) □ Acute TMA, in absence of any other cause □ Acute tubular injury in absence of any other apparent cause – Evidence of current/recent antibody interaction with vascular endothelium, including ≥ 1 of following □ Linear C4d staining in peritubular capillaries (C4d2 or C4d3 by IF on frozen sections or C4d > 0 by IHC on paraffin sections) □ ≥ moderate microvascular inflammation [(g + ptc) ≥ 2)}, unless TCMR present, then g > 1 required; exclude glomerulonephritis as cause of g □ Increased expression of gene transcripts in biopsy tissue indicative of endothelial injury, if validated – Serologic evidence of DSAs (HLA or other antigens) □ C4d staining or expression of validated transcripts/classifiers as noted above may substitute for DSA □ Serum DSA testing still recommended ○ Chronic active AMR (all 3 features must be present for diagnosis) – Morphologic evidence of chronic tissue injury, including ≥ 1 of following □ Transplant glomerulopathy (cg > 0), if no evidence of chronic thrombotic microangiopathy □ Arterial intimal fibrosis of new onset, excluding other causes □ Severe peritubular capillary basement membrane multilayering (PTCBMML) (requires EM): ≥ 7 layers in 1 cortical peritubular capillary and ≥ 5 in 2 additional capillaries – Often manifested by mononuclear cells in PTC (capillaritis) &/or glomeruli (glomerulitis) – Evidence of current/recent antibody interaction with vascular endothelium, including ≥ 1 of following □ Linear C4d staining in peritubular capillaries (C4d2 or C4d3 by IF on frozen sections or C4d > 0 by IHC on paraffin sections) □ At least moderate microvascular inflammation [(g + ptc) ≥ 2]; unless TCMR present, then g > 1 required □ Increased expression of gene transcripts in biopsy tissue indicative of endothelial injury, if thoroughly validated – Serologic evidence of DSA (HLA or other antigens) ○ C4d deposition without evidence of rejection (all features below must be present for diagnosis) – Linear C4d staining in peritubular capillaries (C4d2 or C4d3 by IF on frozen sections or C4d > 0 by IHC on paraffin sections) – Morphologic features of active or chronic active ABMR are not present – No acute or chronic active TCMR or borderline changes

Important to Compare With Previous Biopsies • Progression or resolution of process • Late samples without previous biopsies may be nondiagnostic of cause

NEW APPROACHES Molecular Tests • • • • •

Gene expression in tissue, blood, and urine Proteomics of urine, blood, tissue Systems analysis of pathologic data ("pathomics") May be of clinical use if validated in individual laboratories Indications include biopsies with suspected ABMR ○ Differentiate from pure TCMR, mixed rejection, and no ABMR

Expected Added Value • Assessment of drug effects • Measurement of activity • Assessment of pathways involved in tissue injury

PROTOCOL BIOPSIES (SURVEILLANCE BIOPSIES) Definition • Biopsies taken according to predetermined schedule • Performed independent of graft function

Purpose • Used to monitor status of graft ○ Some centers use surveillance biopsies in routine care • Common in clinical trials to assess efficacy and toxicity • Provides insights into mechanisms and prevalence of graft pathology

Value • Identification of subclinical graft pathology ○ TCMR or ABMR – Subclinical TCMR varies by therapy and sensitization status □ 1-43% at 1-2 months □ 0-15% at 1 year ○ Polyomavirus infection ○ Recurrent or de novo disease ○ Drug toxicity • Follow evolution of chronic progressive diseases

BANFF CLASSIFICATION Background • Classification currently based on LM, IF, or immunohistochemistry and sometimes EM ○ Diagnostic categories defined by semiquantitative scores ○ Opportunity to add other modalities (e.g., gene expression) • Refinement occurs biannually through open meetings to reach consensus on additions/changes based on published evidence • Widely used in drug trials • Specimen adequacy is 10 glomeruli and 2 arteries 70

Evaluation of Allograft Kidney

Caveats • Biopsies may meet criteria for ≥ 2 diagnoses • Detailed criteria established only for rejection categories • Reproducibility of certain categories and features may be limited

BANFF SCORING CATEGORIES Interstitial Inflammation (i) • Mononuclear inflammation in nonfibrotic areas; excludes subcapsular cortex and perivascular infiltrates ○ i0: < 10% of nonfibrotic cortex ○ i1: 10-25%

○ i2: 26-50% ○ i3: > 50% • Do not include fibrotic areas in denominator

Tubulitis (t) • Mononuclear cells in tubules; for longitudinal sections count per 10 tubular epithelial nuclei ○ t0: No mononuclear cells in tubules ○ t1: Foci with 1-4 cells/tubular cross section ○ t2: Foci with 5-10 cells/tubular cross section ○ t3: Foci with > 10 cells/tubular cross section • Need ≥ 2 foci of tubulitis to be present

Kidney Transplantation

– If performed, no molecular evidence for ABMR • (3) Suspicious (borderline) for acute TCMR ○ Foci of tubulitis (t1, t2, or t3) with minor interstitial infiltration (i0 or i1) or interstitial infiltration (i2, i3) with mild (t1) tubulitis – i1 as lower limit of inflammation for diagnosis of borderline rejection is permitted as long as this is clear in pathology report and publications ○ No intimal or transmural arteritis (Banff v0) • (4) TCMR ○ Requires > i1 and ≥ t2 or > v0; C4d(-) for pure TCMR ○ Acute, active TCMR – IA: Interstitial inflammation (> 25% of unscarred cortex) and foci of moderate tubulitis (5-10 mononuclear cells per tubular cross section) – IB: Interstitial inflammation (> 25% of unscarred cortex) and foci of severe tubulitis (> 10 mononuclear cells per tubular cross section) – IIA: Mild to moderate intimal arteritis (< 25% of luminal area) (v1) – IIB: Severe intimal arteritis (> 25% of luminal area) (v2) – III: Transmural arteritis &/or fibrinoid necrosis of medial smooth muscle (v3) ○ Chronic active TMCR – Interstitial inflammation in areas of fibrosis ("i-IFTA") newly recognized in Banff 2017 □ For all i-IFTA categories, other potential causes of iIFTA should be ruled out – Grade IA: ti score 2 or 3 and > 25% of fibrotic cortical parenchyma (i-IFTA score 2 or 3) with moderate tubulitis (t2), not including severely atrophic tubules – Grade IB: Interstitial inflammation involving > 25% of total cortex (ti score 2 or 3) and > 25% of sclerotic cortical parenchyma (i-IFTA score 2 or 3) with severe tubulitis (t3) – Grade 2: Chronic allograft arteriopathy (arterial intimal fibrosis with mononuclear cell inflammation in fibrosis and formation of neointima) □ a.k.a. transplant arteriopathy • (5) Interstitial fibrosis and tubular atrophy, no evidence of any specific etiology ○ Use only when unknown etiology of IF/TA ○ Formerly known as chronic allograft nephropathy ("CAN") • (6) Other ○ Changes considered not due to rejection – Calcineurin inhibitor toxicity, polyomavirus infection, and others

Vascular Inflammation (v) • Mononuclear cells in intima or media of arteries or medial necrosis ○ v0: No arteritis ○ v1: Intimal arteritis in < 25% of lumen (minimum: 1 cell, 1 artery) ○ v2: Intimal arteritis in ≥ 25% of lumen in ≥ 1 artery ○ v3: Transmural arteritis &/or medial smooth muscle necrosis (fibrinoid necrosis) • ‘‘v’’ lesions are only scored in arteries having continuous media with ≥ 2 smooth muscle layers

Glomerulitis (g) • Percentage of glomeruli with increased mononuclear cells in capillaries ○ g0: No glomerulitis ○ g1: < 25% of glomeruli (mostly segmental) ○ g2: 25-75% of glomeruli (segmental to global) ○ g3: > 75% of glomeruli (mostly global)

Interstitial Fibrosis (ci) • Percentage of cortex with fibrosis ○ ci0: ≤ 5% ○ ci1: 6-25% ○ ci2: 26-50% ○ ci3: > 50%

Tubular Atrophy (ct) • Percentage of cortex with atrophic tubules ○ ct0: 0% ○ ct1: ≤ 25% ○ ct2: 26-50% ○ ct3: > 50%

Arterial Fibrointimal Thickening (cv) • Percentage of narrowing of lumen of most severely affected artery ○ cv0: 0% ○ cv1: ≤ 25% ○ cv2: 26-50% ○ cv3: > 50% • Note characteristic lesions of chronic rejection, if present (inflammatory cells in intima, foam cells, lack of fibroelastosis in intima) ○ Chronic allograft arteriopathy only scored in arteries having continuous media with ≥ 2 smooth muscle layers

Transplant Glomerulopathy (cg) • Percentage of glomerular capillary loops with duplication of GBM in most affected nonsclerotic glomerulus by LM 71

Kidney Transplantation

Evaluation of Allograft Kidney ○ cg0: No GBM double contours by LM or EM ○ cg1: ≤ 25% GBM double contours by LM or EM – cg1a: Duplication seen by EM only, in ≥ 3 glomerular capillaries – cg1b: ≥ 1 glomerular capillaries with GBM double contours by LM ○ cg2: 26-50% ○ cg3: > 50%

Mesangial Matrix Increase (mm) • Percentage of glomeruli with mesangial increase, at least moderate increase in ≥ 2 glomerular lobules ○ mm0: 0% ○ mm1: ≤ 25% ○ mm2: 26-50% ○ mm3: > 50%

Arteriolar Hyalinosis (ah) • Circumferential or noncircumferential (focal) hyaline ○ ah0: No arterioles with hyaline ○ ah1: 1 arteriole with noncircumferential hyaline ○ ah2: ≥ 1 arteriole with noncircumferential hyaline ○ ah3: ≥ 1 arteriole with circumferential hyaline • Note if peripheral nodules are present

Peritubular Capillary Inflammation (ptc) • Percentage of cortical PTC with neutrophils or mononuclear cells ○ ptc0: < 10% PTC with > 2 cells/PTC ○ ptc1: > 10% with 3-4 cells/PTC ○ ptc2: > 10% with 5-10 cells/PTC ○ ptc3: > 10% with > 10 cells/PTC • Note whether only mononuclear cells, < 50% neutrophils, or > 50% neutrophils

C4d Score in PTC (C4d) • Percentage of PTC with C4d deposition scored in ≥ 5 HPF ○ C4d0: 0% ○ C4d1: 1-9% ○ C4d2: 10-50% ○ C4d3: > 50% • Note technique used (frozen vs. paraffin)

Total Inflammation (ti) • Includes all cortical inflammation, even subcapsular, perivascular, nodular, and fibrotic areas ○ ti0: < 10% of cortex ○ ti1: 10-25% ○ ti2: 26-50% ○ ti3: > 50%

Inflammation in Areas of Interstitial Fibrosis and Tubular Atrophy (i-IFTA) • Inflammation in scarred cortical parenchyma ○ i-IFTA0: No inflammation or inflammation in < 10% of scarred cortical parenchyma ○ i-IFTA1: Inflammation in 10-25% of scarred cortical parenchyma ○ i-IFTA2: Inflammation in 26-50% of scarred cortical parenchyma ○ i-IFTA3: Inflammation in > 50% of scarred cortical parenchyma 72

Peritubular Capillary Basement Membrane Multilayering • Evaluated by EM • Circumferential basement membrane multilayering in peritubular capillaries ○ Mild: 2-4 layers ○ Moderate: 5-6 layers ○ Severe: ≥ 7 layers • Optimal number of PTCs evaluated by EM to be determined by Banff EM Working Group • Not official Banff score but PTCBMML considered in diagnosis of chronic ABMR ○ Chronic ABMR diagnosis based on severity and number of PTCs involved by PTCBMML

SELECTED REFERENCES 1.

2.

3.

4.

5. 6. 7.

8.

9.

10.

11. 12. 13.

14.

Haas M et al: The Banff 2017 Kidney meeting report: revised diagnostic criteria for chronic active T cell-mediated rejection, antibody-mediated rejection, and prospects for integrative endpoints for next-generation clinical trials. Am J Transplant. ePub, 2017 Becker JU et al: Banff borderline changes suspicious for acute t cellmediated rejection: where do we stand? Am J Transplant. 16(9):2654-60, 2016 Loupy A et al: The Banff 2015 Kidney meeting report: current challenges in rejection classification and prospects for adopting molecular pathology. Am J Transplant. 17(1):28-41, 2016 Haas M et al: Banff 2013 meeting report: inclusion of C4d-negative antibodymediated rejection and antibody-associated arterial lesions. Am J Transplant. 14(2):272-83, 2014 Mengel M et al: Banff 2011 Meeting report: new concepts in antibodymediated rejection. Am J Transplant. 12(3):563-70, 2012 Williams WW et al: Clinical role of the renal transplant biopsy. Nat Rev Nephrol. 8(2):110-21, 2012 Sis B et al: Banff '09 meeting report: antibody mediated graft deterioration and implementation of Banff working groups. Am J Transplant. 10(3):46471, 2010 Solez K et al: Banff '05 meeting report: differential diagnosis of chronic allograft injury and elimination of chronic allograft nephropathy ('CAN'). Am J Transplant. 7(3):518-26, 2007 Furness PN et al: International variation in histologic grading is large, and persistent feedback does not improve reproducibility. Am J Surg Pathol. 27(6):805-10, 2003 Racusen LC et al: Antibody-mediated rejection criteria - an addition to the Banff 97 classification of renal allograft rejection. Am J Transplant. 3(6):70814, 2003 Iványi B et al: Peritubular capillaries in chronic renal allograft rejection: a quantitative ultrastructural study. Hum Pathol. 31(9):1129-38, 2000 Racusen LC et al: The Banff 97 working classification of renal allograft pathology. Kidney Int. 55(2):713-23, 1999 Colvin RB et al: Evaluation of pathologic criteria for acute renal allograft rejection: reproducibility, sensitivity, and clinical correlation. J Am Soc Nephrol. 8(12):1930-41, 1997 Solez K et al: International standardization of criteria for the histologic diagnosis of renal allograft rejection: the Banff working classification of kidney transplant pathology. Kidney Int. 44(2):411-22, 1993

Evaluation of Allograft Kidney

i-IFTA (Left) Interstitial fibrosis and mild tubular atrophy with increased interstitial inflammation is shown. This finding now qualifies for the category of chronic TCMR if it is present in > 25% of the total cortex, in conjunction with moderate to severe tubulitis. (Right) Interstitial fibrosis with inflammation (i-IFTA) is shown. This example shows less dense fibrosis.

i-IFTA With Tubulitis

Kidney Transplantation

i-IFTA

Severe Interstitial Fibrosis and Tubular Atrophy (Left) Mononuclear cell tubulitis ﬈ is seen in partially atrophic tubules, accompanied by interstitial fibrosis with inflammation ﬊, compatible with chronic TCMR. (Right) This example of i-IFTA shows severe interstitial fibrosis and tubular atrophy. Varying densities of interstitial fibrosis and degrees of interstitial inflammation can be seen in transplant biopsies. If accompanied by tubulitis, this example could qualify for a diagnosis of chronic TCMR.

Active ABMR

Transplant Arteriopathy (Left) Moderate peritubular capillaritis ﬈ is shown. In the Banff 2017 schema, this finding in the absence of chronic changes (transplant glomerulopathy or arteriopathy) may be considered “active ABMR" instead of “acute/active ABMR." (Right) Transplant arteriopathy shows narrowing of the arterial lumen and a thickened arterial intima with inflammatory cells ﬈. This pattern may be in chronic ABMR or TCMR. If the latter, this lesion is considered chronic active TCMR grade II by Banff 2017.

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Kidney Transplantation

Evaluation of the Donor Kidney

SURGICAL/CLINICAL CONSIDERATIONS Goal of Consultation • Determine if potential donor kidney is suitable for transplantation • Suitability of kidney depends on following ○ Pathologic features predictive of adequate renal function – ~ 40% of kidneys considered for transplantation under expanded criteria are rejected ○ Donor renal disease – Patients may not have been evaluated for renal disease prior to death – Some donor kidney disease can resolve after transplantation □ IgA nephropathy most common; IgA deposits present in 10% of donor biopsies ○ Donor neoplastic disease – Any suspicious focal lesions are biopsied

Change in Patient Management • Kidney will not be used for transplantation if determined to be unsuitable

○ Donors with Kidney Donor Profile Index (KDPI) of > 85% – Derived from 10 donor factors □ Age □ Height □ Weight □ Ethnicity □ History of diabetes or hypertension □ Cause of death □ Serum creatinine □ Hepatitis C virus □ Donor after cardiac death status ○ By request of surgeon

Kidney Acceptance Criteria • Clinical features of donor and potential recipient and pathologic findings of donor kidney are taken into consideration ○ No absolute cutoff has been established for any pathologic criteria ○ Double transplants possible if concern about renal function

SPECIMEN EVALUATION

Clinical Setting

Gross

• Clinical criteria used to select kidneys most likely to become functional allografts ○ Standard criteria donors (SCDs) – Donors not defined as expanded criteria donors (ECDs) ○ ECDs – All donors age > 60 years – Donors age 50-60 years and have ≥ 2 of following □ Death from cerebrovascular accident □ Hypertension □ Serum creatinine > 1.5 mg/dL – Donation after cardiac death • Organ Procurement and Transplantation Network (OPTN) recommends preimplantation biopsies on subset of potential donors

• Usually small wedge biopsies but can be needle biopsy • Tissue should be gently blotted to remove excess fluid

Frozen Section • Entire specimen used for frozen section • If capsule can be identified, specimen should be oriented so sections are perpendicular to capsule

Reliability • Renal specialist pathologist vs. on-call general pathologist ○ One study found evaluation of donor biopsies by renal pathologist correlated with 1-year graft function and death-censored graft survival – Evaluation by on-call pathologists, however, had no correlation with graft outcome

Donor Biopsy: Frozen Section (Left) This donor kidney biopsy appears edematous and the glomeruli appear hypercellular ﬊. These are common artifacts of frozen sections. The percentage of globally sclerotic glomeruli ﬉ is routinely reported. (Right) Frozen section artifacts are not seen in formalin-fixed permanent sections stained with PAS. This glomerulus has normal cellularity. There is mild intimal hyalinosis of an arteriole ﬈.

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Donor Biopsy: Permanent Section

Evaluation of the Donor Kidney

EVALUATION OF DONOR BIOPSY Sample Adequacy • Size ○ Wedge biopsy 10 mm long x 5 mm wide x 5 mm deep • Structures present ○ ≥ 25 glomeruli should be present, including glomeruli from deep cortex ○ ≥ 2 arteries should be present

Histologic Features • Chronic changes ○ Present in many biopsies ○ Usually increase with donor age • Glomerulosclerosis ○ Glomerulus replaced by solid eosinophilic fibrosis ○ Percentage of globally sclerotic glomeruli important (global glomerulosclerosis) – > 20% associated with higher incidence of delayed graft function (DGF) requiring transient dialysis and higher creatinine (Cr) at 3-24 months □ Variable effect on graft survival ○ No absolute cutoff point for percent of global glomerulosclerosis has been determined ○ Sclerotic glomeruli predominately in subcapsular cortex in arteriosclerosis and often overestimated in wedge biopsies ○ Strong correlation with age • Arteriosclerosis ○ Moderate arteriosclerosis (> 25% luminal narrowing) predictor of worse graft outcome (graft loss, DGF, higher Cr) • Interstitial fibrosis and tubular atrophy ○ Not consistently predictive of graft function • Thrombi in glomeruli and arteries ○ Head trauma in donor can precipitate thrombotic microangiopathy ○ Even with glomerular thrombi, good outcome possible ○ Reporting percentage of glomeruli with thrombi may not represent extent of thrombi in graft

– Percentage of glomerular area involved by thrombi (segmental vs. diffuse) may convey better extent of thrombosis □ Not well studied ○ Cholesterol emboli may be contraindication to transplantation • Other features ○ Renal infarct: May be associated with vascular changes ○ Tumors – Patients may have unsuspected neoplasms – Angiomyolipoma most common benign renal tumor (0.1-0.2% of population) □ Involved kidneys used successfully for transplantation □ May be contraindication to transplantation in some settings – Small, well-differentiated renal cell carcinomas may be resected and kidney used for transplantation ○ Mesangial nodules – Often associated with diabetes ○ Pigmented casts in tubules – Myoglobin casts associated with rhabdomyolysis □ May not be contradiction to transplantation ○ Interstitial inflammation – Lymphocytic infiltrates are common finding □ Very rarely due to lymphoma or leukemia – Granulomatous inflammation may be contraindication for transplantation

Kidney Transplantation

○ Training of on-call pathologists recommended if renal pathologist not available ○ If no renal pathologist available on site, whole slide imaging with remote interpretation by renal pathologist may be option in some centers • Reproducibility ○ Good to fair reproducibility among pathologists for some features – Number of viable and globally sclerotic glomeruli – Percentage of global glomerulosclerosis – Interstitial fibrosis – Arteriosclerosis ○ Poor reproducibility for arteriolar hyalinosis on frozen sections – Significant discrepancy from reproducibility on permanent sections ○ Acute tubular injury shows poor reproducibility on frozen and permanent sections

Maryland Aggregate Pathology Index (MAPI) • Subset (~ 12%) of kidneys from donors unsuitable by clinical criteria have normal histologic features • MAPI can be used to predict likelihood of graft survival at 5 years based on pathologic features • Aggregate score predictive of graft survival ○ Scores 0-7: 90% ○ Scores 8-11: 63% ○ Scores 12-15: 53% • 5 histologic features evaluated, and points for each are totaled ○ Global glomerulosclerosis – If ≥ 15%: 2 points ○ Wall:lumen ratio of interlobular arteries (width of wall at 2 points/diameter of lumen) – If ≥ 0.5: 2 points ○ Periglomerular fibrosis (thickening, wrinkling, and reduplication of Bowman capsule) – If present: 4 points ○ Arteriolar hyalinosis (amorphous, homogeneous eosinophilic deposits in walls of arterioles) – If present: 4 points ○ Scar (focus of sclerosis and renal parenchymal fibrosis and atrophy involving at least 10 tubules) – If present: 3 points

REPORTING Donor Biopsy • Site and type of specimen (wedge or needle core biopsy) • Number of glomeruli, number of globally sclerotic glomeruli, percentage of global glomerulosclerosis 75

Kidney Transplantation

Evaluation of the Donor Kidney Donor Biopsy Reporting and Scoring Sheet Features

Notes

Type of specimen

Wedge biopsy, core needle biopsy

Number of glomeruli

Number of unique glomeruli that are available for evaluation are counted

Number of globally sclerosed glomeruli

Periglomerular sclerosis and focal glomerulosclerosis are included under other findings

Percentage of global glomerulosclerosis

Number of globally sclerosed glomeruli over number of total glomeruli

Number of arteries (not arterioles)

Artery defined as vessel with internal elastic lamina, or diameter > 1/3 diameter of typical glomerulus, or vessel with ≥ 3 layers of smooth muscle

Following Findings Are Scored Score

None

Mild

Moderate

Severe

Interstitial fibrosis

None (< 5% cortex)

Mild (6-25%)

Moderate (26-50%)

Severe (> 50% of cortex involved)

Tubular atrophy

None (0% cortex)

Mild (< 25%)

Moderate (26-50%)

Severe (> 50% of cortical tubules involved)

Interstitial inflammation

None (< 10% cortex)

Mild (10-25%)

Moderate (26-50%)

Severe (> 50% of cortex involved)

Arterial intimal fibrosis (arteriosclerosis)

None (0% luminal narrowing)

Mild (< 25%)

Moderate (26-50%)

Severe (> 50% vascular narrowing)

Arteriolar hyalinosis (hyaline restricted to subendothelial layer)

None

Mild (at least 1 arteriole)

Moderate (> 1 arteriole)

Severe (circumferential, multiple arterioles)

Glomerular thrombi

None

Mild (< 10% of capillaries occluded in most severely affected glomerulus)

Moderate (10-25% Severe (> 25% capillaries occluded) capillaries occluded)

Acute tubular injury or necrosis

None

Mild (epithelial flattening, tubule dilation, nuclear dropout, loss of brush border)

Moderate (focal coagulative type necrosis)

Severe (infarction)

Focal segmental glomerulosclerosis

Nodular glomerulosclerosis

Tumor

Others

Other notable findings

H Liapis et al: Banff Histopathological Consensus Criteria for Preimplantation Kidney Biopsies. Am J Transplant. 22 July 2016.

• Number of arteries (not arterioles) • Histologic features (graded as none, mild, moderate, or severe) ○ Interstitial fibrosis, tubular atrophy, interstitial inflammation, arterial intimal fibrosis, glomerular thrombi • Any other notable feature (e.g., nodular glomerulosclerosis, focal segmental glomerulosclerosis, tumor)

PITFALLS Wedge Biopsies • Sclerotic glomeruli overestimated in wedge biopsies ○ Superficial biopsy may overrepresent globally sclerotic glomeruli, which tend to be subcapsular in patients with arteriosclerosis • Arteries may be absent

Superficial Biopsy

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○ Retraction of epithelial elements can resemble interstitial edema ○ Edema can resemble fibrosis • Tubules ○ Tubules appear retracted – Can be misinterpreted as atrophy or injury ○ Difficult to appreciate acute tubular injury • Red cell casts lyse on freezing

Specimen Handling • Cold ischemic time must be minimized • Biopsies should be kept moist in appropriate preservative ○ Prolonged time in saline can produce artifacts ○ Desiccation can alter appearance

SELECTED REFERENCES 1.

• May only have capsule present

2.

Frozen Section Artifacts

3.

• Glomeruli ○ Glomeruli appear hypercellular • Interstitium

4.

Liapis H et al: Banff histopathological consensus criteria for preimplantation kidney biopsies. Am J Transplant. 17(1):140-150, 2017 Azancot MA et al: The reproducibility and predictive value on outcome of renal biopsies from expanded criteria donors. Kidney Int. 85(5):1161-8, 2014 Haas M: Donor kidney biopsies: pathology matters, and so does the pathologist. Kidney Int. 85(5):1016-9, 2014 Israni AK et al: New national allocation policy for deceased donor kidneys in the United States and possible effect on patient outcomes. J Am Soc Nephrol. 25(8):1842-8, 2014

Evaluation of the Donor Kidney

Donor Biopsy: Wedge Biopsy (Left) Frozen section of a donor biopsy shows apparent interstitial edema, likely the "normal" state of the kidney before dehydration during processing. The tubular morphology is uninterpretable. The evaluation of features predictive of graft function is challenging in specimens with this degree of artifact. (Right) Most donor biopsies are wedge, instead of needle, biopsies, and sample subcapsular tissue (capsule ﬊). This is a reason why intermediate-sized arteries are not often sampled.

Donor Biopsy: Glomerular Thrombi

Kidney Transplantation

Donor Biopsy: Frozen Section

Donor Biopsy: Glomerular Thrombi (Left) Frozen section of a donor biopsy shows fibrin thrombi ﬈ in 2 capillary loops. These can be easily overlooked, especially if they are focal. Thrombi are common in donors who died from stroke or head injury. Scattered thrombi do not contraindicate use of the kidney for transplantation. (Right) Thrombi ﬈ in a glomerulus of a donor biopsy can appear pale on some H&Estained sections, instead of darkly eosinophilic. This can make detection of thrombi difficult.

Donor Biopsy: Arteriosclerosis

Donor Biopsy: Diffuse Global Glomerulosclerosis (Left) Biopsy from a potential donor kidney shows severe arteriosclerosis ﬊. This size artery is not always present in wedge biopsies. A globally sclerotic glomerulus is also seen ﬈. (Right) Donor kidney biopsy shows > 80% globally sclerotic glomeruli ﬈. The biopsy is not from a subcapsular scar. Both the right and left kidneys showed the same finding, a reason for rejecting kidneys for transplantation.

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Kidney Transplantation

Evaluation of the Donor Kidney

Donor Biopsy: Diabetic Donor

Donor Biopsy: Diffuse Diabetic Glomerulosclerosis

Transplanted Kidney Biopsy: Nodular Glomerulosclerosis Not Apparent

Donor Biopsy: Myoglobin Casts

Donor Kidney: Incidental IgA Deposits

Transplanted Kidney Biopsy: Resolving IgA Deposits Post Transplant

(Left) Donor biopsy shows mild arteriolar hyalinosis ﬈, which is difficult to recognize on frozen section. The corresponding permanent section showed diffuse diabetic glomerulosclerosis. (Right) Permanent section of a donor biopsy shows mild to moderate mesangial matrix expansion ﬈, readily recognized on a PAS-stained section. There is also arteriolar hyalinosis ﬊.

(Left) Biopsy 17 days after transplantation of the kidney shows prominent nodular diabetic glomerulopathy ﬊. Both the donor and the recipient were diabetic. The nodules were not apparent in a donor frozen section, even in retrospect. The graft was eventually lost. Donation to a nondiabetic recipient might have been more salutary. (Right) This kidney shows severe tubular injury with granular eosinophilic casts positive for myoglobin ﬈. The cause of death of the donor was trauma.

(Left) Immunofluorescence of a living donor kidney shows IgA deposits in the mesangium ﬉. The donor had no evidence of glomerular disease. This may be an incidental finding in donor biopsies. IgA deposits resolve with time in the recipient after donation. (Right) Immunofluorescence of biopsy 3 months after transplant of a living donor kidney that previously showed prominent IgA deposits in the mesangium now shows loss of the mesangial IgA deposits. The recipient had no clinical evidence of glomerular disease.

78

Evaluation of the Donor Kidney Donor Biopsy: Nodular Arteriolar Hyalinosis (Left) Intimal arteriolar hyalinosis ﬈ is a feature difficult to identify on a frozen section. This feature is more obvious on this permanent section stained with periodic acid-Schiff. (Right) Nodular peripheral arteriolar hyalinosis ﬉ in a donor biopsy looks exactly like the hyalinosis commonly found in chronic calcineurin inhibitor (CNI) toxicity and was once thought to be specific for CNI. This feature, however, is also uncommonly seen in the absence of CNI administration.

Donor Biopsy: Permanent Section

Kidney Transplantation

Donor Biopsy: Arteriolar Hyalinosis

Donor Wedge Biopsy: Frozen Section With Wrinkles (Left) The appearance of kidney parenchyma can be different on frozen section and subsequent permanent section. This biopsy looked markedly edematous on frozen section, and the glomeruli appeared hypercellular. On permanent section, there is only focal, fine interstitial edema and normal-appearing glomeruli ﬈. (Right) Frozen sections are more likely than permanent sections to show wrinkles ﬈ and folds, which can make interpretation more difficult. Additional sections without artifacts should be prepared.

Donor Biopsy: Pediatric Patient

Donor Biopsy: Angiomyolipoma (Left) A donor kidney biopsy from a child shows immature glomeruli ſt with crowded podocytes and 1 glomerulus with focal segmental glomerulosclerosis st, probably developmental. (Right) Unsuspected neoplasms may be found in donor kidneys. This biopsy revealed an angiomyolipoma. The vascular ﬈, smooth muscle ﬉, and adipose tissue ﬊ elements characteristic of this tumor are seen.

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Kidney Transplantation

Evaluation of Transplant Nephrectomy

TERMINOLOGY Synonyms • Kidney explant

Definitions • Irreversible renal allograft failure necessitating nephrectomy

ETIOLOGY/PATHOGENESIS Causes of Early Allograft Loss (< 6 Months Post Transplantation) • Allograft thrombosis ○ Venous thrombosis ~ 2x more frequent than arterial thrombosis ○ Hypercoagulable/thrombophilic states are important underlying causes of thrombosis – Inherited disorders □ Factor V Leiden (G1691A mutation) □ Prothrombin (factor II) mutation □ Protein S, protein C, or antithrombin III deficiency – Acquired disorders □ Tissue factor release from surgery or trauma, antiphospholipid antibodies, hyperhomocysteinemia – Sickle cell disease or trait may give rise to intragraft sickle crisis and graft thrombosis ○ Poor blood flow and vessel wall injury also important factors – Surgical difficulties arising from anatomical discrepancies between donor and recipient vessels – Large vessel injury from torsion, kinks, and compression – Small vessel or endothelial injury related to prolonged ischemia or reperfusion injury • Thrombotic microangiopathy ○ Causes include antibody-mediated rejection and recurrent hemolytic uremic syndrome • Acute allograft rejection (AR) ○ Cell-mediated &/or antibody-mediated rejection

• Recurrent disease, such as primary oxalosis and hemolytic uremic syndrome • Primary nonfunction ○ Clinical term for graft that never functioned ○ Causes include – Acute tubular necrosis (ATN)/injury – Perfusion nephropathy – Atheroembolism arising from plaque rupture at graft harvest – Prolonged cold graft ischemia and marginal donor kidneys are predisposing factors

Causes of Late Allograft Loss (> 6 Months Post Transplantation) • Recurrent disease ○ Focal segmental glomerulosclerosis (FSGS) ○ Glomerulonephritis: IgA nephropathy, membranous nephropathy, membranoproliferative glomerulonephritis • De novo glomerulopathies ○ FSGS ○ Transplant glomerulopathy • AR ○ Antibody-mediated rejection may account for 2/3 of late allograft losses ○ Antibody ± T-cell-mediated rejection • Vascular disease ○ Hypertensive nephrosclerosis • Drug toxicity ○ Calcineurin inhibitor toxicity • Infection ○ Polyomavirus, cytomegalovirus, or adenovirus ○ Recurrent episodes of pyelonephritis • Urinary obstruction • Neoplasms ○ Posttransplantation lymphoproliferative disease ○ Renal cell carcinoma ○ Sarcoma

Transplant Nephrectomy (Left) Hilar arteries, veins, and the renal pelvis and ureter are often absent ſt from kidney transplant nephrectomy specimens. (Right) Allograft loss on day 2 after transplantation demonstrates enlargement (298 g) with diffuse cortical and medullary hemorrhage. Note also the radial pattern of pallor (necrosis) in the cortex ſt.

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Early Allograft Loss

Evaluation of Transplant Nephrectomy

Clinical Presentation • • • •

Oliguria or anuria Flank pain Hematuria Asymptomatic

Clinicopathologic Correlation • Detailed clinical and operative history necessary for final diagnosis • Elective allograft removal often preceded by cessation of immunosuppressive therapy for weeks

MACROSCOPIC General Features • Size ○ Enlargement may be associated with – ATN – AR – Renal vein thrombosis ○ Thrombosis, torsion, and anatomical discrepancies of donor and recipient vasculature are important features ○ Allograft rupture may be seen in first 2-3 weeks after transplantation due to – AR – ATN – Ureteral obstruction – Biopsy procedure – Trauma ○ Small renal allograft is feature of chronic disease (e.g., chronic rejection, infection, and chronic ischemia)

Anatomic Features • Hilar vessels and pelvis frequently not resected at transplant removal • Thrombi may be seen in hilar vascular remnants or in intraparenchymal vessels • Location and extent of necrosis and hemorrhage should be described

Specimen Handling • Save portion of viable cortex for immunofluorescence and electron microscopy

○ Features of AR may be superimposed on other disease processes because of discontinuation of immunosuppression ○ AR may obscure underlying cause of end-stage graft failure, especially if necrosis • Chronic rejection ○ Transplant glomerulopathy and peritubular capillaritis with multilayering of capillary basement membrane ○ Chronic allograft arteriopathy (chronic intimal arteritis) • Allograft fibrosis ○ Infarct scars are wedge-shaped in outer cortex ○ Biopsy sites are linear or band-like coarse scars and may have hemosiderin deposition ○ Large or medium vessel arteriosclerosis &/or stenosis result in chronic allograft ischemia – Subcapsular fibrosis, glomerular obsolescence, and ischemic glomerulopathy ○ Chronic calcineurin inhibitor toxicity results in striped cortical fibrosis – Accompanied by nodular arteriolar hyalinization ○ Chronic rejection or polyomavirus nephropathy: Patchy or diffuse interstitial fibrosis and tubular atrophy

Ancillary Studies • Direct immunofluorescence ○ Detection of immunoglobulins and complement for recurrent and de novo immune complex glomerulonephritis • Indirect immunofluorescence ○ Detection of C4d important in antibody-mediated rejection • Immunohistochemistry ○ Detection of C4d, polyomavirus large T antigen, adenovirus, and cytomegalovirus in paraffin sections • Electron microscopy ○ Podocyte injury detected in recurrent or de novo FSGS ○ Immune complex deposits in recurrent or de novo glomerulonephritis • Serology ○ Donor-specific antibodies to histocompatibility antigens essential diagnostic feature of antibody-mediated rejection

SELECTED REFERENCES 1.

MICROSCOPIC General Features • Necrosis ○ Hemorrhagic or anemic ○ Cortical or both cortical and medullary • Large vessel thrombosis may affect arteries or veins or both • Microvascular thromboses in glomeruli and arterioles are diagnostic of thrombotic microangiopathy • Acute rejection ○ T-cell mediated: Interstitial edema, mononuclear infiltrates, and tubulitis – Intimal or transmural arteritis ○ Antibody mediated: Glomerulitis, peritubular capillaritis, and transmural arteritis with necrosis

Kidney Transplantation

CLINICAL IMPLICATIONS

2. 3. 4.

5.

Morales JM et al: Association of early kidney allograft failure with preformed IgA antibodies to β2-glycoprotein I. J Am Soc Nephrol. 26(3):735-45, 2015 Loupy A et al: The impact of donor-specific anti-HLA antibodies on late kidney allograft failure. Nat Rev Nephrol. 8(6):348-57, 2012 Phelan PJ et al: Renal allograft loss in the first post-operative month: causes and consequences. Clin Transplant. 26(4):544-9, 2012 Sellarés J et al: Understanding the causes of kidney transplant failure: the dominant role of antibody-mediated rejection and nonadherence. Am J Transplant. 12(2):388-99, 2012 El-Zoghby ZM et al: Identifying specific causes of kidney allograft loss. Am J Transplant. 9(3):527-35, 2009

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Kidney Transplantation

Evaluation of Transplant Nephrectomy

Hemorrhagic Necrosis

Reperfusion Injury Mimicking Arteritis

Renal Allograft Venous Thrombosis

Allograft Hilar Venous Thrombus

Hemorrhagic Necrosis From Venous Thrombosis

Sickle Cell Thrombosis

(Left) H&E shows diffuse cortical necrosis with hemorrhage in the pars convoluta from a renal allograft removed post transplant on day 2. Medullary rays have necrosis without hemorrhage. Large arteries have occlusive thrombi. Protein S deficiency and oral contraceptive use contributed to graft thrombosis. (Right) Necrotic arteries often have neutrophils ﬈ and erythrocytes ﬊ in the media and intima and a lot of karyorrhexis. This is a form of reperfusion injury and should not be mistaken for vasculitis.

(Left) A formalin-fixed renal allograft was removed on day 4 post transplant for renal vein thrombosis related to vein injury at implantation. Occlusive thrombi ſt and diffuse pallor of the cortex are consistent with necrosis. The pelvis is filled with a blood clot ﬇. (Right) H&E shows erythrocyte-rich occlusive venous thrombosis in a renal hilar vein from an allograft removed on day 4 post transplant. Layers of fibrin can be seen ﬈.

(Left) Necrosis of the cortex with congestion and hemorrhage ﬈ are seen in a renal allograft removed for renal vein thrombosis on day 5 post transplant. (Right) Vascular thrombi may contain clues to the underlying condition predisposing to graft thrombosis. Multiple refractile, sickled erythrocytes are seen on high-power examination. This patient had a clinically occult sickle cell trait with 26% hemoglobin S.

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Evaluation of Transplant Nephrectomy

Late Allograft Loss With Segmental Scar (Left) Extensive cortical hemorrhage ſt and necrosis is shown in a 160-g renal allograft removed 4 years after transplantation. There was severe arterial ﬇ and arteriolar nephrosclerosis with superimposed severe rejection resulting in hemorrhagic necrosis. (Right) A 186-g renal allograft removed after 8.5 years shows a segmental scar ſt and effacement of the pyramid ﬇. The parenchyma is hemorrhagic. Histologically, there was severe rejection and recurrent lupus nephritis.

Late Rejection Without Immunosuppression

Kidney Transplantation

Late-Onset Allograft Hemorrhagic Necrosis

Plasma Cell-Rich Late Rejection (Left) Late rejection often has interstitial edema, hemorrhage ﬈, plasma cells, eosinophils, and peritubular capillaritis ﬊. C4d was present in the capillary walls. Tubules are atrophic. The findings are indicative of celland antibody-mediated rejection. (Right) Abundant plasma cells are a feature of late-onset allograft rejection. The tubules are severely atrophic, and the glomerulus is obsolescent. These dense infiltrates must be distinguished from posttransplantation lymphoproliferative disorders.

κ-Light Chain RNA In Situ Hybridization

λ-Light Chain RNA In Situ Hybridization (Left) Sheets of infiltrating plasma cells raise the possibility of posttransplantation lymphoproliferative disease. In situ hybridization can differentiate reactive from neoplastic plasma cells. κ-light chain RNA is evident in this section. (Right) λ-light chain RNA can be detected in conjunction with κ-light chain RNA to help differentiate reactive from neoplastic plasma cell infiltrates in renal allografts. Reactive infiltrates have κ:λ ratios of ~ 5-8:1.

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Kidney Transplantation

Evaluation of Transplant Nephrectomy

Late Allograft Nephrectomy

Active Chronic Allograft Arteriopathy

Chronic Allograft Arteriopathy

T Cells in Chronic Arteriopathy

Transplant Glomerulopathy

Chronic Transplant Glomerulopathy

(Left) Renal allograft removed 5.75 years post transplant demonstrates variable thinning and global pallor st of the cortex. Acute and chronic rejection was histologically evident. (Right) Acute and chronic intimal arteritis from an allograft removed after 5 months post transplant shows mononuclear infiltrates deep within the fibrotic intima ﬈. The endothelium is undermined by similar infiltrates ﬊.

(Left) Chronic allograft arteriopathy can have a subtle appearance with intimal fibrosis and scattered nuclei within the fibrotic intima. Immunohistochemistry for CD3 may be necessary to identify T-cell infiltrates. (Right) This artery has CD3(+) T cells deep within the fibrotic intima ﬈, indicative of chronic allograft arteriopathy (chronic intimal arteritis).

(Left) Transplant glomerulopathy has global capillary double contours ﬈ and segmental hypercellularity ﬊. Hyaline arteriolosclerosis is often a prominent feature ﬉. Antibody-mediated rejection accounts for ~ 80% of cases. (Right) Morphologic identification of transplant glomerulopathy raises a differential diagnosis of membranoproliferative glomerulonephritis, chronic thrombotic microangiopathy, and antibody-mediated rejection. Hepatitis C infection often has a contributory role.

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Evaluation of Transplant Nephrectomy

Striped Fibrosis (Left) Chronic allograft ischemia associated with arteriosclerosis may be associated with outer cortical interstitial fibrosis, tubular atrophy, ischemic glomerulopathy ﬈, & global glomerular obsolescence ſt. (Right) A striped pattern of interstitial fibrosis and tubular atrophy ſt is characteristic of chronic calcineurin inhibitor toxicity. Hyaline arteriolosclerosis is found in these cases. Other causes of arteriolosclerosis, such as hypertension and diabetes mellitus, may also give rise to this pattern of fibrosis.

Borderline Infiltrates

Kidney Transplantation

Chronic Allograft Ischemia

Subtle Polyomavirus Infection (Left) Tubulointerstitial mononuclear inflammation affected ~ 20% of the cortex in this 4-year-old allograft. There is also mild tubulitis ﬈. The findings are consistent with a borderline infiltrate by the Banff criteria but raise the differential diagnosis of viral infection. (Right) A kidney allograft with a borderline infiltrate has focal tubular nuclear staining for polyomavirus large T antigen ﬈ indicative of active viral replication without viral cytopathic changes. This is a subtle example of polyomavirus nephropathy.

Acute Pyelonephritis

Recurrent Lupus Glomerulonephritis (Left) This allograft was removed from a 63-year-old man 5 years post transplant. Urine cultures were positive for Escherichia coli. Prominent neutrophilic infiltrates ﬇ and casts ſt are typical of acute pyelonephritis. (Right) Segmental crescentic glomerulonephritis is evident in this allograft removed 7.75 years post transplant. Immunofluorescence showed granular IgG and C3 deposits. The patient had a history of systemic lupus erythematosus. The findings indicate recurrent lupus nephritis.

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Kidney Transplantation

Evaluation of Fibrosis

TERMINOLOGY

MECHANISMS

Definitions

Molecular Mediators

• Interstitial fibrosis: Accumulation of collagen and related molecules in interstitium

• • • • •

VALUE Prognosis • Outcome of wide variety of renal diseases correlates with extent of interstitial fibrosis, even after multivariate analysis ○ Studies show reciprocal correlation between kidney function and fibrosis extent • In renal allografts, extent of fibrosis predicts outcome and may be considered surrogate marker ○ Many applications have been reported ○ Interstitial fibrosis and tubular atrophy (IF/TA) associate with – Cold ischemia time – Clinical and subclinical acute rejection – Preexisting donor damage – Degree of sensitization – Cyclosporine exposure – Renal calcifications ○ IF/TA associated with transplant vasculopathy, ↑ serum creatinine, or transplant glomerulopathy implies poorer prognosis than IF/TA without additional lesions • Fibrosis shows prognostic value in renal donor biopsies ○ ↑ risk of adverse outcome at 6 months – 1.9x greater prediction from age alone with Banff index for IF (ci score > 0) ○ Morphometric interstitial volume: Correlates with graft function at 1 year • Protocol biopsies to assess fibrosis progression can demonstrate baseline state of allograft as well as stepwise changes that occur ○ Useful in clinical trials to assess outcome

Transforming growth factor Bone morphogenetic protein Platelet-derived growth factor Hepatocyte growth factor Recent genomic approaches show altered molecular factors in IF

Cellular Mediators • Epithelial cells • Fibroblasts/myofibroblasts and fibrocytes • Inflammatory cells: Lymphocytes, monocyte/macrophages, dendritic cells, mast cells • Endothelial cells

Epithelial-to-Mesenchymal Phenotype • Chronically injured epithelial cells may undergo transition to mesenchymal cells ○ Process called "epithelial-to-mesenchymal transition" (EMT), but term has fallen out of favor ○ So-called EMT may simply reflect change in protein expression rather than true transition • Injured epithelium may change morphology and express mesenchymal-like markers ○ Actual EMT process not observed in vivo • Mesenchymal markers not entirely specific, making research questionable (per Banff conferences and other publications) ○ Epithelial-to-mesenchymal phenotype may be more appropriate term since changes may represent altered phenotype

METHODS FOR ASSESSMENT Qualitative Visual Assessment • Not all fibrosis is "equal" or "same" in quality and quantity ○ "Early," "young," or "active" fibrosis may have greater potential for remodeling

Collagen III Immunohistochemistry (Left) Collagen III immunohistochemistry stains fibrous tissue brown. (Right) A positive pixel count algorithm is applied to the immunohistochemistry, and a "markup" image shows areas that are strongly positive (red), medium positive (orange), and negative (blue).

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Collagen III Immunohistochemistry Quantitation "Markup"

Evaluation of Fibrosis

• •

•

•

Quantitative Visual Assessment • Most fibrosis scoring systems (notably Banff) based on quantitation of percentage of cortical parenchyma involved ○ Banff fibrosis (termed "ci score") uses following cutoffs – ci0: ≤ 5%, ci1: 6-25%, ci2: 26-50%, ci3: > 50% • Special stains ○ Trichrome – Visual assessment of trichrome-stained slides often standard practice – Stains glomerular and tubular basement membranes in addition to areas of fibrosis ○ Periodic acid-Schiff (PAS) – PAS stains glomerular and tubular basement membranes "hot pink" with less staining in interstitium – Trichrome also stains glomerular and tubular basement membranes; thus, assessment of stains, such as PAS, can take these basement membranes into account and allow pathologist to appreciate only interstitium – Morphometric methods used to "subtract" basement membranes from trichrome using PAS (so-called trichrome-PAS or T-P method) ○ Sirius red – Pinkish red staining of most tissues under white light – Dye molecule intercalates into tertiary groove of collagen I and III – Collagen I and III strongly birefringent when observed under polarized light □ Considered specific for these collagens ○ Collagen immunohistochemistry (IHC) – Collagen type III IHC particularly useful for assessing fibrosis in kidney • Repeat biopsies may show differing levels of fibrosis, presumably due to sampling ○ Decreased IF in 12% of cases according to 1 study

• Visual fibrosis assessment susceptible to inter- and intraobserver variability ○ Some pathologists consistently overgrade and some undergrade ○ Kappa values (statistical measure of interobserver agreement) on order of 0.3-0.6 reported

Morphometric Quantitative Assessment • Analysis in morphometric studies correlated with function (e.g., eGFR) • Point counting techniques on static images traditionally utilized • Computer-assisted morphometry shows utility in analysis of stains (e.g., trichrome, Sirius red, collagen IHC) ○ Many methods use pixel-counting algorithms (also called "positive pixel count algorithm") – Available algorithms supplied by commercial vendors and "open sources" (e.g., ImageJ provided by National Institutes of Health) ○ Customized algorithms written by individual researchers also employed – Kappa value of 0.68 compared to expert Banff quantification obtained with analysis method described by Meas-Yedid et al – Computerized algorithms can apply to other organs and measure other parameters (e.g., liver steatosis, inflammatory cell counts, microvessel density, etc.) ○ Methods currently utilize whole-slide images as opposed to static images • May provide more objective, reproducible measurement method

Kidney Transplantation

•

○ Broad scars: Pyelonephritis and infarcts can produce severe focal injury and parenchymal destruction ○ Diffuse, fine fibrosis: Diffuse disease of glomeruli, tubules, and vessels Fibrosis patterns may have different implications ○ "Striped," patchy fibrosis described with calcineurin inhibitor use, possibly due to preferential medullary ray involvement ○ Chronic obstructive pattern: Atubular glomeruli, dilated tubules, and intratubular Tamm-Horsfall protein casts with interstitial extravasation Inflammation in areas of IF noted to be adverse risk factor for renal disease progression IF/TA typically correlate ○ TA may be profound in renal artery stenosis with little or no accompanying fibrosis ○ IF/TA (graded I-III based on same cutoffs as ci) has replaced term "chronic allograft nephropathy" Subcapsular, perivascular, and periglomerular fibrosis typically not included but objective exclusionary criteria lacking Fibrosis assessment typically focuses on cortex but medullary fibrosis also important

SELECTED REFERENCES 1.

2.

3. 4. 5. 6. 7.

8.

9. 10.

11. 12. 13. 14.

Farris AB et al: Banff fibrosis study: multicenter visual assessment and computerized analysis of interstitial fibrosis in kidney biopsies. Am J Transplant. 14(4):897-907, 2014 Haas M: Chronic allograft nephropathy or interstitial fibrosis and tubular atrophy: what is in a name? Curr Opin Nephrol Hypertens. 23(3):245-50, 2014 Farris AB et al: Renal interstitial fibrosis: mechanisms and evaluation. Curr Opin Nephrol Hypertens. 21(3):289-300, 2012 Farris AB et al: Morphometric and visual evaluation of fibrosis in renal biopsies. J Am Soc Nephrol. 22(1):176-86, 2011 Kriz W et al: Epithelial-mesenchymal transition (EMT) in kidney fibrosis: fact or fantasy? J Clin Invest. 121(2):468-74, 2011 Liu Y: Cellular and molecular mechanisms of renal fibrosis. Nat Rev Nephrol. 7(12):684-96, 2011 Meas-Yedid V et al: New computerized color image analysis for the quantification of interstitial fibrosis in renal transplantation. Transplantation. 92(8):890-9, 2011 Scian MJ et al: Gene expression changes are associated with loss of kidney graft function and interstitial fibrosis and tubular atrophy: diagnosis versus prediction. Transplantation. 91(6):657-65, 2011 Boor P et al: Renal fibrosis: novel insights into mechanisms and therapeutic targets. Nat Rev Nephrol. 6(11):643-56, 2010 Mannon RB et al: Inflammation in areas of tubular atrophy in kidney allograft biopsies: a potent predictor of allograft failure. Am J Transplant. 10(9):206673, 2010 Park WD et al: Fibrosis with inflammation at one year predicts transplant functional decline. J Am Soc Nephrol. 21(11):1987-97, 2010 Zeisberg M et al: Mechanisms of tubulointerstitial fibrosis. J Am Soc Nephrol. 21(11):1819-34, 2010 Serón D et al: Protocol biopsies in renal transplantation: prognostic value of structural monitoring. Kidney Int. 72(6):690-7, 2007 Furness PN et al: International variation in the interpretation of renal transplant biopsies: report of the CERTPAP Project. Kidney Int. 60(5):19982012, 2001

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Kidney Transplantation

Evaluation of Fibrosis

Subcapsular Fibrosis

Subcapsular Fibrosis

"Striped" Fibrosis

Fibrosis Quantitation Approaches Used by Pathologists

Fibrosis Level at Baseline

Quantitation "Markup" of Fibrosis at Baseline Level

(Left) Medium-power view of trichrome of a wedge donor biopsy shows subcapsular fibrosis ſt with chronic inflammation, a finding that is not uncommon. (Right) Higher power trichrome image shows the subcapsular fibrosis with chronic inflammation ſt as well as segmental st and global ﬇ glomerulosclerosis.

(Left) Trichrome-stained kidney section from an autopsy of a patient with a history of lung transplantation and longstanding cyclosporine use shows prominent medullary ray fibrosis ﬈, socalled "striped" fibrosis. (Courtesy S. Rosen, MD.) (Right) Two approaches for assessing fibrosis used by pathologists include (A) the percentage of tissue occupied by fibrous tissue and (B) the percentage of morphologically abnormal tissue.

(Left) A case with an essentially baseline level of fibrosis shows diffuse, fine fibrous tissue st between the renal tubules on trichrome stain. (Right) Quantitation of the blue-staining areas of the trichrome stain is accomplished by using a positive pixel count algorithm tuned to the blue fibrous tissue. Markup image shows areas that are considered positive by the algorithm in orange, resulting in a measurement of the percentage of tissue involved by fibrosis.

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Evaluation of Fibrosis Quantitation "Markup" of Trichrome and PAS in Fibrosis Assessment (Left) Trichrome (upper 1/2) stains focal areas of fibrous tissue ﬈, basement membranes, vessels ﬉, and proteinaceous casts ﬊. The latter 3 can be subtracted through their detection on a PAS stain (lower 1/2) to determine degree of fibrosis. (Right) Markup images detect areas (yellow, orange) on a positive pixel count algorithm tuned for blue on trichrome (upper 1/2) and pink on PAS (lower 1/2). Subtracting PAS from trichrome using so-called "trichrome-PAS" method gives a measurement of interstitium.

Sirius Red Fibrosis Staining

Kidney Transplantation

Trichrome and PAS in Fibrosis Assessment

Sirius Red Fibrosis Areas Detected (Left) Sirius red stains areas of fibrosis red. (Right) Detection of the red-staining areas on a Sirius red stain provides a measurement of the areas of fibrosis. The quantitation in this case is performed without polarization, and this markup image shows the areas considered positive in black, illustrating the fibrous tissue "skeleton" of the kidney. (Courtesy P. Grimm, MD.)

Sirius Red Polarization Detects Fibrosis

Sirius Red Polarized Fibrotic Areas Detected (Left) Polarization of a Sirius red-stained kidney shows areas of fibrous tissue deposition. Note the absence of staining in the glomerular basement membrane st, demonstrating how the glomerular basement membrane is composed of collagen type IV as opposed to collagen types I and III in the interstitium. Interstitial fibrous tissue has a characteristic birefringence ﬇. (Right) Markup image is used in the quantitation of the fibrous tissue birefringent on the Sirius red stain.

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Kidney Transplantation

Protocol Biopsies

TERMINOLOGY

MICROSCOPIC

Synonyms

General Features

• Surveillance biopsies

• Variable findings depending on diagnosis

Definitions

Glomeruli

• Renal allograft biopsies performed at predetermined intervals, unrelated to graft dysfunction

• • • • • •

CLINICAL IMPLICATIONS Value of Protocol Biopsies • Useful in monitoring highly sensitized patients • Ability to detect and potentially treat subclinical disease when it may be reversible ○ Includes subclinical rejection, infection, and recurrent disease • Findings on earlier protocol biopsies may identify cause of later graft loss ○ Specific cause of graft loss can be identified in ~ 95% of cases, largely based on earlier biopsies • Frequency of subclinical rejection low in modern era ○ Other diseases may be detected: Drug toxicity, polyomavirus infection, recurrent disease • Clinical trials ○ Document outcome ○ Detect toxicity • Used more commonly in high-risk (presensitized) patients

Risk of Biopsy Procedure • 0.4% risk of major complications ○ Hemorrhage ○ Peritonitis from bowel perforation ○ Graft loss (< 0.05%)

Timing • Implantation ("time zero") • 3-4, 6, 12, 24 months • Later posttransplant (5 and 10 years) biopsies may give insights into late graft loss

Normal Duplication of glomerular basement membrane Mesangial hypercellularity Focal segmental glomerulosclerosis Immune complex deposition if recurrent or de novo disease Late (~ 10 years) posttransplant biopsies show high rate of glomerular disease ○ Mesangial sclerosis, glomerulomegaly, focal segmental glomerulosclerosis, diabetic glomerulosclerosis, increased global glomerulosclerosis

Tubules • • • •

Normal Tubular atrophy Tubulitis Viral inclusions

Interstitium • Normal • Fibrosis • Inflammation ○ Inflammation in areas of fibrosis associated with increased risk of graft loss ○ Inflammation outside areas of fibrosis may indicate subclinical cellular rejection

Arteries and Arterioles • Normal • Arteriosclerosis • Endarteritis ○ More likely to find endothelialitis without significant tubulointerstitial inflammation ("isolated V lesion") on protocol biopsy than on biopsy for cause • Hyalinosis in arterioles

Peritubular Capillaritis on Protocol Biopsy (Left) Peritubular capillaritis (PTCitis) ﬊ and glomerulitis ﬈, features of subclinical AMR, are seen in this protocol biopsy, 4 months post transplant, from a patient with preformed donor-specific antibody and normal renal function. PTCitis should not be confused with acute cellular rejection. (Right) Acute cellular rejection, Banff type IB, is seen on this 4-month posttransplant protocol biopsy. The serum creatinine was stable at 1.6 mg/dL. The patient was on a steroid-free maintenance immunosuppressive protocol.

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Acute Cellular Rejection on Protocol Biopsy

Protocol Biopsies

Peritubular Capillaries • Normal • C4d(+) peritubular capillaries (PTC) ○ Particularly in ABO blood group-incompatible transplants • Peritubular capillaritis (mononuclear cells and neutrophils)

DIAGNOSES

○ Of patients with moderate fibrosis (ci2) on 1-year protocol biopsy, > 50% show mild or no fibrosis at 5 years ○ Findings may in part represent sampling error ○ Moderate to severe IFTA present in only 12% of 10-year protocol biopsies • Combination of interstitial fibrosis and mononuclear inflammation increases risk of later graft loss ○ Now considered pattern of chronic T-cell-mediated rejection

Chronic Antibody-Mediated Rejection

• 5% prevalence on 1-year protocol biopsies (prior to BK monitoring)

• 40% incidence of subclinical antibody-mediated rejection (sAMR) in protocol biopsies of patients with de novo antidonor-specific antibodies (DSA) • 14% prevalence of sAMR on 1-year protocol biopsies in one series ○ Mostly in patients with DSA present prior to transplant (preformed DSA), only 22% with de novo DSA – Histologically characterized by glomerular &/or peritubular capillaritis □ Minority (32%) are C4d(+) ○ Worse graft survival (56% at 8 years post transplant) compared to subclinical T-cell-mediated rejection – C4d positivity also confers worse graft survival

Chronic Calcineurin Inhibitor Toxicity

Transplant Glomerulopathy

• Arteriolar hyalinosis in 61.3%, 90.5%, and 100% of biopsies at 1, 5, and 10 years, respectively, in patients on cyclosporine in 1 study ○ Not necessarily specific for calcineurin inhibitor (CNI) toxicity • Arteriolar hyalinosis may be less prevalent and less severe in patients on tacrolimus or sirolimus vs. cyclosporine ○ In another study, 19% prevalence of moderate to severe hyalinosis on 5-year protocol biopsies in series of patients on tacrolimus-based immunosuppression – 5% of sirolimus-based, CNI-free immunosuppression showed moderate to severe hyalinosis

• ~ 2-5% prevalence of transplant glomerulopathy in 1-year protocol biopsies of conventional transplant patients ○ Increased risk in patients with DSA ○ Reduced graft survival with TG finding on protocol biopsy

Normal Histology • Comprises majority of protocol biopsies

Subclinical Acute T-Cell-Mediated Rejection • 3-17% in first 5 months • 4-15% of 6-month protocol biopsies • 5-19% in 1-year protocol biopsies ○ Includes borderline/suspicious category in most series • Incidence lower after 1st year and on tacrolimus (5%)

Subclinical BK Polyomavirus Nephropathy

Arteriosclerosis • May be donor disease or de novo ○ Comparison to time-zero (implantation) biopsy is helpful

Interstitial Fibrosis and Tubular Atrophy, Not Otherwise Specified • 81% of cases of graft loss with interstitial fibrosis and tubular atrophy (IFTA) have identifiable cause, in part detected on earlier biopsies • ~ 60% of functioning grafts at 5 years have some IFTA (ct or ci > 0) ○ Moderate to severe IFTA (Banff ci > 1, ct > 1) in 17% of biopsies at 5 years – Associated with previous acute cellular rejection and BK polyomavirus infection episodes • IFTA does not necessarily progress with time ○ In study of conventional transplants, allografts with mild fibrosis (ci1) on 1-year protocol biopsy often had different findings on 5-year biopsy – 39% showed no fibrosis (ci0) on 5-year biopsy, while only 23% showed more severe fibrosis on 5-year biopsy

Kidney Transplantation

○ Moderate to severe arteriolar hyalinosis present in 66% of 10-year posttransplant protocol biopsies (predominantly tacrolimus-based immunosuppression)

Accommodation • No active rejection • C4d(+) PTC • Common in ABO blood group-incompatible transplants

Recurrent Glomerular Disease • Early recurrent membranous glomerulonephritis and IgA nephropathy are commonly subclinical

De Novo Glomerular Disease • Focal glomerulosclerosis and membranous glomerulonephritis are most common • De novo C1q nephropathy common, usually of little clinical significance

SELECTED REFERENCES 1.

2. 3. 4.

5. 6.

Stegall MD et al: Renal allograft histology at 10 years after transplantation in the tacrolimus era: evidence of pervasive chronic injury. Am J Transplant. 18(1):180-188, 2018 Huang Y et al: Protocol biopsies: utility and limitations. Adv Chronic Kidney Dis. 23(5):326-331, 2016 Mehta R et al: Subclinical rejection in renal transplantation: reappraised. Transplantation. 100(8):1610-8, 2016 Yamamoto T et al: De dovo anti-HLA DSA characteristics and subclinical antibody-mediated kidney allograft injury. Transplantation. 100(10):2194202, 2016 Loupy A et al: Subclinical rejection phenotypes at 1 year post-transplant and outcome of kidney allografts. J Am Soc Nephrol. 26(7):1721-31, 2015 Mengel M et al: The molecular phenotype of 6-week protocol biopsies from human renal allografts: reflections of prior injury but not future course. Am J Transplant. 11(4):708-18, 2011

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Kidney Transplantation

Protocol Biopsies

Normal Glomerulus by EM

Normal Peritubular Capillary by EM

Reactive Endothelium by EM

Normal Protocol Biopsy

Mesangial Sclerosis on Late Protocol Biopsy

Arteriolar Hyalinosis

(Left) A normal glomerulus is seen in a protocol biopsy. Podocyte foot processes are preserved ﬈. No glomerular basement membrane (GBM) duplication or subendothelial lucency is present. No immune deposits are noted. Endothelial cells appear normal with preserved fenestrations ﬊. (Right) Here, a normal peritubular capillary is seen in a protocol biopsy specimen. The capillary shows a single basement membrane layer ﬊ and normalappearing endothelial cells.

(Left) Enlarged endothelial cells ﬈ with loss of fenestrations are seen by electron microscopy (EM) on a protocol biopsy 1 month post transplant in a patient with preformed donor-specific antibodies. This finding may precede development of transplant glomerulopathy. The biopsy appeared normal by light microscopy (LM). (Right) A 1-year protocol biopsy specimen in a patient with normal renal function is shown. The biopsy appears normal.

(Left) This 10-year posttransplant protocol biopsy shows moderate mesangial sclerosis ﬈, which can be nodular and is a feature often present on late posttransplant biopsies even in patients without diabetes. (Right) Rare arterioles show intimal ﬉ and peripheral nodular ﬈ hyalinosis, likely due to subclinical calcineurin inhibitor toxicity. Arteriolar hyalinosis becomes more common with time post transplant.

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Protocol Biopsies

De Novo C1q Nephropathy (Left) Segmental granular mesangial ﬈ staining for IgA is seen on this protocol biopsy 5 years post transplant in a patient with a history of IgA nephropathy in the native kidney. The glomeruli showed mild mesangial hypercellularity by LM. (Right) Immunofluorescence (IF) staining for C1q shows 2+ granular mesangial staining ﬈ on a protocol biopsy 10 years post transplant. By LM, glomeruli showed mesangial hypercellularity. This finding is usually of no clinical significance.

Early Transplant Glomerulopathy

Kidney Transplantation

IgA Deposits on Protocol Biopsy

Early Recurrent Membranous Nephropathy (Left) Early transplant glomerulopathy is seen in a 1year protocol biopsy. Glomerulitis ﬈ and segmental GBM duplication ﬊ are seen. This patient also had preformed donor-specific HLA antibody but normal renal function. (Right) This glomerulus from early recurrent membranous nephropathy does not show evidence of GBM abnormalities ("spikes" or "pinholes") on high magnification. No deposits were noted on a trichrome stain.

Early Recurrent Membranous Nephropathy by IF

Early Recurrent Membranous Nephropathy on EM (Left) In early recurrent membranous nephropathy, IF staining for C4d reveals granular GBM staining in a membranous pattern ſt, in contrast to usual and normal glomerular C4d staining restricted to the mesangium. Similar staining was seen for IgG. (Right) A 4-month protocol biopsy specimen in a patient with early recurrent membranous nephropathy and minimal proteinuria shows no electron-dense deposits by EM, despite IgG and complement deposits by IF. Podocyte foot processes are intact ﬈.

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Kidney Transplantation

Accommodation KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Accommodation refers to lack of graft injury in presence of donor-reactive immune response, usually detected by circulating antidonor antibodies ○ Commonly, but not always, occurring in ABOincompatible (ABOi) grafts (> 80%) • Corresponding Banff definition: Peritubular capillary (PTC) C4d deposition without evidence of rejection

• • • •

CLINICAL ISSUES • Normal graft function • No treatment indicated for ABOi transplants • No established treatment in patients with preformed antiHLA DSA and normal graft function • In ABOi living donor grafts, similar graft survival as conventional transplants (87% vs. 92% 9-year survival) ○ Presence of C4d in 12-month protocol biopsies has no prognostic significance in absence of inflammation (borderline or more)

Normal graft biopsy C4d deposition in PTC No capillaritis Normal endothelial cells, glomerular basement membrane (GBM), and PTC by electron microscopy

TOP DIFFERENTIAL DIAGNOSES • Chronic antibody-mediated rejection ○ Glomerulitis and PTC ○ Multilamination of PTC basement membrane and GBM

DIAGNOSTIC CHECKLIST • C4d(+) biopsies generally stable in ABOi grafts in absence of tubulointerstitial inflammation or microvascular inflammation • C4d(+) biopsies in recipients with anti-HLA DSA, even in absence of inflammation, generally progress to transplant glomerulopathy

Normal Cortex

Normal Parenchyma

C4d

Peritubular Capillary

(Left) Protocol biopsy of an ABO-incompatible graft 3 months after transplantation appears entirely normal by light microscopy. (Right) Protocol biopsy is shown from a patient with stable renal function with an HLAincompatible, ABO-compatible graft. The appearance by light microscopy is entirely normal, although C4d was present in peritubular capillaries.

(Left) Protocol biopsy is shown from a patient with stable renal function with an HLAincompatible, ABO-compatible graft. C4d is in the peritubular capillaries ﬈ and glomerular ﬈ basement membranes in the absence of inflammation. (Right) Protocol biopsy of a C4d(+), normal-appearing ABO-incompatible graft 3 months after transplantation shows entirely normal peritubular capillaries endothelium, indicative of accommodation.

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Accommodation

Definitions • Refers to lack of graft injury in presence of donor-reactive immune response, usually by circulating antidonor antibody detection ○ Commonly, but not always, occurring in ABOincompatible (ABOi) grafts • Corresponding Banff definition: Peritubular capillary C4d without evidence of rejection ○ Linear C4d staining in peritubular capillaries (PTC) (C4d2 or C4d3 by immunofluorescence on frozen sections or C4d > 0 by IHC on paraffin sections) ○ Criteria for chronic, active antibody-mediated rejection (ABMR) not met ○ No molecular evidence for ABMR (if molecular studies performed) (Banff 2017) ○ No acute or chronic T-cell-mediated rejection

ETIOLOGY/PATHOGENESIS Endothelial Response to Antibodies Reactive to Surface Components • Increased antiapoptotic molecules (Bcl-xL), complement regulatory proteins (CD55, CD59), and MUC1 in vivo • In cultured endothelial cells, ABO antibodies inactivate ERK1/2 pathway and ↑ CD55 and CD59; anti-HLA ligation activates PI3K/AKT and ↑ hemeoxygenase-1 and ferritin H

Prognosis • In ABOi living donor grafts, similar graft survival as conventional transplants (87% vs. 92% 9-year survival) ○ Presence of C4d has no prognostic significance in absence of inflammation (borderline or more) or microvascular inflammation • Positive-crossmatch (+XM) patients with preformed antiHLA DSA ○ Early protocol biopsies (< 3 months) may show minor changes by light microscopy ○ Majority of +XM recipients develop chronic ABMR, increasing incidence with time post transplant

MICROSCOPIC Histologic Features • Normal graft biopsy, no evidence of rejection • Glomeruli: No glomerulitis, no glomerular basement membrane (GBM) duplication • Interstitium: No interstitial infiltrate (< 10%) • Tubules: No tubulitis (< 5 cells/tubule) • Arteries: No arteritis (endothelialitis) • PTC: No capillaritis

ANCILLARY TESTS Immunofluorescence • Positive C4d staining in PTC

Antibody Production

Electron Microscopy

• Evidence in human ABOi kidney transplants for diminished antiblood group antibody production by peripheral blood B cells

• Lack of reactive endothelial cells in glomeruli and PTC • Lack of multilamination of PTC basement membranes • Lack of GBM duplication

Animal Models • In nonhuman primates, 4 stages of chronic antibodymediated rejection (cAMR) ○ Stage I: Circulating donor-specific antibody (DSA) (accommodation 1) ○ Stage II: DSA + C4d in PTC (accommodation 2) ○ Stage III: DSA + C4d + pathologic changes (subclinical cAMR) ○ Stage IV: DSA + C4d + pathology + functional impairment

CLINICAL ISSUES Epidemiology • Incidence ○ Commonly occurs in ABOi grafts [> 80% C4d(+) on protocol biopsies] ○ Incidence in HLA-incompatible grafts uncertain (2-4% in early protocol biopsies), unstable long term

Presentation • Normal graft function • Positive antibodies to donor HLA or ABO antigens

DIFFERENTIAL DIAGNOSIS Chronic Antibody-Mediated Rejection • Glomerulitis and PTC (Banff ptc + g ≥ 2) • Multilamination of PTC basement membrane and GBM

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Accommodation generally stable in ABOi grafts • Accommodation with anti-HLA DSA, especially if C4d(+) on biopsy, generally progresses to transplant glomerulopathy

SELECTED REFERENCES 1.

2.

3.

4.

Treatment • No treatment indicated for ABOi transplants • No established treatment in patients with preformed antiHLA DSA and normal graft function

Kidney Transplantation

TERMINOLOGY

5.

6.

Masutani K et al: Histological analysis in ABO-compatible and ABOincompatible kidney transplantation by performance of 3- and 12-month protocol biopsies. Transplantation. 101(6):1416-1422, 2016 Okumi M et al: ABO-incompatible living kidney transplants: evolution of outcomes and immunosuppressive management. Am J Transplant. 16(3):886-96, 2016 Bentall A et al: Differences in chronic intragraft inflammation between positive crossmatch and ABO-incompatible kidney transplantation. Transplantation. 98(10):1089-96, 2014 Iwasaki K et al: Comparative study on signal transduction in endothelial cells after anti-a/b and human leukocyte antigen antibody reaction: implication of accommodation. Transplantation. 93(4):390-7, 2012 Wiebe C et al: Evolution and clinical pathologic correlations of de novo donor-specific HLA antibody post kidney transplant. Am J Transplant. 12(5):1157-67, 2012 Smith RN et al: Four stages and lack of stable accommodation in chronic alloantibody-mediated renal allograft rejection in Cynomolgus monkeys. Am J Transplant. 8(8):1662-72, 2008

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Kidney Transplantation

Tolerance KEY FACTS

TERMINOLOGY • Tolerance: State of immunologically specific acceptance of tissue or organs without destructive immune response, without immunosuppressive drugs, and with full immunologic reactivity to other antigens (e.g., microbes or 3rd-party allografts)

ETIOLOGY/PATHOGENESIS • Mixed chimerism ○ Bone marrow and kidney allografts from same living donor given to recipient conditioned with thymic irradiation, cyclophosphamide (or whole-body irradiation), calcineurin inhibitors, rituximab, and T-celldepleting antibodies • Ad hoc withdrawal of immunosuppression ○ In some (unpredictable) cases, allografts continue to function normally ○ Increased number of circulating transitional B cells • Mechanisms

○ Deletional tolerance (central tolerance) – Normal mechanism for preventing autoimmunity ○ Regulatory tolerance (peripheral tolerance) – Mediated by regulatory cells (Treg), FOXP3(+)

MICROSCOPIC • Protocol biopsies of stable kidney allografts off immunosuppression limited in number but reveal spectrum of patterns ○ Normal kidney ○ FOXP3(+) cells in interstitial and perivascular aggregates – Treg-rich organized lymphoid structures ○ Accommodation [C4d(+)] ○ Slowly progressive subclinical chronic antibody-mediated rejection ○ Recurrent glomerular disease

DIAGNOSTIC CHECKLIST • Renal biopsy essential in proving state of tolerance

Normal Kidney Protocol Biopsy

Lymphoid Aggregates in Accepted Allograft

Aggregates of FOXP3(+) Cells in Accepted Allograft

Treg-Rich Organized Lymphoid Structure

(Left) This is a 2-year protocol biopsy on a patient with normal renal function who has been off immunosuppression for 18 months. Tolerance was induced with donor bone marrow and the mixed chimerism conditioning protocol. The kidney is normal. (Right) This protocol biopsy in an accepted renal allograft from a patient with normal renal function off immunosuppression 7 years after transplantation shows aggregates of lymphoid cells st, which typically contain (Treg) FOXP3(+) cells.

(Left) Protocol biopsies from accepted renal allografts off immunosuppression often show aggregates of lymphoid cells rich in FOXP3(+) cells st. Double stain for FOXP3 (blue) and CD4 (brown) is shown. (Right) Treg-rich organized lymphoid structures are common in recipients who are tolerant of their renal allografts. These are seen in human, nonhuman primate, pig, and mouse renal allografts (shown) after tolerance induction by a variety of protocols. Double IHC stain for FOXP3 (blue) and CD3 (brown) is shown.

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Tolerance

Definitions • Tolerance: State of immunologically specific acceptance of tissue or organs ○ Without immunosuppressive drugs ○ Without destructive immune response ○ With full immunologic reactivity to other antigens, such as microbes or 3rd-party allografts • While achieved in inbred mice with variety of protocols, only recently tolerance intentionally induced in human kidney allografts

ETIOLOGY/PATHOGENESIS Protocols to Induce Tolerance in Mice • Many strategies, such as neonatal injection of donor cells (Medawar et al) and costimulation blockade ○ Work in mice ○ Only successful protocol in humans administered donor bone marrow cells • Administration of donor bone marrow with nonmyeloablative regimen in mice leads to stable leukocyte mixed chimerism and tolerance of organ or skin grafts (Sachs and Sykes)

Protocols to Induce Tolerance in Humans • Mixed chimerism (Kawai et al) ○ Bone marrow and kidney allografts from same living donor given to recipient conditioned with thymic irradiation, cyclophosphamide (or whole-body irradiation), calcineurin inhibitors, rituximab, and T-celldepleting antibodies ○ Leads to transient mixed chimerism ○ 2 protocols: Myeloma (HLA-identical) and primary kidney disease (HLA haploidentical) • Mixed chimerism with total lymphoid irradiation [(TLI) Strober et al] ○ Mixed chimerism persists ○ Long-term survival of HLA matched recipients off immunosuppression ○ HLA mismatched recipients maintained on low-dose tacrolimus • Spontaneous tolerance ○ Occasional kidney allograft recipients discontinued immunosuppressive drugs after several years for various reasons – In some, their allografts continued to function normally ○ DESCARTES-Nantes survey identified 61 tolerant recipients from pool of 218,983 patients (0.03%) • Full chimerism (Leventhal et al) ○ Bone marrow ablation and replacement with donor cells ± facilitator cells • Cell therapy (ONE study) ○ Administration of induced Treg (± donor reactive) and other cell populations currently under investigation – Donor-reactive Treg – Polyclonal Treg

Mechanisms • 2 principal physiological mechanisms to avoid autoimmunity or hypersensitivity • Deletional tolerance (central tolerance) ○ Newly formed self-reactive T cells deleted in thymus ○ Irreversible • Regulatory tolerance (peripheral tolerance) ○ Effector T cells inhibited by antigen-specific T cells (Treg) – Best defined subpopulation expresses FOXP3 ○ This form of tolerance may be lost if Treg depleted or if their function impaired ○ Experimental evidence that Treg in allografts can mediate tolerance

Kidney Transplantation

TERMINOLOGY

CLINICAL ISSUES Laboratory Tests • Investigation seeks to identify patients able to withdraw immunosuppressive drugs ○ Rebollo-Mesa et al described 9-gene set signature in blood lymphocytes that needs confirmation in prospective studies

Prognosis • Most recent data from presentations at 3rd International Workshop on Clinical Tolerance ○ Limited experience ○ Studies are small, heterogenous, and protocols evolving • Stanford mixed chimerism series ○ Tolerance achieved in 22/23 HLA-identical recipients off immunosuppression (Stanford TLI) – ~ 10% had acute T-cell-mediated rejection – Recurrent glomerular disease reported □ Lupus nephritis (1/3) □ IgA nephropathy (5/16); 4 on protocol biopsy □ Membranous glomerulonephritis (1/2) □ Focal segmental glomerulosclerosis (1/1) ○ HLA mismatched recipients remain on low-dose tacrolimus – ~ 25% had episode of T-cell-mediated rejection (4/19) – Donor-specific antibodies (DSA) developed in 4/19 □ Lost DSA when resumed immunosuppression □ Chronic antibody-mediated rejection (AMR) with C4d and DSA in ~ 10% ○ No severe infections or graft-vs.-host disease (GVHD) ○ 2 cases of engraftment syndrome • Massachusetts General Hospital mixed chimerism series ○ Long-term graft survival achieved in all HLA-identical and majority of haploidentical recipients after immunosuppressive drugs withdrawn ○ Recurrence of IgA nephropathy and C3 nephropathy reported ○ No severe infections or GVHD ○ Engraftment syndrome at 10-14 days post transplant in early series ○ Minority developed chronic AMR • Chicago full chimerism series ○ 71% of 31 patients followed for > 1 year off immunosuppression ○ GVHD reported (2 cases; 1 fatal) ○ No serious infections or DSA 97

Kidney Transplantation

Tolerance

98

○ No recurrence of glomerular disease [IgA nephropathy (7), membranous nephropathy (2), focal segmental glomerulosclerosis (1)] ○ No engraftment syndrome

MICROSCOPIC Histologic Features • Protocol biopsies of stable kidney allografts off immunosuppression limited in number but reveal spectrum of patterns ○ Normal kidney – Reported in HLA-identical grafts in myeloma patients on mixed chimerism protocol – No appreciable lymphoid infiltrate, no evidence of AMR (C4d) – Compatible with deletional tolerance ○ Treg-rich organized lymphoid structures – FOXP3(+) cells in interstitial and perivascular aggregates – Observed in HLA-haploidentical kidney allografts in humans and nonhuman primates on mixed chimerism protocol – Probable signature pattern of regulatory tolerance ○ Accommodation – Several recipients developed donor-specific HLA antibodies and C4d deposition in peritubular capillaries without simultaneous evidence of rejection either pathologically or clinically ○ Subclinical chronic AMR – Progression to chronic AMR manifested by transplant glomerulopathy has occurred in patients with donorspecific HLA antibodies, over 5 or more years • Other pathology in recipients on tolerance protocols ○ Recurrent glomerular disease – Tolerance-inducing protocols have little or no effect on recurrent disease, such as C3 nephropathy, with possible exception of full chimerism (under investigation) ○ Engraftment syndrome – Occurs in first 1-2 weeks post transplant when chimerism is diminishing – Widespread damage of peritubular capillary endothelium – Sparse T cells in peritubular capillaries, Ki-67(+) ○ Chronic AMR – Develops subclinically over several years as smoldering process – Begins with C4d in peritubular capillaries – Later transplant glomerulopathy and graft dysfunction ○ Acute T-cell-mediated rejection – Rarely observed in these series – 1 case triggered by acute pyelonephritis – 1 case in HLA-identical recipient; able to discontinue immunosuppression later ○ Polyomavirus nephropathy – 1 series had 62% with polyoma infection (Seoul) ○ Donor disease – Arteriosclerosis ○ Interstitial fibrosis and tubular atrophy

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Renal biopsy essential in proving state of tolerance or absence of recurrent disease

SELECTED REFERENCES 1. 2.

3. 4.

5. 6. 7. 8.

9. 10.

11.

12.

13. 14.

15.

16.

17. 18.

19. 20. 21.

Chhabra AY et al: HSCT based approaches for tolerance induction in renal transplant. Transplantation. 101(11):2682-2690, 2017 Feng S et al: Five-year histological and serological follow-up of operationally tolerant pediatric liver transplant recipients enrolled in WISP-R. Hepatology. 65(2):647-660, 2017 Oura T et al: Chimerism-based tolerance in organ transplantation: preclinical and clinical studies. Clin Exp Immunol. 189(2):190-196, 2017 Sprangers B et al: Origin of enriched regulatory T cells in patients receiving combined kidney-bone marrow transplantation to induce transplantation tolerance. Am J Transplant. 17(8):2020-2032, 2017 Xie Y et al: Delayed donor bone marrow infusion induces liver transplant tolerance. Transplantation. 101(5):1056-1066, 2017 Zwang NA et al: Cell therapy in kidney transplantation: focus on regulatory T cells. J Am Soc Nephrol. 28(7):1960-1972, 2017 Chandran S et al: Current status of tolerance in kidney transplantation. Curr Opin Nephrol Hypertens. 25(6):591-601, 2016 Demetris AJ et al: 2016 comprehensive update of the Banff working group on liver allograft pathology: introduction of antibody-mediated rejection. Am J Transplant. 16(10):2816-2835, 2016 Hotta K et al: Induced regulatory T cells in allograft tolerance via transient mixed chimerism. JCI Insight. 1(10), 2016 Leventhal JR et al: Nonchimeric HLA-identical renal transplant tolerance: regulatory immunophenotypic/genomic biomarkers. Am J Transplant. 16(1):221-34, 2016 Massart A et al: The DESCARTES-Nantes survey of kidney transplant recipients displaying clinical operational tolerance identifies 35 new tolerant patients and 34 almost tolerant patients. Nephrol Dial Transplant. 31(6):1002-13, 2016 Taubert R et al: Hepatic infiltrates in operational tolerant patients after liver transplantation show enrichment of regulatory T cells before proinflammatory genes are downregulated. Am J Transplant. 16(4):1285-93, 2016 Ildstad ST et al: Facilitating cells: translation of hematopoietic chimerism to achieve clinical tolerance. Chimerism. 6(1-2):33-9, 2015 Leventhal JR et al: Immune reconstitution/immunocompetence in recipients of kidney plus hematopoietic stem/facilitating cell transplants. Transplantation. 99(2):288-98, 2015 Scandling JD et al: Chimerism, graft survival, and withdrawal of immunosuppressive drugs in HLA matched and mismatched patients after living donor kidney and hematopoietic cell transplantation. Am J Transplant. 15(3):695-704, 2015 Kawai T et al: Long-term results in recipients of combined HLA-mismatched kidney and bone marrow transplantation without maintenance immunosuppression. Am J Transplant. 14(7):1599-611, 2014 Sachs DH et al: Induction of tolerance through mixed chimerism. Cold Spring Harb Perspect Med. 4(1):a015529, 2014 Scandling JD et al: Tolerance and withdrawal of immunosuppressive drugs in patients given kidney and hematopoietic cell transplants. Am J Transplant. 2012 May;12(5):1133-45, 2012 Miyajima M et al: Early acceptance of renal allografts in mice is dependent on foxp3(+) cells. Am J Pathol. 178(4):1635-45, 2011 Newell KA et al: Identification of a B cell signature associated with renal transplant tolerance in humans. J Clin Invest. 120(6):1836-47, 2010 Kawai T et al: HLA-mismatched renal transplantation without maintenance immunosuppression. N Engl J Med. 358(4):353-61, 2008

Tolerance

Negative C4d in Tolerant Recipient (Left) This biopsy taken 783 days post transplant (> 1 year off immunosuppression) is normal without glomerular or vascular disease according to the mixed chimerism protocol (Massachusetts General Hospital). The majority of stable recipients show this pattern on biopsy. (Right) No C4d is detected in this biopsy from a patient on the mixed chimerism protocol (Massachusetts General Hospital). The majority of stable recipients show this pattern on biopsy.

Recurrent C3 Glomerulopathy in Tolerant Recipient

Kidney Transplantation

Normal Kidney in Tolerant Recipient

Subendothelial Deposits in Recurrent C3 Glomerulopathy (Left) Recurrent disease (C3 glomerulopathy) with C3 deposition is evident in this biopsy taken 8 years after transplantation in a recipient on the mixed chimerism protocol (Kawai et al). Recurrent glomerulonephritis does occur in tolerant recipients on mixed chimerism protocols (IgA nephropathy, lupus nephritis, membranous nephropathy). (Right) Duplication of the GBM and amorphous deposits are present in this case of recurrent C3 glomerulopathy (mixed chimerism protocol, Kawai et al).

Subclinical Antibody-Mediated Rejection

Subclinical Antibody-Mediated Rejection (Left) C4d deposition is present in this protocol biopsy from a clinically stable patient on the mixed chimerism protocol 2 years after transplantation (> 1 year off immunosuppression). DSA were detected, but renal function was normal. Light microscopy did not show evidence of rejection. (Right) While light microscopy was normal, electron microscopy reveal minimal duplication of the GBM associated with C4d and DSA in this stable patient in the mixed chimerism protocol 2 years post transplant.

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Kidney Transplantation

Acute Allograft Ischemia KEY FACTS

TERMINOLOGY • Delayed graft function (DGF): Transient dialysis needed • Primary nonfunction: Graft never functions

ETIOLOGY/PATHOGENESIS • Deceased cardiac death donors higher risk than heartbeating donation after brain death • Glomerular & other microvascular injury via complement & other mediators • Anastomotic site complications & ischemic causes such as poor preservation

• Intratubular cellular debris, sloughed cells, & neutrophils • Glomerular injury with variable intracapillary endothelial swelling, neutrophils, & fibrin thrombi • Interstitial edema & minimal inflammation ○ Interstitial fibrosis risk increased • Medullary vessels dilated and filled with leukocytes and erythroid precursors

ANCILLARY TESTS • Proliferation markers positive in tubular cells (PCNA, Ki-67 immunohistochemistry)

CLINICAL ISSUES

TOP DIFFERENTIAL DIAGNOSES

• 95-98% of grafts with DGF recover • Rejection risk increased • Drug toxicity (e.g., calcineurin inhibitors) can augment

• Calcineurin inhibitor toxicity • Acute cellular or antibody-mediated rejection • Acute obstruction

MICROSCOPIC

DIAGNOSTIC CHECKLIST

• Acute tubular injury with tubular epithelial cell flattening, nonisometric vacuolization, mitoses, & brush border loss

• Biopsy recommended to determine cause after 10 days of DGF

Acute Tubular Injury

Loss of Tubular Nuclei and Brush Border

Cell Debris in Tubules With Extreme Thinning

Ultrastructure of Ischemic Tubular Epithelium

(Left) Low-power view shows kidney with ischemic injury. Ischemia is manifested by epithelial injury in a hobnail pattern and dilated tubules ﬈ with a sparse infiltrate. (Right) In a deceased-donor renal allograft with oliguria on day 2, there is acute tubular injury with marked tubular cytoplasmic thinning, cellular loss, and debris with loss of tubular nuclei ſt.

(Left) Typical features of acute tubular injury (ATI) are eosinophilic cellular debris in tubules ſt, loss of nuclei st and brush borders, and interstitial edema ﬇. (Right) One tubular cell shows mitochondrial swelling and loss of cristae st. The cytoplasm has decreased density and the nucleus has early chromatin condensation, all features of a dying cell. The neighboring epithelial cell ﬈ is normal, indicating that this is not a processing artifact.

100

Acute Allograft Ischemia

Definitions • Ischemic graft injury in early posttransplant period ○ Delayed graft function (DGF): Transient dialysis in 1st week post transplant ○ Primary nonfunction (PNF) grafts never function

ETIOLOGY/PATHOGENESIS Acute Ischemic Injury • ↓ oxygen delivery/perfusion ○ ↑ risk with warm ischemia > 40 min or cold ischemia > 24 hours ○ May occur in donor before harvest or afterward • Dying cells release danger signals, activating toll-like receptors, leading to cell recruitment ○ NK and T cells produce mediators such as IFN-γ leading to upregulation of adhesion molecules • Complement activation ○ Colocalization of mannose binding lectins with complement in ischemia-reperfusion injury

Other Causes • Calcineurin inhibitor toxicity • Compromised vascular anastomosis, dissection or thrombosis

CLINICAL ISSUES Epidemiology • Incidence ○ DGF in 2-25% of deceased-donor grafts and PNF in 1-2%; varies with center ○ Asystolic donors have higher rate of DGF (19-84%) and PNF (4-18%) ○ ↑ risk from donor after cardiac death vs. donor after brain death donors

○ Neutrophils correlate with cold ischemia time and subsequent graft loss ○ Fibrin thrombi do not affect overall outcome • Tubules ○ Loss of brush border and nuclei, thinning, and dilation, particularly in proximal tubules – Nonisometric tubular cell vacuolization – Intratubular cellular debris and neutrophils – In severe cases, bare tubular basement membrane from desquamation of tubular epithelial cells (nonreplacement phenomenon) ○ Regeneration features (later): Basophilic cytoplasm, prominent nucleoli, mitosis • Interstitium ○ Edema and minimal inflammation with a few neutrophils and mononuclear cells ○ Donor interstitial fibrosis ↑ risk of DGF by 70% • Vessels ○ Medullary vessels dilated and filled with leukocytes and erythroid precursors ○ Arteries unaffected

ANCILLARY TESTS Immunohistochemistry • Proliferation markers positive in tubular cells (PCNA, Ki-67)

DIFFERENTIAL DIAGNOSIS Calcineurin Inhibitor Toxicity • Difficult to distinguish on histologic grounds alone • Classically, isometric vacuolization plus acute tubular injury (ATI) • Associated with high levels of calcineurin inhibitors

Acute Antibody-Mediated Rejection • Can present as ATI but C4d usually positive in PTC • Donor-specific antibody assays positive in > 90%

Treatment

Acute Cellular Rejection

• Transient dialysis • Reduce calcineurin inhibitors • Complement inhibitors and drugs under investigation

• Interstitial mononuclear inflammation, tubulitis, &/or endarteritis

Prognosis

• Collection ducts dilated • Tamm-Horsfall protein in lymphatics

• 95-98% of grafts with DGF recover ○ DGF typically lasts 10-15 days; < 2% last > 4 weeks • DGF has 50% ↑ risk of T-cell-mediated rejection (16% vs. 11%) or antibody-mediated rejection (10% vs. 7%) ○ 3.5x ↑ risk of graft loss in 1st year, but better than remaining on dialysis (not transplanted)

MACROSCOPIC General Features • Blotchy, mottled appearance with darker areas that are poorly perfused

Acute Obstruction

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Biopsy to determine cause after 10 days of DGF • Rejection risk on antithymocyte globulin low (4.8%)

SELECTED REFERENCES 1.

2.

MICROSCOPIC Histologic Features • Glomeruli variably affected ○ Endothelial swelling and vacuolization, capillary collapse

Kidney Transplantation

TERMINOLOGY

3.

Gill J et al: The risk of allograft failure and the survival benefit of kidney transplantation are complicated by delayed graft function. Kidney Int. 89(6):1331-6, 2016 Heilman RL et al: Progression of interstitial fibrosis during the first year after deceased donor kidney transplantation among patients with and without delayed graft function. Clin J Am Soc Nephrol. 11(12):2225-2232, 2016 Mourão TB et al: Predicting delayed kidney graft function with gene expression in preimplantation biopsies and first-day posttransplant blood. Hum Immunol. 77(4):353-7, 2016

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Kidney Transplantation

Acute Allograft Ischemia

Focal Interstitial Inflammation

Glomerular Thrombus

Hobnailed Tubular Epithelium

Delayed Graft Function

Focal Neutrophilic Infiltrate

Tubular Cell Proliferation

(Left) High-power view shows a focus of interstitial inflammation ſt amid tubules with varying degrees of injury. This is less than that seen in Tcell-mediated rejection. (Right) Trichrome stain shows fibrinous clot material ſt in a glomerulus, a finding that can be observed in acute allograft ischemia or other causes of thrombotic microangiopathy.

(Left) In this case of acute allograft ischemia, trichrome stain shows hobnailed tubular epithelial cells, a sign of intercellular detachment ﬊. (Right) Even at low power, injured epithelium with a hobnail pattern ſt and focal epithelial loss ﬈ can be appreciated in this case of acute allograft ischemia on trichrome stain. There is only a thin connective tissue framework between tubules, confirming the lack of appreciable fibrosis.

(Left) H&E section taken close to the renal pelvis demonstrates an inflammatory infiltrate, including neutrophils ſt, and adjacent injured tubular epithelium with nuclear loss ﬈. (Right) ATI in a renal allograft with delayed graft function (DGF) at day 5 shows frequent tubular nuclei that stain for Ki-67, a marker of proliferating cells ﬈. One study showed that, on average, ~ 8% of the tubular cells are positive, higher than in native kidney ATI (4%).

102

Acute Allograft Ischemia

Myoglobulin Casts (Left) Renal biopsy for DGF 2 days post transplant in a 2 year old who died from trauma is shown. Acute tubular injury is present with eosinophilic granular casts that stained for myoglobin; thus, this is renal failure due to rhabdomyolysis. (Right) Renal biopsy for DGF 2 days post transplant is shown. The 2year-old donor died from trauma. Many of the tubular granular casts ﬈ stain for myoglobin by immunohistochemistry; thus, the renal failure is due to rhabdomyolysis.

Acute Tubular Injury With Necrosis

Kidney Transplantation

Myoglobulin Casts

Negative C4d in ATI (Left) Transplant biopsy for DGF at day 10 is shown. Severe tubular necrosis st is present with neutrophils in the interstitium ﬈ & tubules, resembling acute antibodymediated rejection. However, C4d stain was negative, & the patient recovered graft function. (Right) Transplant biopsy for DGF at day 10 shows severe tubular necrosis with neutrophils, resembling acute antibody-mediated rejection. However, C4d is negative except for staining of sloughed necrotic cells in tubules ſt.

Acute Antibody-Mediated Rejection With ATI

ATI With Positive C4d Due to AMR (Left) This renal allograft reveals ATI with little inflammation, suggesting ischemic injury. However, C4d stain was positive in the peritubular capillaries, and the recipient had anti-donor HLA antibodies, meeting criteria for acute antibody-mediated rejection. (Right) C4d stain is positive in the peritubular capillaries in this day-9 biopsy for oliguria & ATI in a renal transplant patient who had anti-donor HLA antibodies, meeting the criteria of acute antibody-mediated rejection.

103

Kidney Transplantation

Urine Leak KEY FACTS

ETIOLOGY/PATHOGENESIS

IMAGING

• Dysfunctional ureterovesical anastomosis, ischemic injury, or rejection episodes • Laparoscopic techniques previously associated with higher complication rate but has declined as technique matured

• Radiology may demonstrate fluid leak (urinoma) • Renal scintigraphy tracer urinary extravasation • Antegrade pyelograph may allow leak localization

CLINICAL ISSUES • Urine leak complicates 3-5% of renal transplants • Presentation in postoperative period, usually within 4 months (84%), within 24 hours if due to technical error, within 2-3 weeks if ischemia ○ Hematuria, oliguria/renal dysfunction, and fistulas • Treatment: Percutaneous nephrostomy, dilatation/stent placement, surgical exploration/reanastomosis, endoscopic methods, special catheter placement • Prognosis: If corrected, does not typically impact 10-year patient or graft survival

MICROSCOPIC • Interstitial inflammation and tubular injury may be present ○ Inflammation in surrounding soft tissue • Edema in renal parenchyma and in soft tissue surrounding urine leak • Vascular thrombosis of periureteral vessels (80%) ○ Productive cytomegalovirus (15%) or BK virus (8%) infection

TOP DIFFERENTIAL DIAGNOSES • Lymphocele: Creatinine and potassium concentrations lower and sodium concentration higher

Urine Leak

Tubular Injury

Tubular Injury

Focal Inflammation

(Left) Ultrasound of a graft that developed a urine leak, taken 7 days post transplant, shows a collection of perinephric fluid ﬇ along one pole. The dilated pelvis ſt is a sign of obstruction. (Right) Low-power hematoxylin and eosin of a biopsy specimen from a patient with a urine leak shows irregular, dilated renal tubules ﬈ with mild tubular injury.

(Left) Medium power of a biopsy specimen from a patient with a urine leak shows focal nuclear loss ﬈ and tubular irregularities, both of which are findings of mild tubular injury. (Right) Mediumpower trichome shows a focal lymphoid infiltrate ﬈ amidst an edematous stroma and irregular tubules in a urine leak.

104

Urine Leak

Definitions • Urine collection due to numerous causes, often as anastomotic "leak" in setting of transplantation

ETIOLOGY/PATHOGENESIS

Laboratory Tests • Provided good renal excretory function, Cr concentration in leak fluid several times higher than serum Cr

Natural History

Dysfunctional Anastomosis

• Postoperative period: Majority in first 4 months (84%)

• May cause reflux/leakage • Surgical technical error may be cause ○ Misplacement of ureteral sutures (often at ureterovesical location) ○ Insufficient ureteral length ○ Ureter or renal pelvis laceration ○ Often evident within 24 hours

Treatment

Ureteral Ischemic Injury

Prognosis

• Risk of proximal ureter devascularization since renal pelvic vessels provide its blood supply • Ureter ischemic risk increases more distally from kidney • Renal allograft pelvic placement allows minimal length of transplant ureter ○ Special techniques: Psoas hitch, Boari flap, ureteroureterostomy, pyelovesicostomy, and ileal ureter • Lowest risk when "golden triangle" of perirenal fat bordered by ureter and lower renal pole preserved • Ischemic necrosis leaks usually occur 2-3 weeks after transplantation

• Does not typically impact 10-year patient or graft survival

Rejection Episodes

Histologic Features

• With transplant endarteritis, may involve ureteral graft vessels

• Limited data on renal histopathologic changes ○ Interstitial inflammation and tubular injury may be present – Inflammation in surrounding soft tissue ○ Edema in renal parenchyma and soft tissue surrounding urine leak • In study of 25 surgically removed, necrotic, ureteral segments ○ Vascular thrombosis of periureteral vessels (80%), productive cytomegalovirus (15%), or BK virus (8%)

Laparoscopic Technique • Living donor procurement initially associated with high incidence • Complication rate now almost as low as open donors

CLINICAL ISSUES Epidemiology • Incidence ○ 1-15% of renal transplant recipients have urologic complications – Urine leak complicates 3-5% of renal transplants ○ 1-3% of renal transplant recipients have ureteral leak

Site • Renal calyx, bladder, or ureter

Presentation • Oliguria ○ Sudden decrease in urinary output • Hematuria • Urinary fistula ○ May result from necrosis of distal ureter • Creatinine (Cr) and plasma urea increase ○ Due to solute resorption across peritoneum • Scrotal swelling • Wound drain ○ Fluid seepage with Cr several times current serum Cr

• • • • •

Kidney Transplantation

○ Difficult to determine if fluid production from preexisting seroma or lymph draining

TERMINOLOGY

Percutaneous nephrostomy Dilatation/stent placement Urgent surgical exploration/reanastomosis Endoscopic methods 3-way Foley catheter with irrigation that intermittently fills and empties bladder

IMAGING General Features • Ultrasound shows fluid collection (urinoma) but not its source • Renal scintigraphy tracer urinary extravasation • Antegrade pyelograph may allow leak localization

MICROSCOPIC

DIFFERENTIAL DIAGNOSIS Lymphocele • Cr and potassium concentrations lower and sodium concentration higher in lymphocele

SELECTED REFERENCES 1.

2. 3.

4. 5. 6.

Yashwant R et al: Leak from the surface of a decapsulated renal allograft: urine or lymph? excellent response to povidone iodine instillation. Saudi J Kidney Dis Transpl. 25(1):105-8, 2014 Hedegard W et al: Management of vascular and nonvascular complications after renal transplantation. Tech Vasc Interv Radiol. 12(4):240-62, 2009 Kobayashi K et al: Interventional radiologic management of renal transplant dysfunction: indications, limitations, and technical considerations. Radiographics. 27(4):1109-30, 2007 Karam G et al: Ureteral necrosis after kidney transplantation: risk factors and impact on graft and patient survival. Transplantation. 78(5):725-9, 2004 Streeter EH et al: The urological complications of renal transplantation: a series of 1535 patients. BJU Int. 90(7):627-34, 2002 Fontaine AB et al: Update on the use of percutaneous nephrostomy/balloon dilation for the treatment of renal transplant leak/obstruction. J Vasc Interv Radiol. 8(4):649-53, 1997

105

Kidney Transplantation

Lymphocele KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Collections of lymphatic fluid in perinephric space

• Typically, fibrous tissue with internal cystic formation and inflammatory cells in wall ○ Predominantly composed of mononuclear lymphoid cells • Renal biopsy may show changes of obstruction • Collecting duct dilation • Interstitial edema • Dilation of lymphatics, present normally along larger vessels

ETIOLOGY/PATHOGENESIS • Incomplete lymphatic anastomosis ○ Donor renal lymphatics at hilum fail to anastomose with recipient lymphatics ○ Normally, lymphatics spontaneously reconnect • Rejection episodes ○ Lead to increased vascular permeability, edema, lymph fluid, and promote lymphocele formation

CLINICAL ISSUES • Occurs in 2-8% of transplant recipients, 34% are clinically significant • Infection can lead to obstruction and infectious complications if large • Treated by fenestration

DIAGNOSTIC CHECKLIST • Consider lymphocele when no obvious change in biopsy accounts for renal transplant dysfunction ○ Particularly when obstructive features are seen ○ Microscopic features include chronic inflammation, edema, and dilated lymphatics

Lymphocele Fibrous Tissue Wall With Mononuclear Cells

Tubular Injury and Focal Interstitial Inflammation

Tubular Injury

Lymphocele Fibrous Tissue Wall With Focal Chronic Inflammation

(Left) Light microscopy of a lymphocele wall shows that it is composed of fibrous tissue with a focus of mononuclear chronic inflammatory cells ﬈. (Right) Kidney biopsy at low power shows a minimal focal interstitial inflammation ﬈ and tubular injury in a case of lymphocele.

(Left) The tubules show focal epithelial flattening and nuclear loss ﬈, consistent with a mild tubular injury in a case of lymphocele. (Right) Higher power examination of the lymphocele wall shows that the fibrous tissue lining has focal chronic inflammation ﬈.

106

Lymphocele

MACROSCOPIC

Definitions

General Features

• Collection of lymphatic fluid in nonepithelialized cavity in perinephric space, typically in postoperative field

• Perinephric nonsanguineous and nonpurulent fluid collection • Most occur adjacent to lower pole of kidney posterolateral to transplant ureter

ETIOLOGY/PATHOGENESIS Incomplete Lymphatic Anastomoses • Donor renal lymphatics at hilum fail to anastomose with recipient lymphatics • Normally, lymphatics spontaneously reconnect • Careful surgical ligation of lymphatic vessels is necessary to prevent lymphocele formation

Rejection Episodes • Lead to increased vascular permeability, edema, lymph fluid, and promote lymphocele formation

CLINICAL ISSUES Epidemiology • Incidence ○ Most occur within 6 weeks after transplantation ○ Occur in 1-26% of transplant recipients ○ Larger series indicated incidence is usually around 2-8% – Can occur 2-11 years post transplantation – 1/3 had rejection episodes – Another series revealed that some lymphoceles are subclinical ○ Obesity, age, rejection episodes, and use of mTOR inhibitors are risk factors

MICROSCOPIC Histologic Features • Fibrous tissue with internal cystic formation and inflammatory cells in wall ○ Inflammation predominantly composed of mononuclear lymphoid cells ○ No epithelial lining • Biopsy may show changes of obstruction ○ Collecting duct dilation ○ Interstitial edema ○ Dilation of lymphatics, present normally along larger vessels – Lymphatics are in kidney proper or in operative field ○ Mild interstitial mononuclear inflammation (borderline rejection pattern) • Microscopic pathologic descriptions are limited

DIFFERENTIAL DIAGNOSIS Abscess • High numbers of neutrophils are appreciated on aspirate samples • Cultures of aspirates useful

Presentation

Hematoma

• • • •

• Occur earlier, often in immediate postsurgical period

May be painful Urinary frequency may occur if bladder compression Lower extremity edema and deep venous thrombosis Infection can lead to obstruction and infectious complications, if large

Laboratory Tests • Creatinine level is similar to serum

Treatment • Surgical approaches ○ Laparoscopic or open fenestration, sometimes with marsupialization • Ultrasound-guided or CT-guided percutaneous drainage, sometimes combined with sclerotherapy

Urine Leak • Has higher creatinine and potassium concentration and lower sodium concentration

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Lymphocele should be considered when no obvious change in biopsy accounts for renal transplant dysfunction ○ Particularly when obstructive features are seen • May be mimicked by other perigraft fluid collections

SELECTED REFERENCES

Prognosis

1.

• 5-19% recur but does not affect graft survival

2.

IMAGING Ultrasonographic Findings • Most small and detected only on ultrasound • Some large and may be multilocular or multiple in number • Hydronephrosis with dilated calyces and hydroureter due to obstruction of ureter may be present

Kidney Transplantation

TERMINOLOGY

3. 4. 5.

6.

Bzoma B et al: Treatment of the lymphocele after kidney transplantation: a single-center experience. Transplant Proc. 48(5):1637-40, 2016 Kostro JZ et al: The use of tenckhoff catheters for draining of symptomatic lymphoceles: a review of literature and our experience. Transplant Proc. 47(2):384-7, 2015 Ranghino A et al: Lymphatic disorders after renal transplantation: new insights for an old complication. Clin Kidney J. 8(5):615-22, 2015 Giuliani S et al: Lymphocele after pediatric kidney transplantation: incidence and risk factors. Pediatr Transplant. 18(7):720-5, 2014 Khater N et al: Pseudorejection and true rejection after kidney transplantation: classification and clinical significance. Urol Int. 90(4):373-80, 2013 Lucewicz A et al: Management of primary symptomatic lymphocele after kidney transplantation: a systematic review. Transplantation. 92(6):663-73, 2011

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Kidney Transplantation

Renal Artery or Vein Thrombosis KEY FACTS

ETIOLOGY/PATHOGENESIS • • • •

Anastomotic problems Multiple arteries or arterial stenosis Hypercoagulability External compression by hematoma, lymphocele, or other lesions

• • • • • •

Loss of endothelium Platelet adhesion Congestion & sparse neutrophils in capillaries Recanalization if chronic Infarction of cortex, particularly renal artery thrombosis Dissection of media in RAT

CLINICAL ISSUES

TOP DIFFERENTIAL DIAGNOSES

• • • •

Macrohematuria Acute renal failure Prevalence highly variable by center Early treatment essential ○ Surgical correction or drugs • Arterial thrombosis usually occurs within 30 days of transplant • Renal vein thrombosis early or late

• Hyperacute & acute antibody-mediated rejection ○ C4d(+) in peritubular capillaries ○ Usually no large vessel thrombosis ○ Usually more neutrophils • Acute T-cell-mediated rejection ○ Endothelial mononuclear inflammation • Sickle cell trait • Recurrent atypical hemolytic uremic syndrome

MICROSCOPIC

DIAGNOSTIC CHECKLIST

• Thrombosis in vessel lumina

• C4d(-) in peritubular capillaries

Renal Vein Thrombosis

Renal Artery Stenosis

Chronic Renal Vein Thrombosis

Infarction in Renal Artery Stenosis

(Left) Thrombosis of the renal vein st 2 days post living donor transplant is shown. The cortex is congested. Arterial thrombosis developed hours post transplant. Despite an arterial thrombectomy, renal vein thrombosis ensued. (Right) An acute thrombus ﬉ is attached to the internal elastic lamina ſt of the renal artery with loss of endothelium. No inflammation is evident in contrast to acute humoral or cellular rejection.

(Left) Granulation tissue ſt is growing into a renal vein thrombosis, indicating that the thrombus is longstanding. (Right) The renal parenchyma is hemorrhagic and infarcted st in a case of renal artery stenosis. Notice the "ghosting" &/or total loss in appreciable tubules.

108

Renal Artery or Vein Thrombosis

IMAGING

Abbreviations

Radiographic Findings

• Renal vein thrombosis (RVT) • Renal artery thrombosis (RAT)

• Angiography demonstrates thrombosis & loss of cortical perfusion, usually wedge-shaped

ETIOLOGY/PATHOGENESIS Surgical Technical Problems • • • •

Intimal injury during procurement Difficult anastomosis or stenosis of anastomosis Trauma to vessels with dissection Narrow, twisted, or compressed renal vein

Hypercoagulable State • Antiphospholipid antibody syndrome • Nephrotic syndrome • Factor V Leiden mutation

Risk Factors • Multiple arteries ○ Especially if accessory artery supplies lower pole or ureter • Placement of graft on left side • En bloc pediatric donor • Renal artery stenosis can be risk factor for RAT • Compression of renal vein ↑ risk for RVT (e.g., by lymphocele, hematoma, or other lesion)

CLINICAL ISSUES Epidemiology • Incidence ○ Prevalence highly variable by center – Primary arterial thrombosis: 0.2-1.9% – Primary venous thrombosis: 0.1-3.4% ○ Arterial thrombosis usually occurs within 30 days of transplant ○ RVT early or late

Ultrasonographic Findings • Doppler ultrasound may demonstrate thrombus & absent flow • RVT ○ Diastolic reversal of flow in renal artery ○ Kidney may be enlarged with surrounding blood

MACROSCOPIC General Features • RVT: Engorged & purple • RAT: Pale, infarcted

MICROSCOPIC Histologic Features • Both arterial thrombosis & venous thrombosis display ○ Loss of endothelium ○ Adherence of platelets ○ Recanalization, if chronic • Arterial thrombosis in particular displays ○ Dissection of media ○ Infarction of cortex • Cortex ○ Peritubular & glomerular capillary neutrophils ○ Edema ○ Congestion

DIFFERENTIAL DIAGNOSIS Hyperacute & Acute Antibody-Mediated Rejection • C4d(+) in peritubular capillaries • Usually no large vessel thrombosis

Site

Recurrent Atypical Hemolytic Uremic Syndrome

• Hilar vessels ○ Intraparenchymal arteries & glomeruli spared • May extend into inferior vena cava

• Possible extrarenal manifestations of thrombotic microangiopathy

Presentation

• Endothelial mononuclear inflammation

• Macrohematuria • Proteinuria • Sudden anuria/oliguria ○ May result from either RVT or RAT • Pain in allograft • Graft swelling

Treatment • Surgical approaches ○ Thrombectomy or revision of anastomosis ○ Rapid recognition & treatment can be effective (~ 2 hr) • Drugs ○ Thrombolytics, anticoagulation, &/or aspirin

Prognosis • Usually causes graft loss • Early treatment essential

Kidney Transplantation

TERMINOLOGY

Acute T-Cell-Mediated Rejection

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • C4d(-) in peritubular capillaries

SELECTED REFERENCES 1.

2.

3. 4. 5.

Kawano PR et al: A case report of venous thrombosis after kidney transplantation - we can save the graft? time is the success factor. Int J Surg Case Rep. 36:82-85, 2017 Fallahzadeh MK et al: Acute transplant renal artery thrombosis due to distal renal artery stenosis: a case report and review of the literature. J Nephropathol. 3(3):105-8, 2014 Aktas S et al: Analysis of vascular complications after renal transplantation. Transplant Proc. 43(2):557-61, 2011 Ripert T et al: Preventing graft thrombosis after renal transplantation: a multicenter survey of clinical practice. Transplant Proc. 41(10):4193-6, 2009 Aschwanden M et al: Renal vein thrombosis after renal transplantation--early diagnosis by duplex sonography prevented fatal outcome. Nephrol Dial Transplant. 21(3):825-6, 2006

109

Kidney Transplantation

Transplant Renal Artery Stenosis KEY FACTS

TERMINOLOGY

MACROSCOPIC

• Posttransplant stenosis of major renal arteries, typically causing refractory hypertension due to increased renin production • Caused by atherosclerosis, intimal flap, kinking, and chronic rejection

• Stenosis usually occurs at or near anastomosis, such as renal-iliac artery anastomosis

ETIOLOGY/PATHOGENESIS • Surgical complication • Donor artery atherosclerosis • Chronic transplant arteriopathy

MICROSCOPIC • Cholesterol emboli may be present if stenosis due to atheroma • Tubular atrophy with little fibrosis is typical pattern • Acute tubular injury if acute or intermittent stenosis • May have prominent juxtaglomerular apparati (JGA)

TOP DIFFERENTIAL DIAGNOSES

CLINICAL ISSUES • Renal dysfunction/delayed graft function, bruit, hypertension • Clinically significant in 1-5% of recipients • Treatment ○ Percutaneous transluminal angioplasty ± stent ○ Surgical approaches, including anastomosis revision

• Calcineurin inhibitor toxicity • Obstruction

DIAGNOSTIC CHECKLIST • Histologic findings bland, subtle, and nonspecific • Diagnosis easily missed • Cholesterol emboli or prominent JGA are clues

Brush Border Loss in Tubular Injury

Renal Artery Stenosis on MRA

Tubular Casts and Injury

Cholesterol Embolus

(Left) A renal biopsy from a patient with intermittent renal artery stenosis 5 months after renal transplantation shows thinning of the tubules with PAS(+) brush border loss ﬈, typical of an acute tubular injury. (Right) MRA shows stenosis st of a donor renal artery at the site of anastomosis in an allograft due to atherosclerosis. A biopsy showed cholesterol emboli and mild tubular atrophy.

(Left) Periodic acid-Schiff stain of a transplant biopsy from a patient with intermittent renal artery stenosis shows a loss of brush borders, sparse tubular nuclei ﬉, and granular pigmented casts ﬊, indicative of an acute tubular injury. (Right) Periodic acidSchiff stain shows a cholesterol cleft ﬉ in an allograft with stenosis due to atherosclerotic disease. Cholesterol emboli provided a clue to the diagnosis.

110

Transplant Renal Artery Stenosis

IMAGING

Abbreviations

Ultrasonographic Findings

• Transplant renal artery stenosis (TRAS)

• Color flow duplex ultrasound reveals flow abnormalities ○ Stenosis may be incidental finding without hypertension or graft dysfunction ○ "Parvus tardus" waveform and decreased resistive index, pulsatility index, and acceleration index • CT, MR, and conventional angiography also useful

Definitions • Posttransplant stenosis of major renal arteries ○ Typically causing refractory hypertension due to increased renin production

ETIOLOGY/PATHOGENESIS

MACROSCOPIC

Surgical Complication

General Features

• Subintimal dissection or flap created at harvest or reanastomosis • Kinking or twisting of renal artery at transplantation • End-to-side anastomoses at higher risk than end-to-end anastomoses • Trauma may induce arterial intimal fibrosis

• Stenosis usually occurs at or near anastomosis, such as renal-iliac artery anastomosis • Uniform atrophy

Donor Artery Atherosclerosis • Can be complicated by atherosclerosis in recipient • Must be distinguished from de novo atherosclerosis and chronic transplant arteriopathy

Chronic Transplant Arteriopathy • Typically diffuse stenosis • Episodes of acute rejection more common in recipients with TRAS

CLINICAL ISSUES Epidemiology • Incidence ○ Clinically significant in 1-5% of recipients – Up to 23% if milder cases included ○ Most common vascular complication after kidney transplantation

Presentation • • • •

Delayed graft function Hypertension typically refractory to drugs Bruit over transplanted kidney Renal dysfunction ○ Progressive deterioration may be intermittent ○ Renin-angiotensin inhibitors worsen function

Natural History • Usually presents 3-24 months after transplantation

Treatment • Percutaneous transluminal angioplasty ± stent • Saphenous vein or recipient iliac artery grafts • Revision of anastomosis

MICROSCOPIC Histologic Features • Transplant kidney ○ Tubular atrophy with little fibrosis if stenosis chronic ○ Acute tubular injury may occur if stenosis intermittent ○ May have prominent juxtaglomerular apparati (JGA) ○ Cholesterol emboli present if stenosis due to atheroma ○ Normal glomeruli, arteries, and arterioles • Renal-iliac artery anastomosis ○ Rarely sampled for histology ○ Atherosclerosis ○ Medial dissection ○ Intimal flap ○ Intimal hyperplasia with inflammation (allograft arteriopathy)

DIFFERENTIAL DIAGNOSIS Calcineurin Inhibitor Toxicity • Arteriolar hyalinosis and isometric tubular vacuolization

Obstruction • Collecting duct dilation and Tamm-Horsfall protein leakage into interstitium • Dilated lymphatics

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Diagnosis easily missed • Hyperplasia of JGA is clue • Cholesterol emboli provide clue for atherosclerotic stenosis

SELECTED REFERENCES 1.

Prognosis • Outcome of angioplasty &/or stent generally good ○ Reduction of hypertension ○ Improvement of renal function ○ No increased graft loss • Percutaneous transluminal angioplasty restenosis rate 1060%

Kidney Transplantation

TERMINOLOGY

2.

3.

4.

Ali A et al: Long-term outcomes of transplant recipients referred for angiography for suspected transplant renal artery stenosis. Clin Transplant. 29(9):747-55, 2015 Aoyama H et al: Pathologic findings of renal biopsy were a helpful diagnostic clue of stenosis of the iliac segment proximal to the transplant renal artery: a case report. Transplant Proc. 46(2):651-3, 2014 Hurst FP et al: Incidence, predictors and outcomes of transplant renal artery stenosis after kidney transplantation: analysis of USRDS. Am J Nephrol. 30(5):459-67, 2009 Audard V et al: Risk factors and long-term outcome of transplant renal artery stenosis in adult recipients after treatment by percutaneous transluminal angioplasty. Am J Transplant. 6(1):95-9, 2006

111

Kidney Transplantation

Hyperacute Rejection KEY FACTS ○ Becomes swollen, hemorrhagic, and necrotic

TERMINOLOGY • Rejection immediately upon implantation and perfusion of graft

ETIOLOGY/PATHOGENESIS • Preexisting donor-reactive HLA or blood group antibodies at time of implantation

CLINICAL ISSUES • Graft primary nonfunction • Rare now due to pretransplant antibody testing ○ < 0.5% of transplants • Decreased incidence due to improved pretransplant testing for antibody • Becomes apparent hours to few days after graft implantation • No effective treatment currently

MICROSCOPIC • • • • • •

Resemble severe acute humoral rejection Platelet and neutrophil margination in capillaries Thrombi in glomeruli and arterioles Interstitial edema and hemorrhage Cortical necrosis in 12-24 hours Usually C4d-positive peritubular capillaries ○ May be negative

TOP DIFFERENTIAL DIAGNOSES • • • • •

Major vascular thrombosis (renal artery &/or vein) Perfusion nephropathy Donor thrombotic microangiopathy Recurrent atypical hemolytic uremic syndrome Sickle cell trait

MACROSCOPIC • Cyanosis of graft minutes to hours after perfusion

Hemorrhagic and Necrotic Kidney

Glomerular Capillaritis

Glomerular Thrombi

C4d

(Left) Nephrectomy specimen with hyperacute rejection shows edema, as indicated by the glistening cut surface, and hemorrhage. The dark zones at the corticomedullary junction are due to marked congestion ſt. The medullary areas are pale due to ischemia. (Right) Shown here are neutrophils ﬈ within glomerular capillaries within a few hours post implantation in hyperacute rejection. Capillaries are congested and some have lost endothelial nuclei ﬈. These are the 1st histological signs of hyperacute rejection.

(Left) H&E shows glomerular thrombi ﬈ in hyperacute rejection. The differential is between thrombotic microangiopathy, possibly donor disease, or preservation injury. C4d, as well as testing for antidonor antibodies, helps distinguish these possibilities. (Right) C4d immunohistochemistry in a wedge biopsy of hyperacute rejection shows strong staining of peritubular capillaries focally ﬈. Necrotic areas are C4d(-) ﬈. C4d can be negative in early biopsies of hyperacute rejection, probably due to poor perfusion.

112

Hyperacute Rejection

Definitions • Rapid rejection (minutes to hours) upon graft implantation

ETIOLOGY/PATHOGENESIS Antibody Mediated (Usual) • Preexisting antibodies to donor endothelium ○ Antidonor ABO blood group or HLA antibody (class I or class II) – Rare cases due to other or unidentified endothelial antigens ○ Antidonor antibody titers high enough to cause immediate rejection – Lower levels may delay onset as acute antibodymediated rejection (days) • Complement activation by antibody, endothelial activation, platelet activation • Rare cases without complement fixation

T Cell Mediated (Rare) • Primed cytotoxic T cells

Exogenous Antibody (Rare) • Rare cases associated with antithymocyte globulin or 3rd party plasma

CLINICAL ISSUES Epidemiology • Incidence ○ < 0.5% of transplants ○ Decreased incidence due to improved pretransplant testing for antibody against donor

Presentation • Graft primary nonfunction ○ Or within hours after graft implantation • Fever

Treatment • No effective treatment • Preventive therapy in ABO-incompatible or positive crossmatch transplants ○ Plasmapheresis to remove donor-specific antibody ○ Intravenous immunoglobulin ○ Rituximab (anti-CD20) ○ Anticomplement drug (experimental)

○ Platelet and neutrophil margination in glomerular or peritubular capillaries (PTC) ○ Scattered thrombi in glomeruli and arterioles • Later (12-24 hours) ○ Widespread thrombi in glomeruli and arteries ○ Fibrinoid necrosis of arteries – Larger arteries may be spared (e.g., HLA-DR antibodies) ○ Cortical and medullary necrosis • Features resemble severe acute antibody-mediated rejection

ANCILLARY TESTS Immunohistochemistry • C4d and CD61 (platelets) ○ PTC and glomeruli

Immunofluorescence • C4d-positive PTC ○ Negative or granular luminal PTC C4d staining does not exclude diagnosis of hyperacute rejection – Technical difficulties due to poor perfusion early and lack of viable tissue late – Possible C4d-negative antibody-mediated rejection – Possible T-cell-mediated rejection • IgG, IgM, &/or C3 may be present in capillaries ○ IgM most common in ABO-incompatible grafts

DIFFERENTIAL DIAGNOSIS Major Vascular Thrombosis (Renal Artery or Vein) • May be technical issues of anastomosis or due to hypercoagulable state ○ Thrombi due to technical problems often limited to larger vessels

Donor Thrombotic Microangiopathy • C4d negative

Perfusion Nephropathy • Thrombi and congestion within capillaries • C4d negative

Recurrent Atypical Hemolytic Uremic Syndrome • May have history of cardiomyopathy, malignant hypertension, or unusual pregnancy complications

SELECTED REFERENCES

Prognosis

1.

• Rapid graft loss

2.

MACROSCOPIC Gross Pathology • Cyanosis of graft minutes to hours after perfusion ○ Swollen, hemorrhagic with necrosis over 12-24 hours

3. 4.

5.

MICROSCOPIC Histologic Features • Early (1-12 hours)

Kidney Transplantation

TERMINOLOGY

6.

Pereira M et al: Hyperacute rejection in a kidney transplant with negative crossmatch: a case report. Transplant Proc. 48(7):2384-2386, 2016 Jackson AM et al: Multiple hyperacute rejections in the absence of detectable complement activation in a patient with endothelial cell reactive antibody. Am J Transplant. 12(6):1643-9, 2012 Kim L et al: Intragraft vascular occlusive sickle crisis with early renal allograft loss in occult sickle cell trait. Hum Pathol. 42(7):1027-33, 2011 Colovai AI et al: Acute and hyperacute humoral rejection in kidney allograft recipients treated with anti-human thymocyte antibodies. Hum Immunol. 66(5):501-12, 2005 Ahern AT et al: Hyperacute rejection of HLA-AB-identical renal allografts associated with B lymphocyte and endothelial reactive antibodies. Transplantation. 33(1):103-6, 1982 Kissmeyer-Nielsen F et al: Hyperacute rejection of kidney allografts, associated with pre-existing humoral antibodies against donor cells. Lancet. 2(7465):662-5, 1966

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Kidney Transplantation

Hyperacute Rejection

Postperfusion Biopsy

Focal C4d Deposition Postperfusion

Glomerular Capillaritis

Peritubular Capillaritis

C4d

CD61

(Left) This presensitized patient developed hyperacute rejection. Postperfusion biopsy shows neutrophils in peritubular st and glomerular ﬈ capillaries. The C4d stain was focally positive. (Right) C4d can be positive ﬉ on postperfusion biopsies, as shown in this biopsy from a patient with pretransplant donor-specific HLA antibodies. The neutrophils in the capillaries also show some staining.

(Left) Hyperacute rejection, postperfusion biopsy, shows a 3rd kidney in a patient who received a 6-antigen deceased donor match and was crossmatch negative. Neutrophils are prominent in glomeruli ﬈. (Right) H&E shows neutrophils within peritubular capillaries in hyperacute rejection ﬈. This biopsy was taken a few hours after implantation. The peritubular capillaries are markedly congested. Similar but milder congestion may be due to ischemia reperfusion injury and may be present at the time of implantation.

(Left) Postperfusion biopsy (hours after transplantation) in a patient shows patchy C4d staining in peritubular capillaries ﬈. Several capillaries are negative ſt. Immunofluorescence on frozen tissue was nondiagnostic. (Right) Postperfusion biopsy shows prominent CD61 staining in peritubular capillaries, indicating the presence of platelets ﬈. CD61 detects the platelet receptor for fibrinogen (IIb/IIIa) and may be a useful test to detect hyperacute rejection.

114

Hyperacute Rejection

Hyperacute Rejection in Day 1 Biopsy (Left) Loss of nuclei in proximal tubules ﬈ indicates early cortical necrosis. Interstitial hemorrhage due to peritubular capillary destruction is also evident st as well as glomerular thrombi ﬈. This graft was removed 3 days later. (Right) This biopsy one day after transplantation shows the classic features of hyperacute rejection: Interstitial hemorrhage ſt, glomerular thrombi st and neutrophils in peritubular ﬈ and glomerular capillaries, and focal tubular necrosis ﬉.

Hyperacute Rejection in Day 1 Biopsy

Kidney Transplantation

Early Cortical Necrosis in Day 1 Biopsy

Nephrectomy Specimen With Hyperacute Rejection (Left) C4d is focally positive in peritubular capillaries ﬈ in this biopsy taken 1 day after transplantation in a presensitized patient. Glomerular capillaries are filled with diffusely staining material corresponding to fibrin and cell debris. Some neutrophils also stain, which may be an artifact. (Right) This kidney is swollen, hemorrhagic, dusky, and has pale focal areas of necrosis ſt.

Cortical Necrosis

Diffuse Hemorrhage and Necrosis (Left) H&E shows a lowmagnification view of a renal allograft with hyperacute rejection. Cortical necrosis and hemorrhage are widespread ﬈. C4d stain was negative in the necrotic areas, comprising 95% of the sample. Nonnecrotic areas were selected from the paraffinembedded material for C4d staining ſt. (Right) This nephrectomy specimen from a patient with hyperacute rejection 3 days post transplant shows congestion and necrosis involving all elements of the kidney.

115

Kidney Transplantation

Acute T-Cell-Mediated Rejection KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Acute immunologic reaction to renal alloantigens mediated by T cells directed at MHC or non-MHC donor alloantigens

• Mixed mononuclear interstitial inflammation and tubulitis (type I TCMR) • Mononuclear cell accumulation under endothelium of arteries (type II TCMR) • Fibrinoid necrosis of arteries (type III TCMR) • Interstitial edema and sometimes hemorrhage • Glomerular involvement is usually mild • C4d negative in pure TCMR, but positive if antibodymediated component

ETIOLOGY/PATHOGENESIS • Alloreactive T cells against donor antigens expressed on donor cells or on recipient antigen presenting cells • Secondary participants, macrophages, granulocytes, chemokines, cytokines

CLINICAL ISSUES • Acute renal failure, oliguria • Incidence 5-10% in 1st year post transplant in conventional transplants • Acute T-cell mediated rejection (TCMR) type I and borderline cases usually responsive to pulse steroid therapy • Acute TCMR type II usually requires anti-T-cell agent • Important to know if antibody-mediated component

TOP DIFFERENTIAL DIAGNOSES • • • • •

Acute antibody-mediated rejection BK polyomavirus interstitial nephritis Pyelonephritis Acute allergic tubulointerstitial nephritis Posttransplant lymphoproliferative disorder

Acute TCMR, Type I

Tubulitis

Endothelialitis

T Cells in ACR

(Left) This biopsy, taken 3 weeks post transplant for a rising Cr (2.3), shows a patchy interstitial mononuclear infiltrate ﬈ and edema typical of acute T-cellmediated rejection (TCMR). Tubulitis was also present. The patchy nature of the infiltrate makes it important to take 2 cores for diagnostic sensitivity. (Right) Tubulitis ﬈ and interstitial mononuclear inflammation ﬊ are the defining features of type I acute cellular rejection (ACR). The infiltrating cells appear activated, and mitotic figures are present ﬉.

(Left) Endothelialitis (endarteritis) in a renal transplant biopsy, the defining feature of type II TCMR, is shown. Many mononuclear cells ﬈ are present in the intima, primarily T cells and monocytes. (Right) CD3 shows numerous infiltrating T cells, typical of TCMR. A C4d stain was also positive (not shown), indicative of an additional component of acute antibodymediated rejection (AMR).

116

Acute T-Cell-Mediated Rejection

Abbreviations • Acute T-cell-mediated rejection (TCMR)

Synonyms • Acute cellular rejection (ACR)

– ~ 50-80% of type III have evidence of AMR • "Pure" TCMR on indication biopsy ○ TCMR or borderline/suspicious without evidence of AMR (negative DSA and C4d): 28% or 30%, respectively, have doubling of Cr in 2 years

MICROSCOPIC

Definitions

Histologic Features

• Acute immunologic reaction to renal alloantigens mediated by T cells ○ Type I: Tubulointerstitial ○ Type II: Endarteritis ○ Type III: Fibrinoid arterial necrosis or transmural inflammation of arteries

• Glomeruli ○ Usually spared ○ Occasional cases show glomerulitis with mononuclear cells in glomerular capillaries – Glomerulitis more commonly found in AMR, where macrophages predominate – Not currently included as criterion for TCMR ○ Acute allograft glomerulopathy in < 5% – Markedly swollen endothelial cells occluding capillary lumen – Mesangiolysis with PAS(+) webs – When present, usually associated with type II TCMR • Interstitium ○ Mononuclear cell inflammation in interstitium – Diagnosis of rejection by Banff criteria requires > 25% of nonscarred cortex to have mononuclear infiltrate – Lesser degrees of inflammation considered suspicious or borderline for rejection – Cells mostly CD4(+) and CD8(+) T cells and CD68(+) macrophages – Dendritic cells [DC-SIGN(+) cells present from recipient and donor (latter dominant in 1st few months)] □ ↑ DC-SIGN(+) cells correlate with ↓ graft survival ○ Eosinophils, plasma cells, and few neutrophils may also be present in infiltrate – Plasma cell-rich rejection has worse prognosis – Some studies show that eosinophils portend worse outcome ○ Interstitial edema ○ Hemorrhage in more severe cases • Tubules ○ T cells and macrophages within tubule (tubulitis) – Only nonatrophic tubules evaluated by Banff criteria ○ Tubular cell injury (loss of brush border, apoptosis) ○ Tubular basement membranes sometimes rupture with severe tubulitis – May form granulomas • Arteries ○ Mononuclear inflammatory cells beneath endothelium in arteries (endarteritis or endothelialitis) – CD3(+) T cells and CD68(+) monocytes/macrophages – Focal process, affecting ~ 25% of cross sections of arteries and ~ 12% of arterioles – Larger vessels affected more than arterioles – Endothelialitis in arterioles sometimes seen in conjunction with endothelialitis in arteries; has same significance ○ Marginated mononuclear cells along endothelial surface – Does not count for endarteritis, but associated with it ○ Venulitis found in some cases of TCMR but not prognostically significant

ETIOLOGY/PATHOGENESIS T-Cell-Mediated Rejection • Alloreactive T cells against donor antigens ○ MHC (HLA) or non-MHC • Target varies and includes capillary and arterial endothelium, tubules, and glomeruli • Ongoing tubulointerstitial inflammation commonly present in follow-up biopsies 1-2 months after TCMR episode ○ Argues that while TCMR may be of acute onset, inflammation may persist and give chronic cellular rejection phenotype

CLINICAL ISSUES Epidemiology • Incidence ○ 5-10% in 1st year post transplant in conventional transplants – Type I: ~ 65% of TCMR cases – Type II: ~ 30% – Type III: < 5%

Presentation • • • •

Acute renal failure Oliguria or decreased urine output Graft tenderness (severe cases) May also be seen on protocol (surveillance) biopsies with normal renal function

Treatment • Drugs ○ Type I and cases borderline/suspicious for TCMR are usually responsive to pulse steroid therapy ○ Type II usually resistant to pulse steroid therapy; additional treatment may consist of anti-T-cell agent ○ Type III resistant to current therapies

Prognosis • 1-year graft survival ○ Type I without evidence of antibody-mediated rejection (AMR) [C4d(-), DSA absent]: 95-100% ○ Type I with evidence of AMR [C4d(+), DSA present]: ~ 75% ○ Type II without evidence of AMR: 67-100% ○ Type II with evidence of AMR: 63-92% ○ Type III: 20-32% overall, ± AMR

Kidney Transplantation

TERMINOLOGY

117

Kidney Transplantation

Acute T-Cell-Mediated Rejection ○ Endothelium may develop signs of "activation" with basophilic cytoplasm, enlarged active nuclei ○ Transmural inflammation in more severe cases – Fibrinoid necrosis also occasionally seen in severe cases but more commonly associated with AMR

ANCILLARY TESTS Immunofluorescence

• C4d deposition in peritubular capillaries, circulating antidonor antibodies • May be superimposed on acute TCMR ("mixed rejection")

Pyelonephritis • Neutrophilic casts and abscesses • Positive urine culture for bacterial infection

• Generally little or no immunoglobulin or C3 deposition in glomeruli or interstitium ○ Fibrin diffusely present in edematous interstitium • C4d(-) in pure TCMR • TCMR cases with C4d deposition have superimposed acute &/or chronic AMR

Thrombotic Microangiopathy

Electron Microscopy

• May represent allergic drug reaction • Difficult or impossible to distinguish from TCMR unless endarteritis present

• Not generally needed for diagnosis • Glomeruli in acute allograft glomerulopathy show marked endothelial reaction (swollen cytoplasm, loss of fenestrations) • Rare cases of minimal change-like disease, with widespread podocyte foot process effacement and nephrotic syndrome, described in association with TCMR

Molecular Tests • mRNA panels proposed for diagnosis of TCMR (Halloran) ○ Strong signals for interferon-γ inducible and cytotoxic Tcell genes (e.g, CXCL9, CXCL11, GBP1, INDO) • 2 recipient single nucleotide polymorphisms (SNP) associated with 1.6-2.0 ↑ risk of TCMR in 1st year ○ PTPRO (tyrosine kinase) and DEUP1 (cilium)

DIFFERENTIAL DIAGNOSIS Borderline/Suspicious for TCMR • Threshold for TCMR related to intensity of tubulitis (> 4 cells/tubular cross section) and requires > 25% infiltrate in nonscarred areas of cortex • Probably should be treated as TCMR if associated with acute graft dysfunction

Isolated Endothelialitis • Variant of TCMR • Recent study shows that endothelialitis with mild or no interstitial inflammation/tubulitis behaves as TCMR • May be associated with AMR, which worsens prognosis

BK Polyomavirus Interstitial Nephritis • Plasma cells in infiltrate • Intranuclear inclusions in tubular epithelial cells • Inflammation primarily in sites with viral infection (IHC)

Resolving/Partially Treated TCMR • Disproportionately more severe tubulitis compared to interstitial inflammation • Inflammation in areas of interstitial fibrosis

Peritubular Capillaritis • Common finding on protocol biopsy in patients with donorspecific anti-HLA antibody ○ C4d usually negative (~ 90% of cases) • Associated with later development of transplant glomerulopathy 118

Acute Antibody-Mediated Rejection

• Endothelialitis and luminal fibrin may be present • Fragmented erythrocytes in intima, thrombi with minimal endothelial inflammation, and mucoid intimal thickening favor this diagnosis

Acute Allergic Tubulointerstitial Nephritis

Obstruction • Edema and collecting duct dilation may be present • Usually no more than borderline infiltrates (< 25% of cortex) • Dilated lymphatics ± Tamm-Horsfall protein • Can never rule out by morphology

Posttransplant Lymphoproliferative Disorder • Little edema and predominance of atypical B cells rather than T cells ○ Monoclonal B-cell infiltrate most common • Most cases EBER positive

Atheromatous Embolism • Endothelialitis and luminal fibrin may be present • Thorough examination of tissue levels should reveal cholesterol clefts in arteries with inflammation

Acute Transient Arteriopathy • Rare lesion (< 1%); occurs in very early posttransplant period (< 2 weeks) in deceased donor transplants • Markedly reactive endothelial cells ○ Endothelial cell swelling, lifting, and vacuolization ○ No endothelialitis or thrombi, C4d(-) • Usually occurs with acute tubular injury • Lesion resolves on follow-up biopsy

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Multiple levels need to be examined to detect focal lesions, such as endarteritis, which are more specific for TCMR • 1 biopsy core has false-negative rate of ~ 10% • Biopsies may meet criteria for TCMR and AMR, in which case outcome is dominated by latter

SELECTED REFERENCES 1.

2. 3.

Becker JU et al: Banff borderline changes suspicious for acute t cellmediated rejection: where do we stand? Am J Transplant. 16(9):2654-60, 2016 Ghisdal L et al: Genome-wide association study of acute renal graft rejection. Am J Transplant. 17(1):201-209, 2016 Halloran PF et al: Molecular assessment of disease states in kidney transplant biopsy samples. Nat Rev Nephrol. 12(9):534-48, 2016

Acute T-Cell-Mediated Rejection

Category and Criteria

Comments

Borderline/Suspicious for TCMR Foci of tubulitis (t1, t2, or t3) with minor interstitial infiltration (i0 or i1) or interstitial infiltration (i2, i3) with mild (t1) tubulitis

Poor interobserver reproducibility; molecular studies do not distinguish from TCMR; criteria have changed since Banff 1997 (e.g., i0 vs. i1); survey of publications revealed that most use i1t1 (Banff 1997) and include borderline/suspicious in TCMR category for outcome studies.; according to Banff 2015, "retaining the i1 threshold for borderline is permitted, although this must be made transparent in reports and publications"†

Kidney Transplantation

Banff Criteria for Acute T-Cell-Mediated Rejection

Acute TCMR With Tubulointerstitial Inflammation (Type I) Type IA: Interstitial inflammation in > 25% of unscarred cortex (i2 or i3) with foci of moderate tubulitis (4-10 cells/tubular cross section; t2)

Absence of endarteritis

Type IB: Interstitial inflammation in > 25% of unscarred cortex (i2 or i3) with foci of severe tubulitis (> 10 cells/tubular cross section; t3)

Absence of endarteritis

Acute TCMR With Intimal Arteritis (Type II) Type IIA: Presence of intimal arteritis (a.k.a. endarteritis or endothelialitis) involving ≤ 25% of luminal area (v1)

Interstitial inflammation and tubulitis not required; can be manifestation of pure TCMR or mixed with antibody-mediated rejection; C4d(+) or DSA associated with worse prognosis

Type IIB: Presence of intimal arteritis involving > 25% of luminal area (v2)

Interstitial inflammation and tubulitis not required; can be manifestation of pure TCMR or mixed with antibody-mediated rejection; C4d(+) or DSA associated with worse prognosis

Acute TCMR With Transmural Arterial Inflammation (Type III) Type III: Transmural arteritis &/or arterial fibrinoid change and necrosis of medial smooth muscle cells with accompanying lymphocytic inflammation

Often manifestation of antibody-mediated rejection

Tubulitis (t) is scored by mononuclear cells/tubular cross section (t0 = 0, t1 =1-4, t2 = 5-10, t3 = > 10). Interstitial inflammation (i) is scored by % of nonscarred cortex with mononuclear infiltration (i0 < 10%, i1 = 10-25%, i2 > 25-50%, i3 > 50%). Vascular lesions (v) are scored by extent of lumen occupied by subendothelial mononuclear cells in arteries ( v1 ≤ 25%, v2 > 25%) or by the presence of transmural inflammation or fibrinoid necrosis (v3). All categories require C4d stains to be negative for pure TCMR. † A Banff Working Group on TCMR is addressing these issues. Loupy A et al: The Banff 2015 Kidney meeting report: current challenges in rejection classification and prospects for adopting molecular pathology. Am J Transplant. ePub, 2016; Becker JU et al: Banff borderline changes suspicious for acute T cell-mediated rejection: where do we stand? Am J Transplant 16: 2654-2660, 2016. 4.

5.

6.

7. 8.

9. 10.

11.

12. 13.

14. 15.

Lamarche C et al: Efficacy of acute cellular rejection treatment according to Banff score in kidney transplant recipients: a systematic review. Transplant Direct. 2(12):e115, 2016 Loupy A et al: The Banff 2015 kidney meeting report: Current challenges in rejection classification and prospects for adopting molecular pathology. Am J Transplant. 17(1):28-41, 2016 Reeve J et al: Using molecular phenotyping to guide improvements in the histologic diagnosis of T cell-mediated rejection. Am J Transplant. 16(4):1183-92, 2016 Teo RZ et al: Cell-mediated and humoral acute vascular rejection and graft loss: a registry study. Nephrology (Carlton). 21(2):147-55, 2016 Zeng G et al: Antigen-specificity of T cell infiltrates in biopsies with T cellmediated rejection and BK polyomavirus viremia: analysis by next generation sequencing. Am J Transplant. 16(11):3131-3138, 2016 Zhao X et al: Rejection of the renal allograft in the absence of demonstrable antibody and complement. Transplantation. 101(2):395-401, 2016 Batal I et al: Dendritic cells in kidney transplant biopsy samples are associated with T cell infiltration and poor allograft survival. J Am Soc Nephrol. 26(12):3102-13, 2015 Randhawa P: T-cell-mediated rejection of the kidney in the era of donorspecific antibodies: diagnostic challenges and clinical significance. Curr Opin Organ Transplant. 20(3):325-32, 2015 Naesens M et al: The histology of kidney transplant failure: a long-term follow-up study. Transplantation. 98(4):427-35, 2014 Nongnuch A et al: Early posttransplant nephrotic range proteinuria as a presenting feature of minimal change disease and acute T cell-mediated rejection. Transplant Proc. 46(1):290-4, 2014 Sis B et al: Isolated endarteritis and kidney transplant survival: a multicenter collaborative study. J Am Soc Nephrol. ePub, 2014 El Ters M et al: Kidney allograft survival after acute rejection, the value of follow-up biopsies. Am J Transplant. 13(9):2334-41, 2013

16. Williams WW et al: Clinical role of the renal transplant biopsy. Nat Rev Nephrol. 8(2):110-21, 2012

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Acute T-Cell-Mediated Rejection

Tubulitis

Destructive Tubulitis

ACR With Increased Eosinophils

Severe Tubulitis

Tubulitis With Basement Membrane Rupture

Granulomatous Reaction

(Left) Mild tubulitis (t1 lesion) in TCMR type IA is shown. A mononuclear inflammatory cell ﬊ can be recognized by its dark nucleus and surrounding halo. Interstitial inflammation is also present ﬈. (Right) Focally severe tubulitis ﬈ in tubules that are nonatrophic or no more than mildly atrophic are borderline/suspicious for TCMR.

(Left) Numerous eosinophils ﬈ are seen in this case of TCMR. This finding can be part of TCMR and does not necessarily indicate an allergic drug reaction in the transplant. (Right) Severe tubulitis (t3 lesion) in a case of TCMR type IB shows numerous mononuclear cells ﬊ within the tubule. The tubular epithelium is displaced from the tubular basement membrane. This is not uncommonly seen in grafts after withdrawal of immunosuppressive drugs.

(Left) In tubulitis, tubular basement membrane rupture ﬈ is associated with a poorer outcome. A granulomatous response to rupture may develop and should not be confused with an infection. (Right) TCMR may have a prominent granulomatous response to ruptured tubules. This may be confused with an infectious process, particularly adenovirus infection. Remnants of the tubular basement membrane can be seen st.

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Acute T-Cell-Mediated Rejection

Borderline/Suspicious TCMR (Left) 1-year posttransplant protocol biopsy shows focal mild interstitial inflammation ﬊. There was focal severe tubulitis, indicative of a borderline TCMR. No endothelialitis was present. (Right) 1-year posttransplant protocol biopsy shows focal severe tubulitis ﬈ (Banff t3) with tubular basement membrane disruption. There was only mild interstitial inflammation (~ 5%) in the biopsy. The presence of severe tubulitis qualifies the biopsy for the borderline/suspicious category of TCMR.

Unappreciated T-Cell Infiltrate

Kidney Transplantation

Borderline TCMR

Plasma Cell-Rich Acute TCMR and AMR (Left) At left, minimal infiltrate is shown, which would be considered borderline/suspicious at most; however, CD3 stain of the same area (at right) shows extensive T-cell infiltrate in the cortex, but mostly as individual cells that are not readily appreciated on H&E. (Right) A combination of TCMR and acute AMR is shown. Numerous plasma cells within the interstitial infiltrate are present in this variant of acute rejection. Most cases of plasma cell-rich acute rejection are C4d(+).

ACR With Plasma Cells

Plasma Cell Tubulitis (Left) In this example of TCMR, type I, there are increased numbers of plasma cells ﬈ in the interstitial infiltrate. A stain for BK polyoma virus (not shown) was negative. (Right) A lesion of plasma cell tubulitis ﬈ is seen in this example of TCMR, type I with increased interstitial plasma cells. While plasma cell tubulitis is most commonly seen in BK polyoma virus infection, it may occasionally be seen in other entities.

121

Kidney Transplantation

Acute T-Cell-Mediated Rejection

Ongoing TCMR

Ongoing TCMR

Ongoing TCMR

Persistent TCMR

Treated TCMR

Residual Tubulitis and Edema After Treatment

(Left) Follow-up biopsy 6 weeks post TCMR shows early diffuse interstitial fibrosis and tubular atrophy ﬊, as well as interstitial inflammation. (Right) Follow-up biopsy 6 weeks post TCMR shows fine interstitial fibrosis, partial tubular atrophy with thickened tubular basement membranes ﬈, and tubulitis ﬈ in partially atrophic tubules. The patient's creatinine had been persistently elevated at ~ 2.3 mg/dL since the previous biopsy.

(Left) 6 weeks following a biopsy that showed TCMR, trichrome stain highlights the increased interstitial fibrosis. Increased interstitial inflammation ﬈ is present as well. (Right) Silver stain highlights thickened tubular basement membranes; mononuclear cell tubulitis ﬈ is apparent.

(Left) Biopsy of a patient 8 days after being treated with antithymocyte globulin and steroids for TCMR, Banff, type IB shows persistent tubulitis ﬉ with somewhat lesser interstitial inflammation, typical of a treated rejection. (Right) Repeat biopsy from a patient 8 days after being treated with antithymocyte globulin and steroids for TCMR, Banff, type IB shows severe tubulitis ﬉ with somewhat lesser interstitial inflammation. Note interstitial edema ﬈.

122

Acute T-Cell-Mediated Rejection Transmural Arterial Inflammation (TCMR, Type III) (Left) Endothelialitis ﬈ is present on this 4-month posttransplant biopsy. The patient's creatinine was elevated to 1.7 mg/dL (baseline of 1.3-1.7 mg/dL). No significant interstitial inflammation or tubulitis was present. (Right) Transmural inflammation ﬊ is shown in a small artery in an allograft 10 years post transplant. Immunosuppression was reduced 4 months previously due to T-cell PTLD. C4d staining (not shown) was negative.

Endothelialitis

Kidney Transplantation

Isolated Endothelialitis

Fibrinoid Necrosis (Left) An apoptotic cell ﬊ is seen in an artery with endothelialitis. (Right) Fibrinoid necrosis ﬈ is seen in a small artery in an allograft 10 years post transplant. Immunosuppression was reduced 4 months previously due to T-cell PTLD. While fibrinoid necrosis is more commonly associated with humoral rejection, this biopsy was C4d(-).

Endarteritis in HLA Identical Sibling Graft

Endothelialitis (Left) The presence of florid endarteritis st in this HLAidentical sibling graft is evidence that non-MHC antigens can serve as a target of the TCMR process in arteries. Rare cases of HLA identical grafts have been lost to type II TCMR. (Right) Endothelialitis shows CD3(+) T cells ﬈. The majority of the T cells in these lesions are CD8(+).

123

Kidney Transplantation

Acute T-Cell-Mediated Rejection

Cytotoxic T Cells in Endarteritis

Endothelialitis and Arteriosclerosis

v-Only Lesion Followed by Type I TCMR

Incidental v-Only Lesion

Resolving TCMR, Type II

TCMR, Type III

(Left) The cells infiltrating the intima stain for a cytotoxic granule component (TIA-1) ﬉. Studies have shown the predominant cells in these lesions are CD8(+) T cells. (Right) Endothelialitis ﬊ along with inflammation in a thickened arterial intima ﬊ due to donor arteriosclerosis is shown. This may be confused with chronic transplant arteriopathy.

(Left) On the left is an indication biopsy that has endarteritis st. No other lesion was present (no infiltrate). The patient was not treated for rejection. A biopsy 3 days later showed a florid infiltrate (Banff, type IB), shown on the right, suggesting sampling error. (Right) This fragment of a large artery has endarteritis, appreciated in retrospect. The patient was not treated for rejection, and the Cr was stable 1 year later. This suggests that v lesions only can be clinically silent.

(Left) Resolving ACR, type II shows a few inflammatory cells within a "loose" intima ﬈, likely representing early transplant arteriopathy. A biopsy from this patient 2 months earlier showed endothelialitis. (Right) Type III ACR with transmural inflammation of an arcuatesized artery is shown. Inflammatory cells can be seen in the media as round, dark nuclei ﬈.

124

Acute T-Cell-Mediated Rejection

Antibody-Mediated Rejection (Left) PAS shows thrombotic microangiopathy in a native kidney due to factor H deficiency. The intimal inflammation ﬈ resembles endarteritis due to rejection. Endothelial cells are markedly reactive. (Right) Fibrinoid necrosis in a small artery ﬈ is shown. Loss of smooth muscle nuclei is present focally. This patient had acute AMR, which accounts for the majority of cases with type III arterial lesions. When C4d is positive, this is considered acute humoral rejection.

Acute Transient Arteriopathy

Kidney Transplantation

Thrombotic Microangiopathy

Inflammation in Renal Vein Thrombosis (Left) This arterial lesion, which occurs in the very early (< 2 weeks) posttransplant period, may resemble endothelialitis. Endothelial cells are enlarged and show areas of vacuolization ﬈ without endothelialitis. (Right) Arterial inflammation resembles endarteritis in a renal transplant with acute renal vein thrombosis and cortical necrosis; this is not a lesion of rejection.

Infiltrating Mononuclear Cell

Acute Cellular Rejection (Left) Tubulitis with an infiltrating mononuclear cell ﬇ is seen by electron microscopy. The loss of cytoplasmic density indicates injury of the adjacent tubular epithelium ﬈. (Right) ACR can affect glomeruli, as shown in this capillary loop with pronounced endothelial reaction and loss of fenestrae ﬈. Podocytes also show segmental foot process effacement ﬈, which rarely can be extreme, resembling minimal change disease.

125

Kidney Transplantation

Acute T-Cell-Mediated Rejection

Interstitial Inflammation

Combined TCMR and Polyomavirus Nephropathy

TCMR, IB With Focal BK Infection

TCMR, IB With Focal BK Infection

TCMR, IB With Focal BK Infection

TCMR, IB With Focal BK Infection

(Left) Both TCMR and BK polyomavirus infection are manifested by interstitial inflammatory cell infiltrate within the cortex. (Right) On higher magnification of TCMR and BK polyomavirus infection, mononuclear cell tubulitis is apparent ﬈. No viral inclusions are identified in this area. Of note, this patient had low-level BK viremia.

(Left) Kidney biopsy from a patient 4 months post transplant with slightly elevated serum creatinine and BK viremia shows diffuse interstitial inflammation ﬈ and tubulitis. Many of the T cells in BK nephritis are donor alloreactive (Zeng). (Right) Kidney biopsy from a patient 4 months post transplant with slightly elevated serum creatinine and BK viremia shows severe tubulitis ﬈.

(Left) Kidney biopsy from a patient 4 months post transplant with slightly elevated serum creatinine and BK viremia shows severe tubulitis with basement membrane disruption ﬉ (Banff t3). (Right) BK in situ hybridization shows rare BK(+) epithelial cell nuclei ﬊. The presence of diffuse interstitial inflammation, physically separated from the BK(+) cells, allows one to make the diagnosis of both TCMR and focal BK infection.

126

Acute T-Cell-Mediated Rejection

Plasma Cells (Left) Malakoplakia is seen in a transplant biopsy, where there is a diffuse infiltrate of large macrophages ﬈, along with lymphocytes and plasma cells ﬊. (Right) Transplant biopsy shows numerous plasma cells in the interstitium ﬊ and plasma cell tubulitis ﬈, features that raise the possibility of BK polyoma virus infection. The patient had BK viremia, but in situ hybridization was negative for BK polyoma virus. A focal BK infection cannot be excluded.

Posttransplant Lymphoproliferative Disorder

Kidney Transplantation

Malakoplakia

Posttransplant Lymphoproliferative Disorder (Left) Posttransplant lymphoproliferative disorder (PTLD) is in the differential diagnosis of TCMR. This biopsy shows a dense lymphoid infiltrate. (Right) The infiltrating cells of PTLD are atypical, including some large cells. Immunostaining revealed atypical CD20(+) B cells; the patient had a posttransplant marginal zone lymphoma. This case also showed tubulitis with CD3(+) T cells, and so a component of TCMR may have been present. PTLD treatment usually includes decreased immunosuppression.

Acute Allograft Glomerulopathy

Reactive Endothelial Cells in Arteriole (Left) Acute allograft glomerulopathy shows glomerulitis and marked swelling of endothelial cells ﬈, resembling endocapillary hypercellularity of proliferative lupus nephritis. This glomerular finding can be a lesion of cellular rejection and is nearly always associated with endothelialitis. C4d was negative. (Right) This arteriole shows markedly enlarged endothelial cells ﬈. Elsewhere, this biopsy showed acute allograft glomerulopathy and endothelialitis in arteries.

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Kidney Transplantation

Chronic T-Cell-Mediated Rejection KEY FACTS

TERMINOLOGY • Persistent or recurrent T-cell-mediated rejection leading to chronic changes in allograft, including transplant arteriopathy, interstitial fibrosis, and tubular atrophy

ETIOLOGY/PATHOGENESIS • T-cell-mediated injury due to recognition of alloantigens on parenchyma or vessels

○ Mononuclear infiltrate and tubulitis ○ Inflammation in areas of fibrosis • Arteries ○ Intimal fibrosis – Little or no fibroelastosis ○ Mononuclear cells in intima

ANCILLARY TESTS

CLINICAL ISSUES

• Negative stain for C4d in peritubular capillaries

• Presents as chronic renal failure, often with proteinuria and hypertension • May be asymptomatic or seen on protocol biopsy • Interstitial fibrosis with inflammation (i-IFTA) shortens graft survival more than either alone • Presence of chronic transplant arteriopathy also shortens graft survival

TOP DIFFERENTIAL DIAGNOSES

MICROSCOPIC • Tubules and interstitium

• • • • •

Chronic antibody-mediated rejection Chronic calcineurin inhibitor toxicity Late stage of BK polyoma virus nephropathy Hypertensive arteriosclerosis Chronic pyelonephritis

DIAGNOSTIC CHECKLIST • Inflammation in areas of fibrosis correlates with progressive graft injury

Inflammation in Fibrotic Areas

Tubulitis

Transplant Arteriopathy and Endothelialitis

Transplant Arteriopathy

(Left) A late graft biopsy with chronic T-cell-mediated rejection (TCMR) shows a diffuse infiltrate of mononuclear cells in areas with interstitial fibrosis, a combination that has a poor prognosis. (Right) Tubulitis ſt in atrophic tubules in allografts with late dysfunction is often associated with tubulitis in nonatrophic tubules.

(Left) This biopsy shows transplant arteriopathy with superimposed endothelialitis ﬊. Inflammatory cells are seen within the thickened intima ﬈. The biopsy did not show significant tubulointerstitial inflammation. This was a 2year posttransplant protocol biopsy. (Right) Arteries with chronic TCMR have a thickened intima with fibrosis and a sparse infiltrate ſt. In contrast to intimal fibrosis due to hypertension, duplication of the elastica is not prominent.

128

Chronic T-Cell-Mediated Rejection

CLINICAL ISSUES

Abbreviations

Epidemiology

• Chronic T-cell-mediated rejection (TCMR)

• Estimates of prevalence of TCMR in late (> 10 year) biopsies vary from 0% to > 10%

Synonyms • Chronic cellular rejection • Chronic active TCMR (Banff term)

Definitions • Persistent or recurrent T-cell-mediated rejection leading to chronic changes in allograft ○ e.g., transplant arteriopathy, interstitial fibrosis, and tubular atrophy • Chronic active TCMR definition according to Banff 2017 ○ Chronic allograft arteriopathy (arterial intimal fibrosis with mononuclear cell infiltration in fibrosis, formation of neointima) – May represent chronic active antibody-mediated rejection (AMR) as well as TCMR – May also be manifest later in tubulointerstitial compartment ○ i-IFTA, moderate to severe

Presentation • • • •

Chronic renal failure Hypertension Proteinuria May be asymptomatic (subclinical)

Kidney Transplantation

TERMINOLOGY

Prognosis • Interstitial fibrosis with inflammation (i-IFTA) shortens graft survival more than either alone when present in protocol biopsies at 3-12 months (Moresco, Cosio) ○ Inflammation can be in fibrotic or nonfibrotic areas (Gago) ○ i-IFTA correlates with decreased graft survival whether cause is TCMR, AMR, or glomerular disease (Sellares) • Presence of chronic transplant arteriopathy also shortens graft survival

MICROSCOPIC ETIOLOGY/PATHOGENESIS T-Cell-Mediated Injury to Arteries and Tubules/Interstitium • • • •

Alloresponse to human leukocyte antigens Other antigens, including autoantigens, may be relevant Macrophages, mast cells also participate Fibrosis postulated to be from mediators of tubular cells and inflammatory cells ○ Transforming growth factor-β, bone morphogenic protein, platelet-derived growth factor, and hepatocyte growth factor

Antibody-Mediated Component • In chronic TCMR, antibody-mediated component often present ○ Evidence for antibody participation includes transplant glomerulopathy, C4d deposition, and multilamination of peritubular capillary basement membranes ○ Chronic arteriopathy can be mediated by either T cells or antibody ○ Combined chronic AMR and TCMR later post transplant (> 1 year) associated with nonadherence to immunosuppression therapy

Experimental Studies • Chronic allograft arteriopathy in mice has ≥ 3 different pathways ○ T cells in absence of donor-specific antibody (DSA) – Examples: Male to female allografts and B-cell knockout recipients ○ DSA in the absence of T cells – Example: Passive transfer of DSA into T-cell deficient recipients (RAG1 knockout) ○ NK cells in absence of antibody or reactive T cells – Example: Parent to F1 allografts ("hybrid resistance')

Histologic Features • Glomeruli ○ Global glomerulosclerosis ○ Focal segmental glomerulosclerosis ○ Glomerular basement membrane duplication is not typical feature and suggests either chronic AMR or thrombotic microangiopathy • Interstitium ○ Interstitial inflammation in areas of fibrosis (i-IFTA) – Not counted in standard Banff i score – Now counted in Banff i-IFTA score ○ Mast cells correlate with fibrosis • Tubules ○ Tubulitis in nonatrophic or mildly atrophic tubules (Banff t score) • Arteries ○ Intimal fibrosis – Lacks duplication of elastica in intima, typical of hypertensive arteriosclerosis ○ Mononuclear cells in intima – Typically most concentrated under intima – Also present in media and adventitia – May be hard to appreciate without CD3 stain ○ Foam cells (macrophages) in intima – Typically lined up against internal elastica – Not feature of usual atherosclerosis in native kidneys

ANCILLARY TESTS Histochemistry • Elastin stain useful to distinguish chronic allograft arteriopathy from hypertensive changes; latter have fibroelastosis in intima

Immunohistochemistry • CD3 stains can be helpful in appreciating T-cell component 129

Kidney Transplantation

Chronic T-Cell-Mediated Rejection Chronic TCMR Categories Banff 2017 Meeting Report Grade IA Interstitial inflammation involving > 25% of total cortex (ti score 2 or 3) and > 25% of fibrotic cortical parenchyma (i-IFTA score 2 or 3) with moderate tubulitis (t2), not including severely atrophic tubules* Grade IB Interstitial inflammation involving > 25% of total cortex (ti score 2 or 3) and > 25% of sclerotic cortical parenchyma (i-IFTA score 2 or 3) with severe tubulitis (t3), not including severely atrophic tubules* Grade 2 Transplant arteriopathy (chronic allograft arteriopathy): Arterial intimal fibrosis with mononuclear cell inflammation in fibrosis and formation of neointima TCMR = T-cell-mediated rejection; *other known causes of i-IFTA should be ruled out. M Haas et al. The Banff 2017 kidney meeting report: revised diagnostic criteria for chronic active T cell-mediated rejection, antibody-mediated rejection, and prospects for integrative endpoints for next-generation clinical trials. Am J Transplant. ePub 2017.

Immunofluorescence • Negative stain for C4d in peritubular capillaries (PTCs) if no concurrent AMR

Molecular Tests • TCMR transcripts similar to acute TCMR

DIFFERENTIAL DIAGNOSIS Chronic Antibody-Mediated Rejection • Often present in conjunction with chronic TCMR • Features that suggest component of chronic TCMR ○ C4d positive in PTCs – Many chronic AMR cases are C4d negative ○ Donor-specific human leukocyte antibodies ○ Transplant glomerulopathy ○ Multilamination of basement membrane of PTCs • Transplant arteriopathy is pattern seen in chronic AMR or chronic TCMR ○ Histologic patterns indistinguishable

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Chronic TCMR and AMR often coexist • Tubulitis in atrophic tubules is not specific but may be responsible for tubular damage • Inflammation in areas of fibrosis (i-IFTA), at least moderate, now recognized as chronic TCMR pattern (Banff 2017) ○ Other potential causes of i-IFTA must be ruled out • Late stages of diseases often lose their specific diagnostic features

SELECTED REFERENCES 1.

2.

3.

Chronic Calcineurin Inhibitor Toxicity • Severe arteriolar hyalinosis ○ Peripheral nodular hyalinosis • "Striped" fibrosis pattern generally not discriminatory

4.

5.

Hypertensive Arteriosclerosis

6.

• Abundant duplication of elastica in intima • Minimal or no mononuclear infiltrate

7.

Late Stage of BK Polyoma Virus Nephropathy • Prior biopsies showing polyoma virus infection give best clue

8. 9. 10.

Chronic Pyelonephritis • Plasma cells, mononuclear cells, and fewer neutrophils in interstitial infiltrate

11. 12.

Radiation Nephropathy • • • •

130

Fibrosis and tubular atrophy Foam cells in arteries mimic chronic allograft arteriopathy Glomerulosclerosis more prominent than in chronic TCMR Mononuclear infiltrate may be present but with little or no tubulitis

13.

Haas M et al: The Banff 2017 kidney meeting report: revised diagnostic criteria for chronic active T cell-mediated rejection, antibody-mediated rejection, and prospects for integrative endpoints for next-generation clinical trials. Am J Transplant. ePub, 2017 Lefaucheur C et al: T cell-mediated rejection is a major determinant of inflammation in scarred areas in kidney allografts. Am J Transplant. ePub, 2017 Nankivell BJ et al: The causes, significance and consequences of inflammatory fibrosis in kidney transplantation: The Banff i-IFTA lesion. Am J Transplant. ePub, 2017 Halloran PF et al: Disappearance of T cell-mediated rejection despite continued antibody-mediated rejection in late kidney transplant recipients. J Am Soc Nephrol. 26(7):1711-20, 2015 Naesens M et al: The histology of kidney transplant failure: a long-term follow-up study. Transplantation. 98(4):427-35, 2014 El Ters M et al: Kidney allograft survival after acute rejection, the value of follow-up biopsies. Am J Transplant. 13(9):2334-41, 2013 Farris AB et al: Renal interstitial fibrosis: mechanisms and evaluation. Curr Opin Nephrol Hypertens. 21(3):289-300, 2012 Gago M et al: Kidney allograft inflammation and fibrosis, causes and consequences. Am J Transplant. 12(5):1199-207, 2012 Hill GS et al: Donor-specific antibodies accelerate arteriosclerosis after kidney transplantation. J Am Soc Nephrol. 22(5):975-83, 2011 Mannon RB et al: Inflammation in areas of tubular atrophy in kidney allograft biopsies: a potent predictor of allograft failure. Am J Transplant. 10(9):206673, 2010 Park WD et al: Fibrosis with inflammation at one year predicts transplant functional decline. J Am Soc Nephrol. 21(11):1987-97, 2010 Moreso F et al: Subclinical rejection associated with chronic allograft nephropathy in protocol biopsies as a risk factor for late graft loss. Am J Transplant. 6(4):747-52, 2006 Shishido S et al: The impact of repeated subclinical acute rejection on the progression of chronic allograft nephropathy. J Am Soc Nephrol. 14(4):104652, 2003

Chronic T-Cell-Mediated Rejection

T Cells in Thickened Arterial Intima (Left) Biopsy of a renal allograft 22 years posttransplant shows marked thickening of the arterial intima with foamy macrophages st against the internal elastica ﬈ and lymphocytes ſt in the neointima. This patient had a Cr of 2.1 and circulating DSA (class II), but the C4d stain was negative. (Right) The arterial intima in a renal allograft 22 years posttransplant has a marked T-cell infiltrate ſt, meeting the definition of chronic TCMR. The internal elastica ﬈ separates the intima from the media below.

Allograft Arteriopathy vs. Hypertensive Arteriopathy

Kidney Transplantation

Foam Cells in Chronic Allograft Arteriopathy

Chronic Allograft Arteriopathy With CD3(+) Cells (Left) Elastin stains reveal the extensive fibroelastosis of the intima that is characteristic of hypertensive arteriosclerosis (right) and the paucity of elastin fibers in the neointima of chronic allograft arteriopathy (left). (Right) The artery in the right panel has markedly thickened intima, which does not have an obvious infiltrate. However, when the same vessel is stained for CD3 (left panel), a sparse infiltrate of T cells can be appreciated ſt. Internal elastica is indicated st.

Fibrosis With Inflammation in Chronic TCMR

i-IFTA in Chronic Active TCMR (Left) A diffuse mononuclear infiltrate ﬈ is present in the areas with interstitial fibrosis (i-IFTA), which now is recognized as a pattern of chronic TCMR, provided other causes of inflammation are excluded. (Right) This field from a graft 11 years posttransplant shows interstitial fibrosis and tubular atrophy with a diffuse, sparse mononuclear infiltrate. Focal tubulitis is also present st.

131

Kidney Transplantation

Acute Antibody-Mediated Rejection KEY FACTS

TERMINOLOGY • Acute allograft rejection caused by anti-donor-specific antibodies (DSA) reactive to graft endothelium

ETIOLOGY/PATHOGENESIS • DSA usually directed against HLA class I or II on endothelium ○ Activates complement via classical pathway – Early acute AMR in recipients with preformed DSA is complement mediated • Mechanism of AMR likely varies with time post transplant and type of DSA

CLINICAL ISSUES • Acute renal failure • Serum DSA ○ Usually donor specific anti-HLA antibody ○ Acute AMR may occur in ABO blood group-incompatible allografts or with antiendothelial cell or other DSA

• Worse allograft survival in acute AMR compared to C4d(-) TCMR

MICROSCOPIC • • • • • •

Glomerulitis, neutrophils, monocytes, fibrin Glomerular thrombi or mesangiolysis Peritubular capillary neutrophils Dilated peritubular capillaries (PTCs) Acute tubular injury Diffuse, bright positive staining of PTCs for C4d by IF or focal or diffuse for C4d staining by IHC

TOP DIFFERENTIAL DIAGNOSES • • • • • •

Acute T-cell-mediated rejection Acute tubular necrosis Chronic active AMR Recurrent atypical hemolytic uremic syndrome Accommodation Pyelonephritis

Early Acute AMR

C4d(+) Peritubular Capillaries

Acute AMR With Minimal Inflammation

Reactive Endothelial Cells in Acute AMR

(Left) Acute antibodymediated rejection (AMR) is seen 1 week post transplant. Neutrophils are present within peritubular capillaries (PTC) ﬈, and tubules show acute injury ﬊. (Right) Diffuse, bright circumferential staining ﬈ of PTCs for C4d by immunofluorescence (IF) is shown. A positive C4d stain is defined as linear endothelial staining of ≥ 10% of PTC by IF in frozen sections or > 0% of PTC by immunohistochemistry in paraffin-embedded samples.

(Left) Acute AMR can show acute tubular injury with minimal inflammation in PTCs. In this situation, C4d staining is essential for the diagnosis. Note the dilated peritubular capillaries ﬈, which are normally inconspicuous. This biopsy was taken 2 weeks post transplant and was C4d(+). (Right) Endothelial cells are enlarged ﬈ and show loss of fenestrations ﬊ in a transplant from a zero-HLAmismatched kidney.

132

Acute Antibody-Mediated Rejection

Abbreviations • Acute antibody-mediated rejection (acute AMR)

Synonyms • Acute humoral rejection, active AMR, acute/active AMR

Definitions • Acute allograft rejection caused by anti-donor-specific antibodies (DSA) reactive to graft endothelium • Banff 2017: Removal of term "acute" from pathology diagnosis ○ "Acute" confused clinicians in setting of wide range of clinical presentations of "active" AMR

ETIOLOGY/PATHOGENESIS Donor-Specific Antibody • MHC antigens usual target ○ HLA class I or II on endothelium • Non-MHC endothelial antigens ○ ABO blood group antigen in ABO-incompatible grafts ○ Endothelial cell-specific antigens – Acute AMR occurs rarely, in HLA, ABO identical grafts □ Clear evidence for non-MHC target – Endoglin, FLT3L, EDIL3, ICAM4 and others yet to be identified – Most patients with antiendothelial antibodies (AECA) also have anti-HLA alloantibody • Nonendothelial antigens ○ Angiotensin II type 1 receptor

Complement Activation • DSA activates complement via classical pathway ○ C4d is inactive fragment of C4b of classical complement pathway – Covalently bound at site of complement activation on endothelium ○ Complement-fixing DSA and IgG3 subclass of DSA associated with greater acute graft injury

Other Mechanisms • Cell-mediated cytotoxicity (ADCC) ○ NK cells ○ Neutrophils ○ Macrophages via Fc receptors • Direct effects of DSA on endothelium

Mechanisms of Acute AMR Vary • Early acute AMR in positive-crossmatch (+XM) recipients with preformed DSA ○ Probably complement-mediated rejection – C4d deposition in capillaries – Prevention of acute AMR by terminal complement inhibition ○ High serum DSA levels ○ "Pure" acute AMR phenotype, not combined with T-cellmediated rejection • Later active AMR in +XM recipients ○ Histologically manifest as glomerular and peritubular capillaritis on protocol biopsy

○ Variable participation of complement – Often C4d(-) [~ 10% C4d(+) on protocol biopsy, may be focal] – May be mediated by ADCC or direct endothelial damage by alloantibody – May be due to proximal complement pathway components or lower levels of complement activation ○ Usually "pure" AMR phenotype, without concurrent Tcell-mediated rejection (TCMR) • Acute AMR in recipients with de novo DSA ○ Probably varied mechanisms (complement and noncomplement mediated) ○ Often combined with TCMR tubulitis, interstitial inflammation) ○ Major risk factor is nonadherence to immunosuppression ○ Chronic active AMR and TG usually occurs without preceding acute AMR

Kidney Transplantation

TERMINOLOGY

CLINICAL ISSUES Epidemiology • Incidence ○ ~ 25% of acute rejection episodes due to antibody – Overall acute AMR rate: ~ 6% □ Early (< 1 month post transplant) rate: ~ 30-40% among +XM patients with preformed anti-HLA DSA

Presentation • Acute renal failure • Oliguria

Laboratory Tests • Anti-HLA class I or class II DSA ○ Serum DSA level at time of biopsy correlates with severity of biopsy changes • Minority (5-10%) have undetectable DSA ○ May be due to non-HLA antibody ○ Possible antibody absorption by graft

Treatment • • • • •

Plasmapheresis Increased immunosuppression Intravenous immunoglobulin (IVIG) Rituximab (anti-CD20, B cell) Complement inhibition (experimental) ○ C5 inhibitor (eculizumab) ○ C1 esterase inhibitor (C1INH) • Anti-plasma cell therapy (experimental) ○ Proteosome inhibitors (bortezomib) • Splenectomy in resistant cases

Prognosis • Worse allograft survival in acute AMR compared to C4d(-) acute TCMR ○ ~ 30% graft loss within 1 year vs. ~ 4% graft loss for TCMR ○ Plasma cell-rich variant resistant to treatment ○ C4d deposition associated with worse outcome in some but not all studies • Increased risk of developing chronic AMR (transplant glomerulopathy) • Properties of preformed DSA influence risk of acute AMR 133

Kidney Transplantation

Acute Antibody-Mediated Rejection ○ Complement fixing DSA have higher risk of acute AMR ○ IgG3 DSA have higher risk of acute AMR ○ Increased risk of early acute AMR with higher level of DSA – B-cell flow crossmatch channel shift > 359 – Bead assay with molecules of equivalent soluble fluorochrome units of > 34,000

MICROSCOPIC Histologic Features • Glomeruli ○ Glomerulitis, neutrophils, monocytes, fibrin ○ Glomerular thrombi or mesangiolysis – Particularly in ABO blood group-incompatible grafts • Tubules ○ Acute tubular injury ○ Little or no tubulitis ○ Sometimes neutrophils in lumen • Interstitium ○ Edema, sparse infiltrate ○ Hemorrhage occasionally ○ Plasma cell-rich variant – Associated with edema and high interferon-γ • Peritubular capillaries ○ Dilated ○ Neutrophils and mononuclear cells – Termed peritubular capillaritis • Arteries ○ Fibrinoid necrosis in minority of cases ○ Endothelialitis (also feature of TCMR) ○ Thrombotic microangiopathy (TMA) • Banff patterns (grades) of acute AMR ○ Type I: Acute tubular injury, minimal inflammation ○ Type II: PTC &/or glomerular capillary inflammation, &/or thromboses ○ Type III: Arterial fibrinoid necrosis or transmural inflammation (v3 lesion)

ANCILLARY TESTS Immunohistochemistry • Diffuse PTC staining for C4d (C4d > 0) ○ Less sensitive, less reproducible than IF ○ Has artifact of plasma staining

Immunofluorescence • Diffuse, bright C4d(+) staining of PTC (> C4d 1) ○ Small minority of probable acute AMR C4d(-) ○ Focal C4d (10-50%) less commonly has detectable DSA ○ C4d staining remains positive ~ 5-7 days after antibody removal from circulation • Banff classification allows for C4d(-) active AMR ○ Clinically apparent acute AMR nearly always C4d(+) ○ C4d(-) AMR may be immunologically "active" but with more slowly progressive course • C4d positivity correlates with C1q fixation by circulating DSA

Electron Microscopy • PTC and glomerular capillary endothelial changes 134

○ ○ ○ ○ ○ ○

Cell enlargement Loss of fenestrations Microvillous changes Detachment from basement membrane Lysis Apoptosis

DIFFERENTIAL DIAGNOSIS Chronic Active Antibody-Mediated Rejection • Chronic rejection features ○ Transplant glomerulopathy ○ Peritubular capillaropathy ○ Transplant arteriopathy • Usually stable or slowly declining clinical course initially • Mononuclear cells in capillaries with fewer neutrophils, more severe peritubular capillaritis with time post transplant in chronic AMR • Increased Banff PTC score not indicator of clinical acute (vs. chronic) AMR

Acute T-Cell-Mediated Rejection • Interstitial inflammation and tubulitis • 20-30% of TCMR cases C4d(+), indicative of concurrent AMR ○ Late combined acute and chronic TCMR and AMR associated with medication nonadherence

Acute Tubular Necrosis/Injury • C4d(-)

Accommodation • C4d deposition without capillaritis or glomerulitis • Commonly seen in ABO blood group-incompatible grafts

Recurrent Atypical Hemolytic Uremic Syndrome • Thrombotic microangiopathy • May have history of cardiomyopathy, malignant hypertension, or unusual pregnancy complications

Acute Pyelonephritis • Neutrophils and neutrophilic tubulitis may be seen in acute AMR • Neutrophilic casts on biopsy and positive urine culture for bacterial infection favor pyelonephritis • C4d(-)

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Presence of DSA or C4d deposition worsens prognosis of endarteritis

SELECTED REFERENCES 1.

2.

3.

Haas M et al: The Banff 2017 Kidney Meeting Report: Revised Diagnostic Criteria for Chronic Active T Cell-Mediated Rejection, Antibody-Mediated Rejection, and Prospects for Integrative Endpoints for Next-Generation Clinical Trials. Am J Transplant. ePub, 2017 Lefaucheur C et al: IgG donor-specific anti-human HLA antibody subclasses and kidney allograft antibody-mediated injury. J Am Soc Nephrol. 27(1):293304, 2016 Malheiro J et al: Determining donor-specific antibody C1q-binding ability improves the prediction of antibody-mediated rejection in human leucocyte antigen-incompatible kidney transplantation. Transpl Int. ePub, 2016

Acute Antibody-Mediated Rejection

All 3 Features Must Be Present for Diagnosis

Comments

1. Histologic Evidence of Acute Tissue Injury, Including 1 or More of Following Microvascular inflammation (g > 0 &/or ptc > 0)

Recurrent/de novo glomerulonephritis should be excluded

Intimal or transmural arteritis (v > 0)

These arterial lesions may be indicative of AMR, TCMR, or mixed AMR/TCMR; “v” lesions are only scored in arteries having continuous media with 2 or more smooth muscle layers

Acute thrombotic microangiopathy

In absence of any other cause

Acute tubular injury

In absence of any other cause

Kidney Transplantation

Acute/Active Antibody Mediated Rejection: Banff 2013 Criteria

2. Evidence of Current/Recent Antibody Interaction With Vascular Endothelium, Including at Least 1 of Following Linear C4d staining in peritubular capillaries

C4d2 or C4d3 by IF on frozen sections, or C4d > 0 by IHC on paraffin sections

At least moderate microvascular inflammation (g + ptc ≥ 2)

In presence acute TCMR, borderline infiltrates, or evidence of infection, ptc > 2 alone is not sufficient to define moderate microvascular inflammation, and g must be > 1

Increased expression of gene transcripts in biopsy tissue is indicative of At present, only validated molecular markers meeting this criterion are endothelial injury, if thoroughly validated endothelial associated transcripts as used at University of Alberta* 3. Evidence of Donor-Specific Antibodies (HLA or Other Antigens) Usual test is for serum DSA; Banff 2017: PTC C4d staining is indicative of presence of serum DSA, but serum DSA testing is still recommended

Methodology and threshold not specified

AMR = antibody mediated rejection; TCMR = T-cell-mediated rejection; DSA = donor-specific antibody For all AMR diagnoses, it should be specified in the report whether the lesion is C4d(+) (C4d2 or C4d3 by IF on frozen sections; C4d > 0 by IHC on paraffin sections) or without evident C4d deposition (C4d0 or C4d1 by IF on frozen sections; C4d0 by IHC on paraffin sections). Biopsies showing 2 of the 3 features, except those with DSA and C4d without histologic abnormalities potentially related to AMR or TCMR (C4d staining without evidence of rejection) may be designated as “suspicious” for acute/active AMR. *Sis B, et al. Endothelial gene expression in kidney transplants with alloantibody indicates antibody-mediated damage despite lack of C4d staining. Am J Transplant 9: 2312-2323, 2009. Haas M et al: Banff 2013 meeting report: inclusion of C4d-negative antibody-mediated rejection and antibody-associated arterial lesions. Am J Transplant. 14(2):272-83, 2014. 4.

5.

6.

7.

8. 9.

10.

11.

12. 13.

14. 15.

16.

Moktefi A et al: C1q binding is not an independent risk factor for kidney allograft loss after an acute antibody-mediated rejection episode: a retrospective cohort study. Transpl Int. ePub, 2016 Montgomery RA et al: Plasma-derived C1 esterase inhibitor for acute antibody-mediated rejection following kidney transplantation: results of a randomized double-blind placebo-controlled pilot study. Am J Transplant. ePub, 2016 Orandi BJ et al: Presentation and outcomes of C4d-negative antibodymediated rejection after kidney transplantation. Am J Transplant. 16(1):21320, 2016 Viglietti D et al: C1 Inhibitor in acute antibody-mediated rejection nonresponsive to conventional therapy in kidney transplant recipients: a pilot study. Am J Transplant. 16(5):1596-603, 2016 Jackson AM et al: Endothelial cell antibodies associated with novel targets and increased rejection. J Am Soc Nephrol. 26(5):1161-71, 2015 Yell M et al: C1q Binding activity of de novo donor-specific HLA antibodies in renal transplant recipients with and without antibody-mediated rejection. Transplantation. 99(6):1151-5, 2015 Haas M et al: Banff 2013 meeting report: inclusion of C4d-negative antibodymediated rejection and antibody-associated arterial lesions. Am J Transplant. 14(2):272-83, 2014 Loupy A et al: Molecular microscope strategy to improve risk stratification in early antibody-mediated kidney allograft rejection. J Am Soc Nephrol. 25(10):2267-77, 2014 Willicombe M et al: Acute cellular rejection: impact of donor-specific antibodies and C4d. Transplantation. 97(4):433-9, 2014 Lawrence C et al: Preformed complement-activating low-level donor-specific antibody predicts early antibody-mediated rejection in renal allografts. Transplantation. 95(2):341-6, 2013 Loupy A et al: Complement-binding anti-HLA antibodies and kidney-allograft survival. N Engl J Med. 369(13):1215-26, 2013 Jackson AM et al: Multiple hyperacute rejections in the absence of detectable complement activation in a patient with endothelial cell reactive antibody. Am J Transplant. 12(6):1643-9, 2012 Sellarés J et al: Understanding the causes of kidney transplant failure: the dominant role of antibody-mediated rejection and nonadherence. Am J Transplant. 12(2):388-99, 2012

17. Stegall MD et al: The role of complement in antibody-mediated rejection in kidney transplantation. Nat Rev Nephrol. 8(11):670-8, 2012 18. Stegall MD et al: Terminal complement inhibition decreases antibodymediated rejection in sensitized renal transplant recipients. Am J Transplant. 11(11):2405-13, 2011 19. Hidalgo LG et al: NK cell transcripts and NK cells in kidney biopsies from patients with donor-specific antibodies: evidence for NK cell involvement in antibody-mediated rejection. Am J Transplant. 10(8):1812-22, 2010 20. Sis B et al: Endothelial gene expression in kidney transplants with alloantibody indicates antibody-mediated damage despite lack of C4d staining. Am J Transplant. 9(10):2312-23, 2009 21. Burns JM et al: Alloantibody levels and acute humoral rejection early after positive crossmatch kidney transplantation. Am J Transplant. 8(12):2684-94, 2008 22. Lipták P et al: Peritubular capillary damage in acute humoral rejection: an ultrastructural study on human renal allografts. Am J Transplant. 5(12):28706, 2005 23. Desvaux D et al: Acute renal allograft rejections with major interstitial oedema and plasma cell-rich infiltrates: high gamma-interferon expression and poor clinical outcome. Nephrol Dial Transplant. 19(4):933-9, 2004 24. Mauiyyedi S et al: Acute humoral rejection in kidney transplantation: II. Morphology, immunopathology, and pathologic classification. J Am Soc Nephrol. 13(3):779-87, 2002

135

Kidney Transplantation

Acute Antibody-Mediated Rejection

Acute Tubular Injury in Acute AMR

Dilated PTCs

Mesangiolysis and Glomerulitis

Glomerular Thrombus

Plasma Cell-Rich Acute AMR

Plasma Cell-Rich Acute AMR

(Left) In this example of acute AMR 2 weeks post transplant, the tubules are dilated and show flattening of the epithelium ﬈. (Right) PTCs are dilated ﬊ in this example of early acute AMR. There is scant inflammation in the capillaries. C4d stain was positive.

(Left) Mesangiolysis is seen in this example of acute and chronic AMR. Fragmented red blood cells ﬈ are present in the expanded mesangium. (Right) A glomerular thrombus ﬈ is seen in a case of acute AMR 1 week post transplant in a patient with preformed DSA. This pattern of early acute AMR is typically accompanied by high serum donor-specific antibody (DSA) levels and C4d deposition in peritubular capillaries.

(Left) This variant of acute AMR shows marked interstitial edema ﬈ and numerous interstitial ﬊ and PTC ﬊ plasma cells. C4d staining was positive in PTCs. (Right) Interstitial edema ﬈ is seen as pale blue on the trichrome stain in a case of plasma cellrich acute AMR.

136

Acute Antibody-Mediated Rejection

C4d by Immunohistochemistry (Left) Pure acute AMR commonly shows accumulation of neutrophils in PTC ﬈. The histological changes of acute AMR can be subtle. (Right) By IHC, PTCs ﬈ show circumferential positive staining for C4d in this example of acute AMR 1 week post transplant. Glomerular capillaries ﬈ are also positive.

PTC-Reactive Endothelial Cells

Kidney Transplantation

Neutrophils in PTC

Glomerular Endothelial Injury (Left) In acute AMR, PTCs show enlarged endothelial cells by EM. Microvillous projections are seen ﬈, a reactive change. (Right) The glomerular endothelial cells have lost their fenestrations and have a shaggy appearance, a sign of injury and activation ﬉. Segmental dehiscence from the GBM is also seen ﬈. This patient received a liver and kidney transplant 2 weeks previously and had pretransplant DSA to donor class II HLA.

Arterial Fibrin in Acute AMR

Endarteritis in Acute AMR (Left) This case of severe acute AMR shows fibrin ﬊ in an artery along with intimal edema and inflammation. C4d stain was positive. (Right) Endarteritis in patients with DSA may be due to antibodymediated vascular injury, as shown in animal studies. Here the infiltrate in the intima includes neutrophils ﬉ and eosinophils ﬈, not usual features of T-cell-mediated rejection (TCMR).

137

Kidney Transplantation

Acute Antibody-Mediated Rejection

Cortical Necrosis

Combined Acute AMR and Acute TCMR

Resolution of Acute AMR

Normal Glomerulus After Acute AMR

Peritubular Capillaritis in Smoldering AMR

Chronic AMR Sequela of Acute AMR

(Left) Cortical necrosis is manifested by karyolysis in tubules ﬊ and in glomeruli ﬈. This case had both acute and chronic AMR. There was focal C4d staining in PTCs; C4d cannot be assessed in necrotic areas. (Right) Acute AMR and TCMR is often seen in patients with de novo DSA and medication nonadherence. "Pure" AMR is more common in presensitized patients. Interstitial inflammation ﬈, tubulitis ﬈, and peritubular capillaritis ﬊ are present. C4d was positive.

(Left) Follow-up biopsy was performed 90 days post transplant, after an episode of acute AMR at 2 weeks. The congestion and neutrophils in peritubular capillaries have disappeared since the 14-day biopsy. Glomeruli appear normal. Full recovery can be observed in acute AMR, but patients with persistent DSA often show continued capillaritis. (Right) Follow-up biopsy occurred 90 days post transplant, after an episode of acute AMR at 2 weeks. Here, endothelium has recovered ﬈ from the previous biopsy with acute AMR.

(Left) Severe peritubular capillaritis ﬈ is seen in a 1year protocol biopsy in a positive-crossmatch transplant with stable creatinine of 1.2 mg/dL. C4d was negative. Counterintuitively, severe capillaritis does not indicate acute AMR but instead is seen in chronic or smoldering AMR. (Right) TG developed 6 years after an acute AMR episode. Glomeruli show many mononuclear leukocytes ﬈ and duplicated GBM. Anticlass II DSA was present. Chronic AMR may develop ± preceding acute AMR.

138

Acute Antibody-Mediated Rejection

Absence of C4d in Area of Necrosis (Left) The most extreme injury in acute AMR is cortical necrosis, here illustrated in a PAS-stained slide. The necrosis is on the left ﬇. Some viable cortex can also be seen ﬈, as well as thrombi in arteries ſt. (Right) C4d usually fails to stain peritubular capillaries in areas of necrosis ﬇. Lack of staining may be due to lack of perfusion or sloughing of endothelial cells. C4d stains necrotic tubular epithelial cells to a variable degree st. Adjacent more viable cortex does show positive PTC C4d deposition ﬈.

C4d-Negative Acute AMR (C4d0)

Kidney Transplantation

Acute AMR With Cortical Necrosis

Acute AMR C4d1 Medulla (Left) This case of C4dnegative acute AMR does show capillaritis ﬉, meeting one of the criteria for diagnosis. This IHC stain is less sensitive than IF and is more likely to be negative. Certain fixatives also affect IHC. (Right) This case of acute AMR had only the lowest level of C4d staining, C4d1 (1-9% of PTCs). Only 3 capillaries are positive in this field ﬉. The medulla can sometimes be more positive than the cortex.

Acute AMR C4d2

Acute AMR C4d3 (Left) This area of cortex has dilated PTC with little capillaritis. The extent of C4d staining was judged to be ~ 25-30% of peritubular capillaries ﬈, which is scored as Banff C4d2. The glomerular capillaries are also positive ﬉. (Right) This is a low-power view of a biopsy with extensive C4d deposition, approaching 100% of peritubular capillaries. This is a common patten in acute AMR. Capillaritis can also be appreciated even at low power because the C4d stain identifies the capillaries.

139

Kidney Transplantation

Chronic Antibody-Mediated Rejection KEY FACTS

TERMINOLOGY

• Arteries: Neointimal proliferation (transplant arteriopathy)

• Chronic allograft injury mediated by donor-specific antibodies (DSA) reactive to endothelium, particularly glomerular and peritubular capillaries

ANCILLARY TESTS

• Episodic antibody-mediated endothelial injury/activation/repair • DSA against HLA class II most common

• Immunofluorescence or IHC ○ PTC C4d may be diffuse, focal, or negative ○ ~ 60-70% of TG cases C4d(-) on biopsy • Electron microscopy ○ Duplication or multilayering of GBM ○ Multilayering of PTC basement membranes

CLINICAL ISSUES

TOP DIFFERENTIAL DIAGNOSES

• Insidious onset > 1 year post transplant; later post transplant with de novo DSA ○ Proteinuria, chronic renal failure

• Thrombotic microangiopathy • Recurrent or de novo glomerulonephritis • Arteriosclerosis

MICROSCOPIC

DIAGNOSTIC CHECKLIST

• Glomeruli ○ Duplication of GBM (transplant glomerulopathy, TG) ○ Glomerulitis, mostly mononuclear cells • Peritubular capillaritis, mostly mononuclear cells

• Diagnostic criteria (Banff) ○ Histologic evidence of chronic tissue injury ○ Evidence for antibody-mediated injury in tissue ○ Serologic evidence of DSA

ETIOLOGY/PATHOGENESIS

Transplant Glomerulopathy Due to Chronic AMR

Transplant Glomerulopathy

PTC Capillaritis With C4d

Multilamination of Peritubular Capillary Basement Membrane

(Left) Transplant glomerulopathy (TG) with glomerular basement membrane (GBM) duplication ﬈ also showed transplant arteriopathy (TA). Both TG & TA can be seen in chronic AMR but do not necessarily occur together. (Right) Duplication of the GBM ﬉ is striking in this case of chronic antibodymediated rejection (AMR) 10 years post transplant. The endothelium is reactive, with loss of fenestrations ﬈. Electron-dense deposits are not conspicuous. A lymphocyte in the lumen is in contact with the endothelium ﬊.

(Left) Immunoperoxidase stain of chronic humoral rejection (CHR) shows extensive staining of PTCs for C4d; > 90% were positive. Cells in PTC were also evident ﬈. (Right) Chronic AMR characteristically has multilamination of PTC basement membranes, as shown here in a severe example with 9-10 layers ﬉. A pericyte is embedded between the layers ﬉. The endothelium is markedly reactive with increased cytoplasm and organelles ﬈. The lumen contains a lymphocyte.

140

Chronic Antibody-Mediated Rejection

Abbreviations • Chronic antibody-mediated rejection (AMR)

○ ○ ○ ○

Nonadherence Presensitization Younger age, higher HLA mismatches Prior acute AMR or TCMR

Synonyms

Presentation

• Chronic humoral rejection (CHR) • Chronic active AMR

• Insidious onset > 1 year post transplant; later post transplant (5-8 years) with de novo DSA • Indolent dysfunction (38%) • Stable function (32%) (protocol biopsy) • Proteinuria (86% ≥ 0.5 g/d) • Hypertension

Definitions • Chronic allograft injury mediated by donor-specific antibodies (DSA) reactive to endothelium, particularly glomerular and peritubular capillaries (PTCs)

Kidney Transplantation

TERMINOLOGY

Laboratory Tests

ETIOLOGY/PATHOGENESIS Donor-Specific Antibody and Complement • DSA against HLA antigens ○ Chronic AMR strongly associated with class II DSA – May occur with class I DSA alone • Episodic antibody-mediated endothelial injury/activation/repair • DSA activates complement by classical pathway in active AMR ○ C4d deposition in graft is marker of complement activation • DSA may also mediate injury via Fc receptors on NK or monocytes or via early complement components (C3) ○ Progressive capillaritis and transplant glomerulopathy occurs even when patients receive terminal complement inhibitor (anti-C5) ○ Evidence for complement-independent, NK-celldependent mechanism in chronic AMR in murine heart transplants (transplant arteriopathy)

Capillaritis • Microvascular inflammation (MVI) • T cells and monocytes/macrophages • NK cells ○ Increased NK-cell transcripts and NK cells in capillaries of chronic AMR biopsies

Experimental Studies • Sequential stages observed in protocol biopsies of nonhuman primates with de novo DSA ○ Stage I: Circulating DSA, no tissue abnormality ○ Stage II: C4d deposition &/or capillaritis without obvious injury ○ Stage III: Transplant glomerulopathy, PTC lamination, and arteriopathy but normal renal function ○ Stage IV: Proteinuria, loss of function • Antibody-mediated chronic allograft arteriopathy requires NK cells (mice) ○ Complement fixation not required ○ NK cells need cytolytic activity and ability to produce interferon-γ

CLINICAL ISSUES

• Serum donor-specific HLA antibody ○ May be undetectable at time of biopsy ○ Single antigen bead (Luminex) most sensitive – False-positives due to denatured antigens on beads

Natural History • Prospective studies of de novo DSA (Wiebe, Everly) ○ ~ 15-25% develop de novo DSA – Mean: 4.6 years post transplant (6-130 months) in one study – Median: 1.6 years post transplant in another study ○ IgM DSA precedes development of IgG DSA ○ Proteinuria follows onset of DSA by ~ 9 months ○ Elevated Cr follows DSA by ~ 12 months • Chronic AMR with preformed DSA in positive-crossmatch (+XM) kidney transplant recipients ○ Glomerulitis and peritubular capillaritis prevalent on protocol biopsies, precede development of transplant glomerulopathy (TG) – Glomerulitis in ~ 30%, peritubular capillaritis in ~ 60% at 1 year post transplant – C4d deposition present in only ~ 10% of protocol biopsies with capillaritis – Capillaritis present more frequently post transplant in patients with anti-HLA class II DSA – Capillaritis and TG tend to increase in severity with time post transplant • Protocol biopsies after 6 months in stable recipients with de novo DSA show AMR in 51% (Eskandary) ○ Chronic AMR in 36% and acute AMR in 15% – ~ 55% C4d(+) – MFI of DSA is predictor of AMR, no added value of C1q ○ 10% had borderline lesions ○ 9% glomerulonephritis (mostly IgA nephropathy) ○ None had TCMR

Treatment • Drugs ○ No known effective treatments ○ Rituximab (anti-CD20) ○ Intravenous immunoglobulin (IVIG) ○ Bortezomib (proteosome inhibitor) to deplete plasma cells that produce alloantibody

Epidemiology

Prognosis

• ~ 60% of late graft dysfunction due to chronic AMR at some centers • Risk factors

• 50% graft loss rate at 5 years after diagnosis of TG ○ C4d(+) cases have poorer graft survival 141

Kidney Transplantation

Chronic Antibody-Mediated Rejection ○ Subset with molecular markers of endothelial activation shows poorer survival, even if C4d(-) • Greater graft loss with acute (57%) or indolent dysfunction (40%) compared with stable function (0%) over mean of 19 months follow-up

MICROSCOPIC Histologic Features • Glomeruli ○ Duplication of glomerular basement membrane (GBM) – TG (chronic allograft glomerulopathy) ○ Mononuclear cell glomerulitis often present ○ May have segmental glomerulosclerosis, mesangial expansion, or glomerular hypertrophy • Tubules and interstitium ○ No specific changes; tubular atrophy and interstitial fibrosis may be present – Correlates with loss of PTCs ○ Advanced TG with graft dysfunction may occur with minimal interstitial fibrosis/tubular atrophy • Peritubular capillaries ○ Peritubular capillaropathy – Duplicated/thickened PTC BM appreciable by light microscopy when severe – Disappearance of PTCs over time □ Loss of PTCs appreciated with immunostain for endothelial cells (e.g., CD34) □ PTC loss correlates with increasing serum creatinine ○ Peritubular capillaritis – Mononuclear cells in PTCs, particularly moderate to severe (Banff ptc > 1), often seen in chronic AMR – May precede development of TG or other chronic AMR features in presensitized patients with normal graft function □ Often seen on protocol biopsy of presensitized patients (sometimes termed "smoldering" or "indolent" AMR) • Arteries ○ Transplant arteriopathy (chronic allograft arteriopathy) – Fibrous intimal thickening of arteries – Inflammatory cells present within thickened intima □ CD3(+) T cells &/or CD68(+) monocytes/macrophages – Some cases may be histologically indistinguishable from fibrous intimal thickening due to hypertension

ANCILLARY TESTS Immunohistochemistry • C4d deposition in PTCs ○ Can be quite focal or absent ○ Not required for diagnosis of chronic AMR if other evidence of antibody interacting with endothelium (e.g., capillaritis) • Glomerular capillary C4d ○ Glomerular staining for C4d in paraffin sections suggestive of chronic AMR – Also seen in immune complex glomerulonephritis • Cells in peritubular and glomerular capillaries are CD16(+) (many), CD68(+) (many), CD56(+) (few), CD3(+) (few) 142

Immunofluorescence • PTC C4d may be diffuse, focal, or negative ○ Not required for diagnosis of chronic AMR if other evidence of antibody interacting with endothelium (e.g., capillaritis) ○ ~ 60% of TG cases are C4d(-) on biopsy – Antibody levels and C4d deposition can fluctuate with time – May be less active process at time of biopsy – C4d(-) cases may represent non-complement-fixing DSA • IF more sensitive and easier to interpret than IHC on formalin-fixed, paraffin-embedded tissue except for glomeruli ○ Mesangium normally positive for C4d in frozen tissue, not fixed tissue ○ Formalin-fixed, paraffin-embedded tissue may have artifact due to fixation of plasma C4 in lumen of capillaries • Immunoglobulins usually negative, except for segmental IgM and C3 ○ Chronic AMR can be associated with de novo membranous glomerulonephritis

Electron Microscopy • Duplication of GBM, often with multilamination extending circumferentially ○ Hypertrophied endothelium, loss of fenestrae and vacuolization ○ Mesangial cell interposition ○ Early changes detected by EM before development of TG by light microscopy (Banff cg 1a) – Subendothelial lucency – Subendothelial serration of GBM with early GBM duplication – Present as early as 3 months post transplant • Circumferential PTC basement membrane multilamination (PTCBMML) ○ Grading – Mild: 2-4 layers – Moderate: 5-6 layers – Severe: ≥ 7 layers ○ Greater specificity for chronic AMR with higher grade ○ Proposed criteria for severe PTCBMML – 1 PTC with ≥ 7 layers and at least 2 with ≥ 5 layers – Based on 3 most affected of 15-20 PTC examined – Present in 83% with C4d(+) chronic AMR – 1% of native kidneys

Gene Expression • Endothelial gene expression (mRNA microarrays) ○ VWF, DARC, CD31, CD34, CD62e, CAV1, and others • NK gene expression (CXCR1, NKp80, MYBL1, and others) • Can detect evidence of chronic AMR in absence of C4d • Confirmed in formalin-fixed, paraffin-embedded biopsy samples by Nanostring technology (Adam)

DIFFERENTIAL DIAGNOSIS Transplant Glomerulopathy • Chronic thrombotic microangiopathy

Chronic Antibody-Mediated Rejection

Transplant Arteriopathy • Arteriosclerosis ○ Fibroelastosis more prominent than transplant arteriopathy – Elastic fibers can be identified on elastic stain ○ Paucity of inflammatory cells ○ Related to hypertension – May be donor disease – May coexist with transplant arteriopathy ○ Transplant arteriopathy may be indistinguishable from arteriosclerosis due to hypertension • May be due to AMR, T-cell-mediated rejection (TCMR), or both ○ C4d and presence of DSA argue for component of chronic AMR • Atheromatous embolism ○ May show inflammatory response and fibrin ○ Cholesterol clefts may be visible on deeper tissue sections ○ Uncommon finding in allograft • Thrombotic microangiopathy ○ Arteries may show endothelialitis and intimal thickening

REPORTING Banff 2015 Diagnostic Criteria for Chronic AMR • Require 3 elements ○ Histologic evidence of chronic injury ○ Evidence of current or recent antibody interaction with vascular endothelium ○ Serologic evidence of DSA • All 3 criteria required for diagnosis; if only 2 present, considered "suspicious" for chronic AMR

SELECTED REFERENCES 1.

2.

3.

4.

5. 6.

7.

Peritubular Capillaropathy • Mild or segmental PTCBMML seen in various nonspecific causes (e.g., acute tubular injury)

Peritubular Capillary Margination • Acute TCMR type 1 ○ 50% of TCMR has capillaritis ○ Also has interstitial inflammation and tubulitis ○ May be superimposed on chronic AMR • Acute AMR ○ Usually more neutrophils and less capillaritis ○ Clinical presentation of acute renal failure ○ May superimposed on chronic AMR

8.

9.

10. 11. 12. 13.

14.

Accommodation • C4d deposition without histologic evidence of rejection ○ 2-6% of HLA incompatible grafts in protocol biopsies – Probably not "stable" accommodation – Considered stage II of chronic AMR ○ Present in > 90% of ABO incompatible grafts – Apparently stable if no concurrent anti-HLA DSA

15.

16. 17.

18.

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Less extensive (or negative) C4d deposition in chronic AMR compared with acute AMR • When present, C4d deposition is highly specific finding for DSA ○ Arguably, if positive, can substitute for DSA, when DSA testing not available (debated at Banff 2015)

Kidney Transplantation

○ Lacks capillaritis and C4d in PTC • Recurrent or de novo immune complex glomerulonephritis (MPGN, HCV, and others) ○ GBM immune deposits seen by immunofluorescence and electron microscopy

19.

20.

21.

22.

Haas M et al: The Banff 2017 Kidney Meeting Report: Revised diagnostic criteria for chronic active T cell-mediated rejection, antibody-mediated rejection, and prospects for integrative endpoints for next-generation clinical trials. Am J Transplant. 18(2):293-307, 2018 Halloran PF et al: A probabilistic approach to histologic diagnosis of antibody-mediated rejection in kidney transplant biopsies. Am J Transplant. 17(1):129-139, 2017 Adam B et al: Multiplexed color-coded probe-based gene expression assessment for clinical molecular diagnostics in formalin-fixed paraffinembedded human renal allograft tissue. Clin Transplant. 30(3):295-305, 2016 Eskandary F et al: Diagnostic contribution of donor-specific antibody characteristics to uncover late silent antibody-mediated rejection-Results of a cross-sectional screening study. Transplantation. ePub, 2016 Halloran PF et al: Identifying subphenotypes of antibody-mediated rejection in kidney transplants. Am J Transplant. 16(3):908-20, 2016 Lin CM et al: Interferon gamma and contact-dependent cytotoxicity are each rate limiting for natural killer cell-mediated antibody-dependent chronic rejection. Am J Transplant. 16(11):3121-3130, 2016 Parkes MD et al: Evidence for CD16a-mediated NK Cell Stimulation in Antibody-mediated Kidney Transplant Rejection. Transplantation. ePub, 2016 Cornell LD et al: Positive crossmatch kidney transplant recipients treated with eculizumab: outcomes beyond 1 year. Am J Transplant. 15(5):1293-302, 2015 Haas M et al: Banff 2013 meeting report: inclusion of c4d-negative antibodymediated rejection and antibody-associated arterial lesions. Am J Transplant. 14(2):272-83, 2014 Bentall A et al: Five-year outcomes in living donor kidney transplants with a positive crossmatch. Am J Transplant. 13(1):76-85, 2013 Everly MJ et al: Incidence and impact of de novo donor-specific alloantibody in primary renal allografts. Transplantation. 95(3):410-7, 2013 Hirohashi T et al: A novel pathway of chronic allograft rejection mediated by NK cells and alloantibody. Am J Transplant. 12(2):313-21, 2012 Liapis G et al: Diagnostic significance of peritubular capillary basement membrane multilaminations in kidney allografts: old concepts revisited. Transplantation. 94(6):620-9, 2012 Wiebe C et al: Evolution and clinical pathologic correlations of de novo donor-specific HLA antibody post kidney transplant. Am J Transplant. 12(5):1157-67, 2012 Baid-Agrawal S et al: Overlapping pathways to transplant glomerulopathy: chronic humoral rejection, hepatitis C infection, and thrombotic microangiopathy. Kidney Int. 80(8):879-85, 2011 Hill GS et al: Donor-specific antibodies accelerate arteriosclerosis after kidney transplantation. J Am Soc Nephrol. 22(5):975-83, 2011 Gaston RS et al: Evidence for antibody-mediated injury as a major determinant of late kidney allograft failure. Transplantation. 90(1):68-74, 2010 Loupy A et al: Outcome of subclinical antibody-mediated rejection in kidney transplant recipients with preformed donor-specific antibodies. Am J Transplant. 9(11):2561-70, 2009 Sis B et al: Endothelial gene expression in kidney transplants with alloantibody indicates antibody-mediated damage despite lack of C4d staining. Am J Transplant. 9(10):2312-23, 2009 Issa N et al: Transplant glomerulopathy: risk and prognosis related to antihuman leukocyte antigen class II antibody levels. Transplantation. 86(5):6815, 2008 Smith RN et al: Four stages and lack of stable accommodation in chronic alloantibody-mediated renal allograft rejection in Cynomolgus monkeys. Am J Transplant. 8(8):1662-72, 2008 Mauiyyedi S et al: Chronic humoral rejection: identification of antibodymediated chronic renal allograft rejection by C4d deposits in peritubular capillaries. J Am Soc Nephrol. 12(3):574-82, 2001

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Chronic Antibody-Mediated Rejection Banff 2017 Criteria for Chronic Acti Antibody-Mediated Rejection Feature

Comments

Histologic Evidence of Chronic Tissue Injury (≥ 1 of Following) Transplant glomerulopathy (cg > 0)

If no evidence of chronic thrombotic microangiopathy; includes changes evident by EM only (cg 1a)

Severe peritubular capillary basement membrane multilayering

Requires EM; ≥ 7 layers in 1 cortical peritubular capillary and ≥ 5 in 2 additional capillaries, avoiding portions cut tangentially

Arterial intimal fibrosis of new onset, excluding other causes

Leukocytes within sclerotic intima favor chronic AMR if no prior history of biopsy-proven TCMR with arterial involvement (but not required)

Evidence of Current/Recent Antibody Interaction With Vascular Endothelium (≥ 1 of Following) Linear C4d staining in peritubular capillaries

C4d2 or C4d3 by IF on frozen sections, or C4d > 0 by IHC on paraffin sections

At least moderate microvascular inflammation (g + ptc ≥ 2)

In presence of acute TCMR, borderline infiltrate, or infection, ptc ≥ 2 alone is not sufficient and g must be ≥ 1

Increased expression of gene transcripts in biopsy tissue indicative of endothelial injury

If thoroughly validated

Serologic Evidence of DSAs (HLA or Other Antigens) Biopsies suspicious for AMR on basis of meeting criteria 1 and 2 should prompt expedited DSA testing Banff 2017: C4d deposition in PTCs is sufficient evidence of circulating DSA, although serum testing for DSA is still recommended All 3 features must be present for diagnosis. Lesions of chronic active AMR can range from primarily active lesions with early transplant glomerulopathy (TG) evident only by EM (cg 1a) to those with advanced TG and other chronic changes in addition to active microvascular inflammation. Haas M et al: The Banff 2017 Kidney Meeting Report: Revised diagnostic criteria for chronic active T cell-mediated rejection, antibody-mediated rejection, and prospects for integrative endpoints for next-generation clinical trials. Am J Transplant. 18(2): 293-307, 2018.

Scoring of Chronic AMR Lesions Score

Description

Transplant Glomerulopathy cg 0

No GBM double contours by light microscopy or EM

cg 1a

No GBM double contours by light microscopy but GBM double contours in ≥ 3 glomerular capillaries by EM with associated endothelial swelling &/or subendothelial electron lucent widening

cg 1b

≥ 1 glomerular capillary with GBM double contours in 1 glomerulus by light microscopy; EM confirmation recommended, if available

cg 2

GBM double contours in 26-50% of peripheral capillary loops in most affected nonsclerotic glomerulus

cg 3

GBM double contours in > 50% of peripheral capillary loops in most affected nonsclerotic glomerulus

Peritubular Capillaritis* ptc 0

Peritubular capillary (PTC) inflammation in < 10% of cortex

ptc 1

PTC inflammation in ≥ 10% of cortex, with 3-4 marginated luminal inflammatory cells in most affected capillary

ptc 2

PTC inflammation in ≥ 10% of cortex, with 5-10 marginated luminal inflammatory cells in most affected capillary

ptc 3

PTC inflammation in ≥ 10% of cortex, with > 10 marginated luminal inflammatory cells in most affected capillary

Peritubular Capillaropathy (by EM) Mild

≤ 3 circumferential layers in 3 PTCs

Moderate

4-6 circumferential layers in 1 or 2 PTCs

Severe

1 PTC with ≥ 7 layers and at least 2 with ≥ 5 layers by EM

*Peritubular capillaritis is considered an "active" lesion, commonly found in chronic AMR. Haas et al: Banff 2013 Meeting Report: Inclusion of c4d-negative antibody-mediated rejection and antibody-associated arterial lesions. Am J Transplant 14: 272, 2014; Liapis et al: Diagnostic significance of peritubular capillary basement membrane multilaminations in kidney allografts: old concepts revisited. Transplantation 94: 620, 2012.

144

Chronic Antibody-Mediated Rejection

Peritubular Capillaritis (Left) From low magnification, peritubular capillaritis may be inconspicuous; here, focal PTCs are dilated ﬈ and contain marginated inflammatory cells ﬊. There is minimal interstitial fibrosis. (Right) Intracapillary mononuclear cells ﬈ are typically prominent in chronic AMR, with fewer neutrophils. PTCs are dilated in both, as shown here. This patient was 4 years post kidney/liver transplantation and had TG and positive C4d. The liver does not completely protect the kidney from AMR.

Focal C4d Positivity in Chronic AMR

Kidney Transplantation

Capillaritis in Chronic AMR

Prominent C4d in Chronic AMR (Left) Immunoperoxidase stain of paraffin-embedded tissue from a patient with TG and donor-specific antibodies (DSA) shows rare PTCs positive for C4d ﬈. In contrast, the frozen tissue, which is usually more sensitive, was more extensively positive. (Right) C4d may be extensive (as in this case) or even negative in chronic AMR. Diffuse C4d in this setting is associated with noncompliance, a more acute onset of graft dysfunction and later graft loss. This patient also had de novo membranous GN.

Macrophages in PTC

NK Cells in PTC (Left) The cells in PTC are mostly mononuclear cells, which stain for CD68, typical of monocytes/macrophages. The other cells detected by IHC include NK cells and T cells. (Right) Occasional mononuclear CD56(+) cells ﬉, presumably NK cells, can be seen in PTC in chronic AMR. CD56 is on immature NK cells and is lost as the NK cell becomes activated and increases CD16. NK cells are difficult to identify by IHC or IF, since they have no unique surface marker.

145

Kidney Transplantation

Chronic Antibody-Mediated Rejection Transplant Glomerulopathy and Glomerulitis

Glomerulitis and Transplant Glomerulopathy

CD16(+) (FcγRIII) Cells in Chronic AMR

Duplication of GBM

Advanced Transplant Glomerulopathy

Starburst Activated Endothelium

(Left) GBM duplication ﬈ is seen in this example of TG due to chronic AMR 11 years after positive-crossmatch kidney transplantation. Endothelial cells are enlarged. Endocapillary hypercellularity is also present ﬉. By IF, there was focal C4d positivity in PTC. (Right) PAS shows TG with prominent intracapillary mononuclear inflammatory cells (glomerulitis) ﬈, a common feature. Duplication of the GBM is also evident ſt. Patient had class II DSA, and PTCs were positive for C4d.

(Left) Numerous leukocytes stain for the FcγRIII receptor (CD16) in glomerular ﬈ and peritubular capillaries ﬉ in a case of chronic AMR 23 years post transplant. Monocytes and NK cells express CD16. (Right) TG is defined as duplication of the GBM, as shown here in a biopsy 5 years post transplant. The endothelium is reactive with increased organelles and loss of fenestrations. A lymphocyte is in the lumen in contact with the endothelium. C4d was focally positive.

(Left) Some capillary loops show GBM duplication and multilamination ﬊. Note the enlarged endothelial cell ﬈ with processes extending into crevasses in the GBM layers ﬉. (Right) Activation of the glomerular endothelium is manifested by loss of fenestrations and increase in the volume and organelles of the cytoplasm ﬈. Cell processes extend ﬉ into the neo-GBM in a distinctive starburst pattern.

146

Chronic Antibody-Mediated Rejection Stage II: Peritubular Capillaritis and Glomerulitis (Left) Chronic AMR evolves through stages, beginning with production of de novo DSA and progressing to clinically evident chronic AMR. C4d and DSA may be intermittently present during this time. The time between stages is unknown. (Right) A 4month protocol biopsy from a positive-crossmatch transplant patient with stable renal function is shown. Mononuclear cells are marginated within peritubular capillaries ﬈. Glomeruli show no evidence of TG. C4d stain was negative.

Stage II With C4d and No Injury

Kidney Transplantation

Stages of Chronic AMR

Early Ultrastructural Lesions (cg 1a) (Left) This protocol biopsy is from a patient with normal renal function and positive DSA (class II). The only abnormality is the presence of prominent C4d in PTC ﬉ and glomeruli. No capillaritis was evident. 4 years later, the patient developed chronic AMR. (Right) Electron microscopy can reveal early features predictive of later development of TG. Subendothelial lucency ﬉, enlarged endothelial cells ﬈, and corrugation of the GBM ﬊ are seen here, before duplication becomes evident on light microscopy.

Acute AMR With Normal Glomerulus

Acute AMR Sequelae of Chronic AMR (Left) A normal glomerulus is seen on biopsy 6 months post transplant from a patient with graft dysfunction due to acute AMR. There was class II DSA and C4d deposition. (Right) Evolution of glomerular changes is seen 6 years after an acute AMR episode. The patient presented with elevated Cr, proteinuria, persistent class II DSA, and positive C4d. Glomerulitis ﬈ and GBM duplication ﬊ are prominent in this hypertrophied glomerulus. The graft was lost 18 months later.

147

Kidney Transplantation

Chronic Antibody-Mediated Rejection

Peritubular Capillaropathy

Peritubular Capillaropathy

Reactive Endothelium in PTCs

Severe Peritubular Capillaropathy

Mixed TCMR and AMR

Mixed TCMR and AMR

(Left) Sometimes peritubular capillaropathy can be seen on silver stains with basement membrane duplication ﬈ or multilayering, although this finding is best seen by EM. Peritubular capillaritis ﬉ is also seen here. (Right) PTC basement membrane duplication and multilayering ﬈ is seen on this silver stain. These capillaries do not show marginated mononuclear cells, although they are typically present.

(Left) Mononuclear cells ﬊ in PTC are associated with reactive endothelium ﬈. This patient presented with proteinuria (2.8 g/day) and elevated Cr (2 mg/dL) 6 years post transplant. C4d was positive. (Right) This case shows circumferential PTCBMML ﬊ along with activated endothelial cells and marginated mononuclear leukocytes ﬊ within the capillary. Endothelial cells are enlarged and show loss of fenestrations, microvillus changes ﬈, and a starburst pattern ﬉ of extension into the neo-BM.

(Left) Mixed chronic TCMR and AMR is shown with interstitial inflammation and early fibrosis, tubulitis ﬊ in partially atrophic tubules, and peritubular capillaritis ﬈. C4d stain was positive in focal PTCs by IF. (Right) Tubulitis ﬈ is present in partially atrophic tubules, a feature often seen in mixed cellular and humoral rejection that is ongoing. Peritubular capillaritis ﬊ is present as well.

148

Chronic Antibody-Mediated Rejection

Chronic Thrombotic Microangiopathy (Left) Venn diagram shows differential diagnosis of TG. Most cases are due to CHR [57% C4d(+), DSA], but other causes include chronic TMA (13%), hepatitis C virus (4%), MPGN (2%), and idiopathic (24%). (Right) Renal transplant biopsy with TG is due to TMA, associated with calcineurin inhibitor toxicity and severe arteriolar hyalinosis collapsing FSGS. Duplication of GBM and intracapillary leukocytes are conspicuous, but the C4d stain was negative.

Collapsing Glomerulopathy

Kidney Transplantation

Differential Diagnosis of Transplant Glomerulopathy

Membranoproliferative Glomerulonephritis (Left) Collapsing glomerulopathy and thrombotic microangiopathy are shown in an allograft with chronic calcineurin inhibitor toxicity. GBM duplication ﬈ due to TMA can resemble chronic AMR and is one of the causes of transplant glomerulopathy. (Right) Membranoproliferative glomerulonephritis (MPGN), which may mimic transplant glomerulopathy, is distinguished by prominent mesangial and subendothelial deposits by EM ﬈ and C3 ± immunoglobulin by IF.

Transplant Glomerulopathy With De Novo Membranous GN

De Novo Membranous GN and Chronic AMR (Left) This biopsy from a nonadherent recipient 6 years post transplant shows duplication of the GBM ﬉, endothelial swelling and loss of fenestrations ﬈, and foot process effacement ﬈. The serrated subepithelial GBM ﬊ is due to de novo membranous GN. (Right) This patient developed proteinuria 6 years post transplant (original disease obstructive uropathy). The prominent IgG along the GBM is not typical of chronic AMR alone. PTCs were C4d(+). EM showed subepithelial serrations and occasional deposits.

149

Kidney Transplantation

Chronic Antibody-Mediated Rejection

Acute and Chronic TCMR and AMR

Transplant Arteriopathy

Arteriosclerosis

Accelerated Arteriosclerosis

Transplant Arteriopathy

Thrombotic Microangiopathy

(Left) This case shows features of acute and chronic rejection, with T-cell-mediated and antibody-mediated rejection. Intimal thickening with inflammatory cells ﬈ is a lesion of chronic rejection, while fibrin ﬊ is a feature seen in acute AMR. C4d stain was positive. (Right) TA has severe narrowing of the arterial lumen. This biopsy also showed TG. C4d was negative; the patient had a donor-specific HLA class II antibody.

(Left) A typical lesion of arteriosclerosis due to hypertension shows intimal thickening ﬈ but without inflammation. (Right) Biopsy from a positive-crossmatch transplant patient shows accelerated arteriosclerosis with a thickened intima ﬈ of an artery. This appearance can be identical to arteriosclerosis due to hypertension but still may represent transplant arteriopathy.

(Left) This artery shows fibrous intimal thickening and segmental endothelialitis ﬈; the minimal endothelialitis represents a lesion of rejection (either AMR or TCMR). (Right) This example of arterial intimal thickening resembles TA, even with inflammatory cells in the thickened intima ﬈, but was seen in a native kidney. This lesion is suggestive of antiphospholipid antibody syndrome in the native kidney.

150

Chronic Antibody-Mediated Rejection Sequential Ultrastructural Changes in Chronic AMR: 21 Months (Left) Protocol biopsy shows no ultrastructural abnormalities. The endothelium has normal fenestrations ﬉ and the GBM ﬈ is not laminated; ptc0, g0, C4d0, DSA negative, Cr 1.7. (Right) Protocol biopsy was normal by light microscopy and electron microscopy, except for minor segmental duplication of the GBM, of uncertain significance st; ptc0, g0, C4d0, DSA negative, Cr 2.1.

Sequential Ultrastructural Changes in Chronic AMR: 32 Months

Kidney Transplantation

Sequential Ultrastructural Changes in Chronic AMR: 2 Months

Sequential Ultrastructural Changes in Chronic AMR: 58 Months (Left) Protocol biopsy was normal by light microscopy but has very segmental endothelial reaction with processes into the thickened subendothelial space ﬉; ptc0, g0, C4d0, DSA positive, Cr 2.7. (Right) Protocol biopsy had no definite glomerular abnormalities by light microscopy, but by electron microscopy had segmental duplication of the GBM ﬉ with loss of endothelial fenestrations ﬈; cg1a, ptc1, g0, C4d1, DSA positive, Cr 2.0, urine protein/Cr 0.45.

Sequential Ultrastructural Changes in Chronic AMR: 75 Months

Sequential Ultrastructural Changes in Chronic AMR: 81 Months (Left) Protocol biopsy had focal duplication of the GBM by light microscopy and electron microscopy st with loss of endothelial fenestrations ſt. Spikes of endothelial cytoplasm intrude into the subendothelial matrix ﬈; cg1, ptc1, g1, C4d0, DSA anti-class II present, Cr 2.6. (Right) Duplication of the GBM is now florid by electron microscopy st. Reactive endothelial cells have lost fenestrations and have increased cytoplasm ſt; cg3, ptc1, g0, C4d0, DSA anti-class II present, Cr 2.9, urine protein/Cr 3.2.

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Kidney Transplantation

Recurrent Diseases in the Allograft KEY FACTS

CLINICAL ISSUES • Recurrent glomerular disease is 3rd leading cause of graft failure ○ Recurrence rates vary by specific disease ○ Focal segmental glomerulosclerosis (FSGS), atypical hemolytic uremic syndrome, and membranoproliferative glomerulonephritis (MPGN) have greatest impact on graft survival ○ Timing of recurrence varies from minutes (FSGS) to years (diabetic glomerulopathy) • Diseases may recur subclinically • Documentation of primary cause of ESRD essential to diagnose recurrence

• Superimposed features of chronic rejection or drug toxicity may be present • Early stages of recurrent disease appreciated on early protocol biopsies or biopsies performed for other indication (e.g., with acute rejection)

TOP DIFFERENTIAL DIAGNOSES • De novo disease ○ Early onset (< 1-2 years) favors recurrence of immune complex GN • Acute transplant glomerulitis • Transplant glomerulopathy (chronic humoral rejection, hepatitis C infection, &/or chronic thrombotic microangiopathy)

MICROSCOPIC

DIAGNOSTIC CHECKLIST

• Pathology resembles primary disease but not identical ○ Diagnosis of recurrent glomerular disease usually requires IF and EM

• Must ascertain primary kidney disease • Prepare to perform IF and EM, particularly if signs of glomerular disease on biopsy > 1 year post transplant

Recurrent MGN

Recurrent MGN, IgG

Recurrent FSGS

Diffuse Foot Process Effacement

(Left) PAS shows marked thickening of the glomerular basement membranes (GBMs) ﬊ with a vacuolated appearance, characteristic of membranous glomerulonephritis (MGN). Other injuries include segmental glomerular scarring ﬈ with a fibrous attachment st to the Bowman capsule and marked arteriolar hyalinosis ﬉ in this 7-year-old allograft. (Right) Immunofluorescence for IgG shows strong granular staining of the glomerular capillary walls ſt in an allograft with recurrent MGN.

(Left) PAS highlights collapsed glomerular tufts ﬈ with prominence of podocytes ﬊. Recurrence of collapsing focal segmental glomerulosclerosis (FSGS) may manifest with either collapsing or noncollapsing segmental sclerosis. (Right) EM shows extensive effacement of the podocyte foot processes ﬈ in this biopsy with recurrence of FSGS within 2 weeks after renal transplantation.

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Recurrent Diseases in the Allograft

Abbreviations • • • •

Membranous glomerulonephritis (MGN) Focal segmental glomerulosclerosis (FSGS) Membranoproliferative glomerulonephritis (MPGN) Hemolytic uremic syndrome (HUS)

Definitions • Recurrence of original cause of end-stage renal disease after kidney transplantation

ETIOLOGY/PATHOGENESIS Proposed Mechanisms • Pathogenesis of recurrence presumably same as original disease ○ Recurrence often as evidence for circulating factor – Give insights into mechanism and early stages ○ Immunosuppressive medications and allogeneic kidney may alter pathogenesis and disease course in allograft • Primary FSGS (nonfamilial) ○ Humoral or permeability factor postulated due to rapid recurrence after kidney transplantation in nonhereditary etiologies – No definitive identification of putative factor □ Recent significant doubts on specificity of soluble urokinase-type plasminogen activator receptor for primary FSGS □ Cardiotrophin-like cytokine-1 also possible candidate ○ Hereditary causes generally do not recur – NPHS1 (nephrin) mutation: Antinephrin alloantibodies lead to podocyte injury and proteinuria (not true "recurrence") – NPHS2 (podocin) mutation: Recurrence may be steroid responsive, even if original disease was steroid resistant • MGN ○ Circulating antibodies against antiphospholipase A2 receptor (PLA2R) in most adults – Anti-PLA2R at transplant associated with disease recurrence in allograft • MPGN ○ Varying causes; some due to defect in complement regulation ○ Immunoglobulin and complement deposits in glomeruli • C3 glomerulopathy ○ Includes dense deposit disease and C3 GN ○ Due to inherited or acquired defect in regulation of alternative complement pathway ○ Immunofluorescence (IF) staining for C3 only or C3 with sparse immunoglobulin deposits – Native kidney disease often misdiagnosed as postinfectious GN or MPGN before description of C3 GN ○ Can recur predominantly as thrombotic microangiopathy/atypical HUS • Atypical HUS ○ CFH (complement factor H) and CFI (complement factor I) mutations

– Recurrence rate is 80% for CFH and 90% for CFI mutations – CFH and FI synthesized by liver – Simultaneous liver and kidney transplantation is treatment option ○ CD46 (membrane cofactor protein) mutation – 20% recurrence rate – Normal CD46 in kidney allograft corrects defect ○ Some immunosuppressive medications cause thrombotic microangiopathy – May potentiate underlying propensity to form thrombi • Proliferative GN with monoclonal IgG deposits (PGMID) ○ Deposition of monoclonal immunoglobulin in glomeruli of allograft ○ Monoclonal protein may not be detectable in serum or in urine pre- or post transplant – Detectable in allograft – IgG3-κ (most common) ○ Recurrent PGMID with more aggressive clinical course – May recur in absence of detectable monoclonal protein • Primary hyperoxaluria type 1 ○ Gene defect for liver peroxisomal enzyme alanine:glyoxylate aminotransferase (AGT) ○ Impaired metabolism of glyoxylate to glycine ○ End-stage renal disease due to oxalate nephropathy ○ Liver transplantation cures enzyme defect – Performed in combination with kidney transplantation for patients not responsive to pyridoxine therapy – Kidney transplantation alone considered for patients with pyridoxine responsiveness

Kidney Transplantation

TERMINOLOGY

CLINICAL ISSUES Epidemiology • Incidence ○ Recurrent disease causes > 15% of allograft loss – Most common recurrent diseases that cause graft loss are FSGS, IgA nephropathy, MGN, and MPGN

Presentation • Acute renal failure • Chronic renal failure • Proteinuria ○ MGN – High recurrence rate (~ 42%) – Recurrence detected by biopsy as early as 2 weeks even in absence of proteinuria □ Histologic or immunophenotypic recurrence precedes proteinuria ○ FSGS – Proteinuria, may be nephrotic range – Recurrence 5x more likely in childhood-onset FSGS vs. adult-onset FSGS □ Independent of age at transplantation – Increased recurrence rate with living-related organ donation – Severe proteinuria may occur immediately post transplant 153

Kidney Transplantation

Recurrent Diseases in the Allograft □ Lesser proteinuria may develop and become progressive over first 1-2 years post transplant ○ Amyloidosis ○ Diabetic glomerulosclerosis – Common cause of end-stage renal disease – Recurs later post transplant (5-10 years) □ Common in 5- and 10-year allograft protocol biopsies □ Recurrence more prevalent with increased graft survival ○ Other glomerular diseases • Hematuria ○ IgA nephropathy ○ MPGN ○ Lupus nephritis ○ Pauci-immune [antineutrophil cytoplasmic antibody (ANCA)-associated] crescentic GN ○ Antiglomerular basement membrane (GBM) nephritis

Laboratory Tests • Serologic tests ○ ANCA titer ○ Anti-GBM antibody titer ○ Donor-specific antibody ○ PLA2R antibody ○ Complement levels • Serum or urine protein electrophoresis, serum-free light chains

Treatment • Drugs ○ Treatment depends on specific glomerular disease – Increased steroids – Cyclophosphamide – Rituximab – Eculizumab (experimental for atypical HUS and some complement-mediated glomerulopathies) • Plasmapheresis ○ FSGS ○ Anti-GBM disease ○ Pauci-immune (ANCA-associated) crescentic GN ○ HUS • Metabolic replacement transplant ○ Some recurrent metabolic diseases can be prevented by simultaneous liver transplantation – Primary hyperoxaluria – Familial HUS patients with CFH or CFI mutations

Prognosis • Variable ○ Depends on original renal disease ○ Lupus nephritis tends not to recur or recurs with minor clinical significance • 3 diseases with worst prognosis ○ Primary FSGS ○ MPGN ○ Atypical HUS

154

MICROSCOPIC Histologic Features • Morphologic features similar to primary disease ○ May be seen at earlier stages due to concurrent immunosuppression • MGN ○ Early recurrent MGN – Normal glomeruli by light microscopy – IF: Granular capillary wall staining for C4d, IgG, κ, λ □ C3 often negative or faint – EM: No or very few tiny subepithelial deposits ○ Late recurrent MGN – Thickened appearance of GBM – GBM spike formation seen with silver stains – IF: Granular capillary wall IgG, κ, λ, C4d □ Unlike native kidneys, C3 often negative or faint on IF – EM: Well-formed subepithelial deposits, like MGN in native kidney • FSGS ○ Podocyte injury or foot process effacement precedes histologic finding of segmental sclerosis – Appears as minimal change disease prior to development of FSGS lesions ○ Segmental glomerular sclerosis with either similar or different features to FSGS prior to transplantation • Lupus nephritis ○ Mesangial hypercellularity or sclerosis ○ Crescent formation or fibrinoid necrosis ○ Segmental sclerosis or prominent podocyte injury may represent unusual manifestation of recurrent disease • IgA nephropathy ○ Glomerular alterations range from normal to marked mesangial hypercellularity with cellular crescent formation similar to disease in native kidneys ○ IF: Glomerular IgA-dominant deposits • Complement-mediated glomerular disease

DIFFERENTIAL DIAGNOSIS De Novo Glomerular Disease • Biopsy documentation helps distinguish de novo from recurrent disease • De novo FSGS ○ Develops > 1 year ○ Generally of "secondary" or maladaptive type ○ Glomerular hypertrophy ○ Arteriolar hyalinosis ○ Contribution of calcineurin inhibitors suspected – Seen in native kidneys in heart, liver, and lung transplant recipients • De novo MGN ○ Often occurs late (~ 5 years) post transplant ○ Associated with features of chronic antibody-mediated rejection ○ Negative for PLA2R • De novo C1q nephropathy ○ C1q-dominant deposits in glomeruli ○ Usually clinically insignificant

Recurrent Diseases in the Allograft 7.

8.

Acute Transplant Glomerulitis

9.

• Prominent glomerular capillary leukocytic infiltration ○ Resembles endocapillary hypercellularity • No glomerular immune complex deposition • Usually associated with endothelialitis

10.

11.

Chronic Antibody-Mediated Rejection • Duplication of GBM with loss of endothelial fenestrations • Usually, but not always, has C4d deposition in peritubular capillaries • Glomerulitis usually present • Multilamination of peritubular capillary basement membranes • Donor-specific circulating antibody • Rare immune complex deposits may be present

12.

Donor-Transmitted Glomerular Diseases

17.

• Implantation (time-zero) biopsy helps diagnose donor disease • IgA nephropathy ○ Glomerular IgA deposits in ~ 10% of normal donor biopsies ○ Donor IgA nephropathy with mesangial proliferation may occur – IgA deposits disappear with time post transplant • MGN • Diabetic glomerulosclerosis

13.

14. 15.

16.

18. 19. 20. 21. 22.

23.

Thrombotic Microangiopathy

24.

• Associated with calcineurin inhibitor toxicity • Increased risk with CFH mutations

25.

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Knowledge of original cause of end-stage renal disease usually necessary for accurate evaluation • Granular C4d glomerular capillary wall deposition raises consideration of de novo or recurrent immune complex GN • Significant glomerular alterations (endocapillary proliferation, basement membrane alterations, increased circulating leukocytes) should trigger additional IF and EM studies ○ Characteristic features may not be present in early stages of recurrent GN

SELECTED REFERENCES 1. 2.

3. 4. 5. 6.

Allen PJ et al: Recurrent glomerulonephritis after kidney transplantation: risk factors and allograft outcomes. Kidney Int. 92(2):461-469, 2017 Cosio FG et al: Recent advances in our understanding of recurrent primary glomerulonephritis after kidney transplantation. Kidney Int. 91(2):304-314, 2016 Delville M et al: B7-1 Blockade does not improve post-transplant nephrotic syndrome caused by recurrent FSGS. J Am Soc Nephrol. 27(8):2520-7, 2016 Kattah AG et al: Temporal IgG subtype changes in recurrent idiopathic membranous nephropathy. Am J Transplant. 16(10):2964-72, 2016 Roberti I et al: Immune-mediated nephropathies in kidney transplants: recurrent or de novo diseases. Pediatr Transplant. 20(7):946-951, 2016 Çeltİk A et al: Recurrent lupus nephritis after transplantation: clinicopathological evaluation with protocol biopsies. Nephrology (Carlton). 21(7):601-7, 2016

26.

27.

28. 29. 30. 31.

32.

33. 34. 35.

36. 37.

Barbour S et al: Advances in the understanding of complement mediated glomerular disease: implications for recurrence in the transplant setting. Am J Transplant. 15(2):312-9, 2015 Kattah A et al: Anti-phospholipase A2 receptor antibodies in recurrent membranous nephropathy. Am J Transplant. 15(5):1349-59, 2015 Green H et al: Recurrent membranoproliferative glomerulonephritis type I after kidney transplantation: a 17-year single-center experience. Transplantation. 99(6):1172-7, 2014 Von Visger JR et al: The risk of recurrent IgA nephropathy in a steroid-free protocol and other modifying immunosuppression. Clin Transplant. 28(8):845-54, 2014 Wühl E et al: Renal replacement therapy for rare diseases affecting the kidney: an analysis of the ERA-EDTA Registry. Nephrol Dial Transplant. 29 Suppl 4:iv1-8, 2014 Zand L et al: Clinical findings, pathology, and outcomes of C3GN after kidney transplantation. J Am Soc Nephrol. 25(5):1110-7, 2014 Kowalewska J: Pathology of recurrent diseases in kidney allografts: membranous nephropathy and focal segmental glomerulosclerosis. Curr Opin Organ Transplant. 18(3):313-8, 2013 Servais A et al: C3 glomerulopathy. Contrib Nephrol. 181:185-93, 2013 Blosser CD et al: Very early recurrence of anti-Phospholipase A2 receptorpositive membranous nephropathy after transplantation. Am J Transplant. 12(6):1637-42, 2012 Canaud G et al: Recurrence from primary and secondary glomerulopathy after renal transplant. Transpl Int. 25(8):812-24, 2012 Rodriguez EF et al: The pathology and clinical features of early recurrent membranous glomerulonephritis. Am J Transplant. 12(4):1029-38, 2012 Nasr SH et al: Proliferative glomerulonephritis with monoclonal IgG deposits recurs in the allograft. Clin J Am Soc Nephrol. 6(1):122-32, 2011 Lorenz EC et al: Recurrent membranoproliferative glomerulonephritis after kidney transplantation. Kidney Int. 77(8):721-8, 2010 Ponticelli C et al: Posttransplant recurrence of primary glomerulonephritis. Clin J Am Soc Nephrol. 5(12):2363-72, 2010 Said SM et al: C1q deposition in the renal allograft: a report of 24 cases. Mod Pathol. 23(8):1080-8, 2010 Czarnecki PG et al: Long-term outcome of kidney transplantation in patients with fibrillary glomerulonephritis or monoclonal gammopathy with fibrillary deposits. Kidney Int. 75(4):420-7, 2009 El-Zoghby ZM et al: Identifying specific causes of kidney allograft loss. Am J Transplant. 9(3):527-35, 2009 Boyer O et al: Complement factor H deficiency and posttransplantation glomerulonephritis with isolated C3 deposits. Am J Kidney Dis. 51(4):671-7, 2008 Ivanyi B: A primer on recurrent and de novo glomerulonephritis in renal allografts. Nat Clin Pract Nephrol. 4(8):446-57, 2008 Jeong HJ et al: Progression of renal allograft histology after renal transplantation in recurrent and nonrecurrent immunoglobulin A nephropathy. Hum Pathol. 39(10):1511-8, 2008 Meehan SM et al: Pauci-immune and immune glomerular lesions in kidney transplants for systemic lupus erythematosus. Clin J Am Soc Nephrol. 3(5):1469-78, 2008 Joshi K et al: Recurrent glomerulopathy in the renal allograft. Transplant Proc. 39(3):734-6, 2007 Casquero A et al: Recurrent acute postinfectious glomerulonephritis. Clin Nephrol. 66(1):51-3, 2006 Choy BY et al: Recurrent glomerulonephritis after kidney transplantation. Am J Transplant. 6(11):2535-42, 2006 Little MA et al: Severity of primary MPGN, rather than MPGN type, determines renal survival and post-transplantation recurrence risk. Kidney Int. 69(3):504-11, 2006 Braun MC et al: Recurrence of membranoproliferative glomerulonephritis type II in renal allografts: The North American Pediatric Renal Transplant Cooperative Study experience. J Am Soc Nephrol. 16(7):2225-33, 2005 Couser W: Recurrent glomerulonephritis in the renal allograft: an update of selected areas. Exp Clin Transplant. 3(1):283-8, 2005 Kowalewska J et al: IgA nephropathy with crescents in kidney transplant recipients. Am J Kidney Dis. 45(1):167-75, 2005 Soler MJ et al: Recurrence of IgA nephropathy and Henoch-Schönlein purpura after kidney transplantation: risk factors and graft survival. Transplant Proc. 37(9):3705-9, 2005 Floege J: Recurrent glomerulonephritis following renal transplantation: an update. Nephrol Dial Transplant. 18(7):1260-5, 2003 Briganti EM et al: Risk of renal allograft loss from recurrent glomerulonephritis. N Engl J Med. 347(2):103-9, 2002

Kidney Transplantation

• De novo diabetic glomerulosclerosis ○ Often occurs late post transplant ○ May be due to steroid use

155

Kidney Transplantation

Recurrent Diseases in the Allograft Recurrent Disease After Kidney Transplantation Renal Disease

Recurrence Rate

5- to 10-Year Graft Loss

Additional Features

MGN

40-50%

10-15% (10 years)

Presence of PLA2R Ab pretransplant has ~ 70% risk of recurrence vs. 28-30% if PLA2R antibody is negative

IgA nephropathy/Henoch-Schönlein purpura

13-50%

10% (10 years)

IgA deposits may be seen in protocol biopsies in patients with normal function; higher rate of disease recurrence reported in patients on steroid-free regimens; HSP and IgA nephropathy have similar rates

MPGN type I (polyclonal Ig)

30-35%

10-40%

Recurrence rate increases with successive allografts

Lupus nephritis

Up to 30%

< 5%

Glomerular immune complex deposits may be seen in protocol biopsies in patients with normal function; podocytopathy or FSGS may represent manifestation of recurrent lupus nephritis

FSGS

30-35%

15-20% (10 years)

May recur with features of collapsing glomerulopathy

Diabetic nephropathy

> 50%

5%

Recurs late post transplant (5-10 years)

Dense deposit disease (formerly MPGN type II) > 80%

10-20%

10-20% of living-related grafts do better than deceased-donor grafts

C3 GN

67%

50% (~ 3 years)

Can recur predominantly as atypical HUS

Atypical HUS (or non-Shiga toxin)

33-82%

40-50%

High recurrence rate for CFH and CFI mutations; 20% recurrence for CD46 mutation

Amyloidosis, AL type

10-30%

35%

Recurrence depends on treatment responsiveness of underlying disease

Amyloidosis, AA type

< 10%

Rare

Immune Complex Mediated

Nonimmune Complex Mediated

Complement Mediated

Deposits With Substructure or Monoclonality

Proliferative GN with monoclonal IgG deposits 66%

50% (2-3 years)

May recur even in absence of detectable circulating monoclonal protein

Fibrillary GN

50%

20%

Monoclonal gammopathy-associated fibrillary GN likely to recur; other fibrillary GN less likely to recur; fibrillary GN may recur very late post transplant (> 15 years)

Immunotactoid glomerulopathy

Rare

Not available

Monoclonal immunoglobulin deposition disease

70-85%

> 50%

Antiglomerular basement membrane disease

< 5%

Rare

Pauci-immune (ANCA-associated) crescentic GN

0-20%

10% (10 years)

Crescentic Glomerulonephritis Recurrent disease may spare kidney allograft

Genetic/Metabolic Disorders Primary hyperoxaluria type 1

90-100% if 80-100% kidney transplant alone in pyridoxineresistant patients

Liver transplantation is curative and may obviate need for kidney transplantation; excretion of oxalate may simulate disease recurrence

Fabry disease

Low

Rare

Decreasing recurrence with advent of enzyme replacement therapy

Cystinosis

Rare

0%

Macrophages with cystine crystals may deposit in interstitium or mesangium; cystine accumulates in other organs

Sickle cell nephropathy

Rare

Not available

Sickle cell crisis common within 1st year of transplantation

Percentages are approximate, some based on combined reported series. Recent review: Cosio FG and Cattran DC. Recent advances in our understanding of recurrent primary glomerulonephritis after kidney transplantation. Kidney Int 91: 304-314, 2017.

156

Recurrent Diseases in the Allograft

IgA (Left) Mild mesangial hypercellularity ﬈ is the predominant finding in this glomerulus from an allograft with recurrent IgA nephropathy (PAS). (Right) IF for IgA shows strong granular mesangial ſt staining in a kidney allograft with recurrent IgA nephropathy. This is identical to the pattern of involvement in the native kidneys.

Recurrent Lupus Nephritis

Kidney Transplantation

Recurrent IgA Nephropathy

Recurrent Lupus Nephritis (Left) PAS shows focal mesangial hypercellularity ﬈ and mesangial sclerosis ﬊, which are common alterations in the early phase of recurrent lupus nephritis. When glomeruli are normal on light microscopy, immunofluorescence microscopy may detect the presence of immune complexes in patients with lupus. (Right) Segmental fibrinoid necrosis ﬈ and endocapillary hypercellularity ﬊ are present in this case of recurrent lupus nephritis.

IgG

C4d (Left) IF for IgG reveals discrete granular and confluent staining along the capillary walls and some mesangial regions ſt in this kidney allograft with lupus nephritis with proliferative and membranous features. (Right) IF shows granular C4d staining along the GBM, which is suggestive of lupus MGN. This staining pattern is distinct from the mesangial C4d staining that may be present in normal glomeruli.

157

Kidney Transplantation

Recurrent Diseases in the Allograft

Early Recurrent MGN

Recurrent MGN

Recurrent MPGN

Recurrent MPGN

Recurrent FSGS

Electron Microscopy in Recurrent FSGS

(Left) At 2 months post transplant, this glomerulus appears normal by light microscopy. By IF, there was bright granular GBM staining for C4d and less staining for IgG, κ, and λ (with negative C3), indicative of recurrent MGN. No deposits were identified by EM. (Right) PAS reveals thick GBMs, characteristic of MGN in an allograft with recurrent disease. Duplicated GBMs ﬈ indicate chronic transplant glomerulopathy.

(Left) There is severe infiltration by inflammatory cells ﬉ within this glomerulus and duplication of the GBMs ﬈ in an allograft with recurrence of membranoproliferative glomerulonephritis (MPGN). (Right) PAS reveals a cellular crescent ﬈ that accompanies marked endocapillary hypercellularity with numerous inflammatory cells ﬊ and duplication of the GBM ﬉ in an allograft with recurrent MPGN type I.

(Left) PAS shows segmental occlusion of glomerular capillaries by hyaline and matrix ﬊ in this allograft with recurrent FSGS 6 years after transplantation. Focal duplication of GBM ﬈ suggests chronic transplant glomerulopathy. (Right) In this biopsy from an adult man with recurrent FSGS in a transplant, a podocyte ﬈ appears to have "fallen off" the GBM. Urinary podocyte excretion has been found to be increased in patients with FSGS.

158

Recurrent Diseases in the Allograft

Recurrent Pauci-Immune GN (Left) Recurrent antineutrophil cytoplasmic antibody (ANCA)associated disease has a fibrocellular crescent ﬈ and a spared segment of the glomerulus ﬊. The Bowman capsule is disrupted ﬉, a useful sign of crescents. The patient had posttransplant pulmonary hemorrhage, positive myeloperoxidase antibody, and recurrent pauciimmune crescentic GN. (Right) PAS reveals a cellular crescent ſt in this allograft from a 57year-old woman with recurrent ANCA-associated pauci-immune crescentic GN.

Recurrent Diabetic Nephropathy

Kidney Transplantation

Recurrent Pauci-Immune GN

Arteriolar Hyalinosis (Left) PAS shows diffuse mesangial ﬊ and nodular sclerosis ﬈, which characterizes recurrence of diabetic nephropathy in an allograft. (Right) Severe arteriolar hyalinosis is present ﬈ in this allograft with recurrent diabetic nephropathy 6 years after transplantation. Calcineurin inhibitor toxicity and hypertension may also contribute, as their histologic features can be indistinguishable from diabetic vascular injury.

Recurrent Amyloidosis

Amyloid A Immunohistochemistry (Left) H&E shows prominent deposition of amorphous eosinophilic material ﬈ in a hilar arteriole, suggestive of recurrent amyloidosis in this 9year-old allograft from a 47year-old man with ankylosing spondylitis. (Right) IHC confirms the presence of amyloid A deposits in a 47year-old man with ankylosing spondylitis. Note the prominent amyloid deposition within the arterioles ﬈ and much less involvement of mesangial areas ﬊ in the glomeruli.

159

Kidney Transplantation

De Novo Focal Segmental Glomerulosclerosis KEY FACTS

TERMINOLOGY

CLINICAL ISSUES

• Definition ○ Focal segmental glomerulosclerosis (FSGS) or its variants in kidney transplant patients with original disease not due to FSGS

• • • •

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• Hyperfiltration ○ Longstanding allografts with severe nephron loss ○ Pediatric kidneys transplanted into adult recipients • Severe vascular disease ○ De novo collapsing glomerulopathy (CG) associated with deceased donor kidneys ○ Zonal distribution of CG • Drug induced ○ Higher frequency with calcineurin inhibitors (CNI) – CNI may contribute via microvascular disease ○ Mammalian target of rapamycin inhibitors • Donor APOL1 risk alleles

• FSGS or CG • Arterial or arteriolar sclerosis

Proteinuria, nephrotic range Chronic renal failure De novo FSGS: 40% graft loss within 5 years of diagnosis De novo CG: 50% loss of graft within 1 year of diagnosis

TOP DIFFERENTIAL DIAGNOSES • • • • •

Recurrent FSGS Calcineurin inhibitor toxicity Recurrent atypical hemolytic uremic syndrome Chronic transplant glomerulopathy Immune complex-mediated glomerulonephritis, de novo or recurrent

Prominent Epithelial Cells

Segmental Sclerosis

Collapsing Focal Segmental Glomerulosclerosis

Severe Arteriolar Hyalinosis

(Left) Collapsing-type de novo focal segmental glomerulosclerosis in a renal transplant patient with nephrotic-range proteinuria shows prominent adjacent visceral epithelial cells (podocytes) ſt. (Right) Protein reabsorption droplets ﬇ accentuate a few prominent podocytes, which correlate with proteinuria. Duplication of the glomerular basement membranes is present ﬈, which raises the consideration of chronic transplant glomerulopathy (Jones methenamine silver).

(Left) Prominent podocytes ﬈ overlie a collapsed tuft in this case of de novo collapsing glomerulopathy in a transplant patient with severe proteinuria, whose original disease was diabetic nephropathy. (Right) Prominent subendothelial hyalinosis ﬈ of an arteriole is present in an allograft biopsy with collapsing glomerulopathy, implying that it may be related to calcineurin inhibitor toxicity.

160

De Novo Focal Segmental Glomerulosclerosis

Abbreviations • Focal segmental glomerulosclerosis (FSGS)

Definitions • FSGS or its variants in kidney transplant patients with original disease not due to FSGS

ETIOLOGY/PATHOGENESIS Hyperfiltration • Pediatric kidneys transplanted into adult recipients • Longstanding allografts with severe nephron loss

Severe Vascular Disease • De novo collapsing glomerulopathy (CG) associated with deceased donor kidneys • Zonal distribution of CG

Drug Induced • Higher frequency with calcineurin inhibitors (CNI) ○ CNI may contribute via microvascular disease • Mammalian target of rapamycin inhibitors

Donor Risk Factors • APOL1 risk alleles in donor kidney

CLINICAL ISSUES Presentation • Proteinuria ○ Variable, unlike recurrent FSGS • Chronic renal failure

Laboratory Tests • Urinalysis • 24-hour urine protein collection

○ Zonal distribution • Global glomerulosclerosis • Interstitial fibrosis and tubular atrophy ○ Often severe • Interstitial foam cells can be correlate of severe proteinuria • Arteriolar hyalinosis ○ Subendothelial hyalinosis ○ Adventitial hyaline nodules may be present

ANCILLARY TESTS Immunofluorescence • IgM and C3 in segmental scars

Electron Microscopy • Podocyte foot process effacement • Separation of podocytes from glomerular basement membrane (GBM) if CG

Genetics • Allografts that develop collapsing FSGS may have APOL1 risk alleles • These can be assayed in renal biopsy tissue

DIFFERENTIAL DIAGNOSIS Recurrent Focal Segmental Glomerulosclerosis • Usually presents earlier (< 1 year)

Calcineurin Inhibitor Toxicity • Isometric vacuolization of tubular epithelial cells • Adventitial hyaline nodules

Chronic Transplant Glomerulopathy • Duplication of GBM without immune complex deposition • Variable C4d peritubular capillary deposition • May occur concurrently with de novo FSGS

Treatment

Immune Complex-Mediated Glomerulonephritis, De Novo or Recurrent

• Not well defined • If CNI implicated, alter drug regimen

• Positive immunofluorescence staining for immunoglobulins (immune complex deposition)

Prognosis • De novo FSGS ○ 40% graft loss within 5 years of diagnosis • De novo CG ○ 50% loss of graft within 1 year of diagnosis

MICROSCOPIC Histologic Features • Segmental glomerulosclerosis with any of following findings ○ Synechial or fibrous attachments to Bowman capsules ○ Obliteration of glomerular capillary lumina by foam cells &/or hyaline ○ Segmental accumulation of mesangial matrix ○ Prominence of visceral epithelial cells (podocytes) ○ Protein reabsorption droplets in podocytes ○ Foot process effacement • CG (variant of FSGS) ○ Collapse of glomerulus with podocyte hypercellularity ○ Associated with severe vascular disease and CNI toxicity

Kidney Transplantation

TERMINOLOGY

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • FSGS should trigger immunofluorescence and electron microscopy work-up • CG may mimic crescentic glomerulonephritis

SELECTED REFERENCES 1.

2. 3.

4. 5.

6.

Shah PB et al: APOL1 polymorphisms in a deceased donor and early presentation of collapsing glomerulopathy and focal segmental glomerulosclerosis in two recipients. Am J Transplant. 16(6):1923-1927, 2016 Ponticelli C et al: De novo glomerular diseases after renal transplantation. Clin J Am Soc Nephrol. 9(8):1479-87, 2014 Ikeda Y et al: A case of de novo focal segmental glomerulosclerosis occurred one and half years after kidney transplantation supposed to be caused by calcineurin inhibitor. Clin Transplant. 26 Suppl 24:76-80, 2012 Nadasdy T et al: Zonal distribution of glomerular collapse in renal allografts: possible role of vascular changes. Hum Pathol. 33(4):437-41, 2002 Cosio FG et al: Focal segmental glomerulosclerosis in renal allografts with chronic nephropathy: implications for graft survival. Am J Kidney Dis. 34(4):731-8, 1999 Meehan SM et al: De novo collapsing glomerulopathy in renal allografts. Transplantation. 65(9):1192-7, 1998

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De Novo Membranous Glomerulonephritis KEY FACTS

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• May represent unusual manifestation of chronic antibodymediated rejection ○ Associated with C4d peritubular capillary deposition and anti-HLA-DQ ○ 1 autopsy showed de novo membranous glomerulonephritis (MGN) involving only kidney allograft and not native kidneys ○ May recur in subsequent renal allografts

• Glomerular basement membrane (GBM) thickening ○ Diffuse or segmental ○ GBM thickening "spike" formation – ± swiss cheese appearance depending on MGN stage • Mesangial hypercellularity in 1/3 of cases • Granular IgG staining in glomerular capillary walls ○ C4d and C3 may stain in similar pattern ○ Careful evaluation of C4d glomerular staining pattern if only antibody tested in transplant kidney biopsies ○ C4d in peritubular capillaries found in ~ 70% of cases • Subepithelial electron-dense deposits seen by EM ± basement membrane "spike" formation

CLINICAL ISSUES • 0.5-9.0% of kidney transplant patients ○ Manifests late (> 3 years) ○ Proteinuria ○ Renal dysfunction ○ Unfavorable prognosis – 67% require renal replacement therapy

TOP DIFFERENTIAL DIAGNOSES • Recurrent MGN • Donor-derived MGN • Chronic transplant glomerulopathy

Subepithelial "Spike" Formation

Duplication of Glomerular Basement Membrane

IgG

Subepithelial Deposits

(Left) Jones methenamine silver shows a hint of subepithelial "spike" formation ﬈ along some of the glomerular basement membranes (GBMs). (Right) Chronic transplant glomerulopathy or duplication of the GBMs ﬈ can affect 50% of de novo membranous glomerulonephritis (MGN), as demonstrated in this glomerulus from a pediatric patient with renal transplant due to renal dysplasia (Jones methenamine silver).

(Left) IF microscopy for IgG demonstrates granular staining of all glomerular capillaries. Clinical correlation is necessary to confirm whether this represents recurrent or de novo MGN. (Right) EM of a glomerulus with de novo MGN shows very small deposits in contact with the podocyte ﬈ with only minimal "spike" formation, typical of early and mild MGN. The patient was transplanted 4 years previously for diabetic nephropathy.

162

De Novo Membranous Glomerulonephritis • Intimal arteritis ○ Acute (type 2) rejection found in subset

Abbreviations

ANCILLARY TESTS

• Membranous glomerulonephritis (MGN), de novo

Definitions

Immunofluorescence

• MGN in kidney allograft when primary cause of end-stage renal disease is not MGN

• Positive granular capillary wall staining for IgG, κ-, and λlight chains ○ IgG1 predominant subclass – IgG4 predominant subclass in primary MGN ○ Variable capillary wall staining for C4d, C3, C1q, and IgM • C4d(+) peritubular capillaries in ~ 70%

ETIOLOGY/PATHOGENESIS Allo-/Autoantibody • May represent unusual manifestation of chronic antibodymediated (humoral) rejection ○ Associated with C4d peritubular capillary deposition and anti-HLA-DQ ○ 1 autopsy showed de novo MGN involving only kidney allograft without MGN in native kidneys – May recur in subsequent renal allografts ○ 1 de novo MGN case with donor-specific antibodies against HLA-DQ7 • Can occur in HLA identical grafts, presumably due to nonHLA antigen ○ Rat model of de novo MGN occurs only in transplant, not native kidney • No autoantibodies to PLA2R1

CLINICAL ISSUES

Electron Microscopy • Subepithelial amorphous electron-dense deposits ○ Often small and relatively sparse ○ Stage I (Ehrenreich-Churg) deposits common • Duplication of GBMs ○ Subendothelial space widening when injured endothelial cells detach from GBM

DIFFERENTIAL DIAGNOSIS Recurrent Membranous Glomerulonephritis • Clinical history of MGN • Earlier onset (< 3 months) • IgG4 predominant

Donor-Derived Membranous Glomerulonephritis

Epidemiology

• Present in donor biopsy, disappears in months

• Incidence ○ 0.5-9.0% of kidney transplant patients

Chronic Transplant Glomerulopathy

Presentation • Manifests late (> 3 years) • Proteinuria ○ 2nd most common cause in renal allograft patients ○ Often nephrotic range (> 3 g/24 hours), may be intermittent or persistent

Prognosis • 5-year graft loss in > 50% • 67% progress to renal failure

MACROSCOPIC General Features • Renal vein thrombosis occasional present ○ Less common than idiopathic MGN in native kidneys

• • • •

Duplication of GBM without immune complexes Occurs concurrently in 50% of de novo MGN cases C4d(+) in peritubular capillaries Multilamination of peritubular capillaries on EM

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Significant proteinuria should trigger additional evaluation by IF &/or EM • Thickening or duplication of GBMs or segmental glomerular sclerosis should trigger additional IF &/or EM • C4d immunofluorescence microscopy may reveal granular glomerular capillary wall staining

SELECTED REFERENCES 1.

MICROSCOPIC Histologic Features • Glomerular basement membrane (GBM) thickening ○ Focal &/or segmental thickening common • Glomerular capillaritis in ~ 50% • Mesangial hypercellularity in ~ 33% • Double contours or duplication of GBM in 50% ○ Possibly due to concurrent chronic transplant glomerulopathy (chronic antibody-mediated rejection) • Prominent interstitial inflammation ○ Often sufficient for diagnosis of acute (T-cell-mediated) rejection

Kidney Transplantation

TERMINOLOGY

2. 3.

4. 5. 6.

7.

Wen J et al: HLA-DR, and not PLA2R, is expressed on the podocytes in kidney allografts in de novo membranous nephropathy. Medicine (Baltimore). 95(37):e4809, 2016 Ponticelli C et al: De novo glomerular diseases after renal transplantation. Clin J Am Soc Nephrol. 9(8):1479-87, 2014 Patel K et al: De novo membranous nephropathy in renal allograft associated with antibody-mediated rejection and review of the literature. Transplant Proc. 45(9):3424-8, 2013 Honda K et al: De novo membranous nephropathy and antibody-mediated rejection in transplanted kidney. Clin Transplant. 25(2):191-200, 2011 Lal SM: De novo membranous nephropathy in renal allografts with unusual histology. Arch Pathol Lab Med. 131(1):17, 2007 Monga G et al: Membranous glomerulonephritis (MGN) in transplanted kidneys: morphologic investigation on 256 renal allografts. Mod Pathol. 6(3):249-58, 1993 Truong L et al: De novo membranous glomerulonephropathy in renal allografts: a report of ten cases and review of the literature. Am J Kidney Dis. 14(2):131-44, 1989

163

Kidney Transplantation

De Novo Membranous Glomerulonephritis

Chronic Transplant Glomerulopathy

Segmental Sclerosis

Segmental IgG

C4d

C4d Peritubular Capillary Deposition

C4d Immunohistochemistry

(Left) Glomerulus with de novo MGN and chronic antibodymediated rejection shows prominent and widespread duplication of the GBM. (Right) PAS of a biopsy with de novo MGN reveals thick GBMs with a vacuolated appearance ﬊ in a glomerulus with segmental accumulation of matrix and hyaline ﬈, which obscures the capillary lumina.

(Left) IF for IgG highlights the segmental distribution of immune complex deposition along the glomerular capillaries ﬇. Several glomerular capillaries or segments of GBM show no significant staining ſt. A similar pattern, but less intense staining, was also seen with κ- and λ-light chains. (Right) IF shows granular C4d staining along the GBMs, which may be the only hint of MGN and should trigger additional IF and EM studies.

(Left) C4d deposition is present in the peritubular capillaries in a patient with donor-reactive HLA antibodies and de novo MGN. The peritubular capillaries have widespread linear deposits of C4d, typical of antibodymediated rejection ſt. (Right) C4d deposits are detected in the GBM and in peritubular capillaries in this patient with de novo MGN in a 2nd transplant. The 1st graft was lost to de novo MGN and chronic antibody-mediated rejection. Prior de novo MGN is a risk factor for a 2nd episode in a subsequent allograft.

164

De Novo Membranous Glomerulonephritis

Subepithelial "Spikes" (Left) EM reveals segmental distribution of many small, subepithelial, electron-dense deposits ﬈. The podocyte foot processes demonstrate diffuse effacement ﬊. (Right) EM of a glomerulus with de novo MGN shows very small deposits in contact with the podocyte ſt with only minimal "spike" formation, typical of early and mild MGN. This pattern is not uncommon in de novo MGN.

Segmental Distribution of "Spikes"

Kidney Transplantation

Subepithelial Immune Complexes

Subepithelial Deposits (Left) EM reveals many discrete, electron-dense deposits in subepithelial locations. Focal separation of the endothelial cell from the GBM ﬈ is noted, but duplication of the GBM is not apparent in this or other glomerular capillaries. (Right) EM of a patient with de novo MGN from a patient transplanted 6 years ago for focal segmental glomerulosclerosis shows subepithelial deposits ﬉ surrounded by GBM "spikes" ﬈, typical of stage II MGN.

Subepithelial and Intramembranous Deposits

Peritubular Capillary Basement Membrane Multilayering (Left) EM of a glomerulus from a patient with both chronic antibody-mediated rejection and de novo MGN shows duplication of the GBM and amorphous, electron-dense deposits within the GBM ﬈ and in subepithelial locations ﬉. (Right) EM shows multilamination of the peritubular capillary basement membrane ﬈ in a patient with donor-reactive HLA antibodies and de novo MGN.

165

Kidney Transplantation

Anti-GBM Disease in Alport Syndrome KEY FACTS

TERMINOLOGY • Alport posttransplant nephritis

ETIOLOGY/PATHOGENESIS • Antibodies develop against α-3, α-4, or α-5 chains of collagen IV of transplant kidney ○ Larger COL4A5 deletions may be more susceptible to posttransplant antiglomerular basement membrane (anti-GBM) disease • X-linked AS patients target NC1 domain of COL4A5 • Autosomal recessive AS patients target NC1 domain of COL4A3 or COL4A4

CLINICAL ISSUES • 3-5% of Alport recipients reported to develop de novo antiGBM after kidney transplantation • 0.4% of AS patients develop de novo anti-GBM disease in recent series (2014) • Acute renal failure

○ 75% of cases occur in 1st posttransplant year • Hematuria • Male predominance • 90% of grafts fail within months after onset ○ Anti-GBM disease often recurs in subsequent allografts with accelerated course • Plasmapheresis

MICROSCOPIC • Cellular crescents &/or fibrinoid necrosis, typically > 80% of glomeruli • Strong linear IgG immunofluorescence staining of GBM • Intratubular red blood cell casts • Acute tubular injury

TOP DIFFERENTIAL DIAGNOSES • Pauci-immune (ANCA-associated) crescentic glomerulonephritis • Diabetic nephropathy

Transplant Nephrectomy

Red Blood Cell Casts

Cellular Crescent

Linear IgG Deposition

(Left) Gross photograph shows an allograft nephrectomy from a 37-year-old woman with ESRD due to X-linked Alport syndrome. Eighteen months post transplant, the patient presented with allograft tenderness and renal failure. Anti-GBM titers were equivocal by ELISA and western blot. (Right) H&E shows several red blood cell and pigmented casts ﬈ in the cortex, widespread interstitial fibrosis, and tubular injury in this Alport patient with antiGBM nephritis after kidney transplantation.

(Left) H&E-stained tissue section from a patient with de novo anti-GBM disease 18 months after transplantation for Alport syndrome shows a cellular crescent ﬈ compressing the glomerulus. (Right) Immunofluorescence microscopy of a glomerulus with de novo anti-GBM disease from this Alport syndrome patient after transplantation shows bright linear staining of the GBM for IgG. The excess IgG staining vs. albumin is typical of anti-GBM nephritis.

166

Anti-GBM Disease in Alport Syndrome

Abbreviations • Antiglomerular basement membrane (anti-GBM) disease in Alport syndrome (AS)

Synonyms • Alport posttransplant nephritis • De novo anti-GBM nephritis

Prognosis • 90% of kidney allografts fail within weeks to months after onset of anti-GBM disease ○ Anti-GBM disease often recurs in subsequent allografts with accelerated course • Overall graft and patient survival after transplant similar to non-Alport recipients

MICROSCOPIC

Definitions

Histologic Features

• Glomerulonephritis (GN) mediated by anti-GBM antibodies in AS renal transplant recipients

• Cellular crescents &/or fibrinoid necrosis (typically > 80% of glomeruli) ○ Glomerular tufts not involved by crescent formation or fibrinoid necrosis appear normal • Acute tubular injury • Red blood cell casts • T cell-mediated or antibody-mediated rejection may also be present

ETIOLOGY/PATHOGENESIS Exposure to Nonendogenous Glomerular Basement Membrane Antigens After Kidney Transplantation • Antibodies develop against NC1 domains of intact α345NC1 (hexamer of α-3, α-4, or α-5 chains of collagen IV) of allograft in AS patients due to mutated endogenous antigen ○ Larger deletions of COL4A5 at higher risk for GN ○ Target quaternary epitopes of intact α345NC1 – Target alloantigen of all alloantibodies that mediate de novo anti-GBM nephritis – Distinct antigenic target from Goodpasture syndrome – X-linked AS target NC1 domain of COL4A5 – Autosomal recessive AS patients target NC1 domain of COL4A3 or COL4A4

CLINICAL ISSUES Epidemiology • Incidence ○ De novo anti-GBM disease reported after kidney transplantation in 3-5% of Alport recipients in older series ○ 0.4% of AS patients developed clinical de novo anti-GBM disease in 2014 series • Sex ○ Male predominance ○ Females with X-linked AS have 1 normal allele of α chains of collagen IV and rarely develop anti-GBM disease – Only 2 reports involving females with autosomal recessive AS

Presentation • Acute renal failure ○ 75% within 1st year after kidney transplantation • Hematuria

Laboratory Tests

ANCILLARY TESTS Immunofluorescence • Strong, linear IgG immunofluorescence staining of GBMs ○ Observed in up to 15% of AS patients after kidney transplantation ○ Kappa and λ-light chains and often C3 stain GBMs in similar pattern with lower intensity ○ Intensity of linear IgG staining > > > albumin ○ Bowman capsules and distal tubular basement membranes may also show linear staining

DIFFERENTIAL DIAGNOSIS Pauci-Immune (ANCA-Associated) Crescentic GN • Lacks strong linear immunoglobulin deposition in GBM

Diabetic Nephropathy • Strong linear IgG and albumin IF staining of GBM

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Linear IgG GBM staining may be present without nephritis

SELECTED REFERENCES 1. 2.

3. 4.

• Serum anti-GBM assay by western blot or ELISA

Treatment • Drugs ○ Cyclophosphamide ○ High-dose corticosteroids ○ Bortezomib (experimental) ○ Rituximab (experimental) • Plasmapheresis

Kidney Transplantation

TERMINOLOGY

5.

6. 7. 8. 9.

Kelly YP et al: Outcomes of kidney transplantation in Alport syndrome compared with other forms of renal disease. Ren Fail. 1-4, 2016 Mallett A et al: End-stage kidney disease due to Alport syndrome: outcomes in 296 consecutive Australia and New Zealand Dialysis and Transplant Registry cases. Nephrol Dial Transplant. 29(12):2277-86, 2014 Olaru F et al: Quaternary epitopes of α345(IV) collagen initiate Alport posttransplant anti-GBM nephritis. J Am Soc Nephrol. 24(6):889-95, 2013 Temme J et al: Outcomes of male patients with Alport syndrome undergoing renal replacement therapy. Clin J Am Soc Nephrol. 7(12):196976, 2012 de Sandes-Freitas TV et al: Late presentation of Alport posttransplantation anti-glomerular basement membrane disease. Transplant Proc. 43(10):40001, 2011 Pedchenko V et al: Molecular architecture of the Goodpasture autoantigen in anti-GBM nephritis. N Engl J Med. 363(4):343-54, 2010 Kashtan CE: Renal transplantation in patients with Alport syndrome. Pediatr Transplant. 10(6):651-7, 2006 Browne G et al: Retransplantation in Alport post-transplant anti-GBM disease. Kidney Int. 65(2):675-81, 2004 Byrne MC et al: Renal transplant in patients with Alport's syndrome. Am J Kidney Dis. 39(4):769-75, 2002

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Kidney Transplantation

Hyperperfusion Injury KEY FACTS

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• 10% of pediatric kidneys (< 10 years of age) develop this injury ○ Infant kidneys (< 1 year of age) more susceptible ○ En bloc (2 kidneys) transplantation may be less susceptible • Possible contributing factors ○ Higher blood volume and systemic blood pressure in adults compared with children ○ Smaller glomeruli and thinner glomerular basement membrane (GBM) thickness in children compared to adults ○ Altered composition of α chains of collagen IV in pediatric glomeruli

• Global &/or segmental glomerulosclerosis • Variable mesangial sclerosis &/or hypercellularity • Interstitial fibrosis and tubular atrophy

CLINICAL ISSUES

ANCILLARY TESTS • Immunofluorescence microscopy ○ Indirect IF for α-3 and α-5 chains of collagen IV demonstrates strong linear staining of GBM (normal) • Electron microscopy ○ Prominent GBM alterations – Diffuse lamellation and splitting – Marked thinning ○ Foot process effacement and microvillous transformation of podocytes

TOP DIFFERENTIAL DIAGNOSES

• Renal dysfunction • Proteinuria

• Alport syndrome • Recurrent focal segmental glomerulosclerosis

Glomerular and Tubulointerstitial Scarring

Mesangial Sclerosis/Prominent Podocytes

Segmental Sclerosis

Prominent Glomerular Basement Membrane Alterations

(Left) PAS shows 2 globally sclerotic glomeruli ﬈ surrounded by severe and diffuse interstitial fibrosis and tubular atrophy in this pediatric kidney allograft. (Courtesy T. Nadasdy, MD.) (Right) Jones methenamine silver stain shows mesangial sclerosis ﬈ with prominence of the adjacent visceral epithelial cells ﬊. There is also segmental mesangial hypercellularity. (Courtesy T. Nadasdy, MD.)

(Left) Jones methenamine silver stain shows a segmentally sclerotic glomerulus with accumulation of the matrix, loss of capillary lumina, and fibrous adhesions ſt to the Bowman capsule. (Courtesy T. Nadasdy, MD.) (Right) Electron micrograph shows prominent lamellation and splitting ﬈ of the glomerular basement membrane with diffuse effacement of the overlying podocyte foot processes. (Courtesy T. Nadasdy, MD.)

168

Hyperperfusion Injury

Synonyms

○ Variable prominence of podocytes • Tubulointerstitium ○ Interstitial fibrosis and tubular atrophy

• Hyperfiltration injury

Definitions • Acute and chronic glomerular injury caused by transplantation of very young pediatric kidneys into adult recipients

ETIOLOGY/PATHOGENESIS Hyperperfusion Injury • 10% of pediatric kidneys (< 10 years of age) develop hyperperfusion injury when transplanted into adult recipients ○ Infant kidneys (< 1 year of age) more susceptible ○ En bloc (2 kidneys) transplantation may be less susceptible • Possible contributing factors ○ Greater blood volume in adults compared with children ○ Higher systemic blood pressure in adults compared with children ○ Smaller glomeruli in children compared to adults ○ Thinner glomerular basement membrane (GBM) in children compared to adults – GBM thickness reaches maximum thickness at age 1213 ○ Altered composition of α chains of collagen IV in pediatric glomeruli – Embryonic GBM consists of α-1 and α-2 chains of collagen IV, gradually replaced by α-3, α-4, and α-5 chains as glomerulus matures

CLINICAL ISSUES Epidemiology • Incidence ○ Rare

Presentation • Renal dysfunction • Proteinuria ○ Often nephrotic range • May develop as early as 10 weeks after transplantation

ANCILLARY TESTS Immunofluorescence • Indirect IF for α-3 and α-5 chains of collagen IV demonstrates strong linear staining of GBM (normal)

Electron Microscopy • Prominent GBM alterations ○ Diffuse lamellation and splitting ○ Marked thinning • Podocyte injury ○ Diffuse foot process effacement ○ Microvillous transformation

DIFFERENTIAL DIAGNOSIS Alport Syndrome • Full-thickness diffuse lamellation and splitting of GBM without normal segments by EM • Indirect IF shows absence of GBM α-3 &/or α-5 chains of collagen IV • Extensive donor work-up minimizes this clinical scenario

Recurrent Focal Segmental Glomerulosclerosis • History of primary focal segmental glomerulosclerosis as original native renal disease • Diffuse effacement of podocyte foot processes • Normal GBM by EM

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Clinical information of kidney donor needed to raise consideration of hyperperfusion injury • EM essential for diagnosis • Without EM finding of GBM alterations, light microscopic features of glomerular and tubulointerstitial scarring are nonspecific

SELECTED REFERENCES 1.

Laboratory Tests • None ○ Kidney biopsy with electron microscopy only method to establish diagnosis

Prognosis • Higher vascular complication rate • Poor ○ Infant donor kidneys often fail within 1 year of transplantation

Kidney Transplantation

TERMINOLOGY

2. 3.

4.

5.

6.

MICROSCOPIC

7.

Histologic Features

8.

• Glomerulus ○ Global &/or segmental glomerulosclerosis – Variable mesangial sclerosis &/or hypercellularity

9.

Al-Shraideh Y et al: Single vs dual (en bloc) kidney transplants from donors ≤ 5 years of age: a single center experience. World J Transplant. 6(1):239-48, 2016 Feltran Lde S et al: Does graft mass impact on pediatric kidney transplant outcomes? Pediatr Nephrol. 29(2):297-304, 2014 Friedersdorff F et al: Outcome of single pediatric deceased donor renal transplantation to adult kidney transplant recipients. Urol Int. 92(3):323-7, 2014 Borboroglu PG et al: Solitary renal allografts from pediatric cadaver donors less than 2 years of age transplanted into adult recipients. Transplantation. 77(5):698-702, 2004 Nadasdy T et al: Diffuse glomerular basement membrane lamellation in renal allografts from pediatric donors to adult recipients. Am J Surg Pathol. 23(4):437-42, 1999 Ratner LE et al: Transplantation of single and paired pediatric kidneys into adult recipients. J Am Coll Surg. 185(5):437-45, 1997 Satterthwaite R et al: Outcome of en bloc and single kidney transplantation from very young cadaveric donors. Transplantation. 63(10):1405-10, 1997 Hayes JM et al: The development of proteinuria and focal-segmental glomerulosclerosis in recipients of pediatric donor kidneys. Transplantation. 52(5):813-7, 1991 Truong LD et al: Electron microscopic study of an unusual posttransplant glomerular lesion. Arch Pathol Lab Med. 115(4):382-5, 1991

169

Kidney Transplantation

Engraftment Syndrome KEY FACTS

TERMINOLOGY

ANCILLARY TESTS

• Systemic syndrome described in association with hematopoietic cell transplantation ○ Primarily observed in combined nonmyeloablative bone marrow transplant (BMT)/kidney transplant recipients ○ Seen in variety of autologous or allogeneic transplants • Synonyms: Capillary leak syndrome, marrow activation, or recovery syndrome

• Ki-67(+) in many cells in PTCs • Cells include T cells [CD3(+), CD8(+)], macrophages [CD68(+)], & granulocytes [MPO(+)] • Cells in capillaries of recipient origin by XY FISH • EM shows severe damage to PTC endothelium & fibrin tactoids • C4d usually negative

CLINICAL ISSUES

TOP DIFFERENTIAL DIAGNOSES

• Renal dysfunction, fever, pulmonary edema, rash • Recent regimens have less pronounced engraftment syndrome (ES)

• • • •

MICROSCOPIC • Acute tubular injury (ATI) • Interstitial hemorrhage • Congestion of peritubular capillaries (PTCs)

ATI due to ischemia Acute antibody-mediated (humoral) rejection Acute T-cell-mediated rejection Thrombotic microangiopathy

DIAGNOSTIC CHECKLIST • Ki-67(+) cells in glomeruli & PTCs distinguish ES from rejection & ATI

Tubular Injury, Mitotic Activity, and Peritubular Capillary Cells

Glomerular Capillary Loop Inflammatory Cells

Erythrocyte Stasis in Peritubular Capillaries

Ki-67(+) Cells in Glomeruli and Peritubular Capillaries

(Left) PAS stain of a simultaneous bone marrow/kidney transplant patient shows loss of brush borders, flattened epithelial cells, cells in peritubular capillaries (PTCs) ﬊, and tubular mitoses ﬈. (Right) Light microscopy of a simultaneous bone marrow transplant/kidney transplant tolerance protocol patient shows a slightly hypercellular glomerulus with occasional cells in glomerular capillary loops ſt.

(Left) EM of a simultaneous bone marrow/kidney recipient in a protocol devised to induce tolerance shows "sludging/stasis" of erythrocytes in the PTCs and an attenuated PTC endothelium ſt. (Right) Ki-67 stain of a bone marrow/kidney transplant recipient in a protocol devised to induce tolerance shows numerous positive cells in PTCs and glomeruli ﬈.

170

Engraftment Syndrome

Abbreviations • Engraftment syndrome (ES)

Definitions • Systemic inflammatory syndrome described after bone marrow transplant (BMT)

ETIOLOGY/PATHOGENESIS Clinical Setting • Autologous or allogeneic BMT ○ Usually no more than mild renal impairment • More severe forms occur in BMT + kidney recipients ○ Often in non-HLA identical renal-BMT allografts – Also in HLA identical graft, suggesting non-MHC antigens can be target

Postulated Mechanisms • • • • •

Cytokine release during recipient bone marrow recovery Auto-/alloreactivity to endothelium Homeostatic proliferation of lymphocytes ↑ vascular sensitivity to calcineurin inhibitors Cyclophosphamide may contribute

CLINICAL ISSUES Epidemiology • Incidence ○ Renal dysfunction (~ 20% of cases) in which BMT, either autologous or allogenic, is only procedure – Occurs during recovery of donor marrow – Biopsies not typically performed to explain renal dysfunction in isolated BMT ○ ES common in nonmyeloablative BMT combined with renal transplant in early studies – Occurs during recovery of recipient marrow, as chimerism declines – Omission of cyclophosphamide may ↓ incidence

Presentation • Acute renal failure/insufficiency ○ 10-12 days after combined BMT/kidney transplant • Fever, pulmonary edema (noncardiogenic), rash

Treatment • Supportive, sometimes ↑ or ↓ in immunosuppression

Prognosis • Typically transient

MICROSCOPIC Histologic Features • Glomeruli ○ Glomerular capillary mononuclear cells & neutrophils ○ No thrombi or hypercellularity • Tubules ○ Acute tubular injury (ATI) • Peritubular capillaries (PTCs) ○ Congestion & occasional inflammatory cells, including neutrophils & mononuclear cells

• Interstitium ○ Interstitial hemorrhage, focal ○ Little or no interstitial infiltrate • Arteries ○ Usually normal ○ Endarteritis in minority of cases ○ Thrombi not conspicuous

ANCILLARY TESTS

Kidney Transplantation

TERMINOLOGY

Immunohistochemistry • PTCs ○ Ki-67 shows numerous positive cells in PTCs & glomeruli ○ Intracapillary cells positive for CD3, CD68, & MPO ○ T cells are frequently CD8(+) with rare CD4(+) T cells • Glomeruli ○ Similar to PTC: Ki-67(+), CD3(+), CD8(+), & CD68(+) cells

Immunofluorescence • C4d usually negative ○ C4d(+) in association with donor-reactive antibodies

In Situ Hybridization • XY FISH shows intracapillary cells of recipient origin • Endothelial cells of donor origin

Electron Microscopy • Severe PTC endothelial damage & loss • Fibrin tactoids can be seen in many PTCs

DIFFERENTIAL DIAGNOSIS ATI From Other Causes • ATI from other causes, such as ischemia, typically have fewer Ki-67(+) cells in PTC

Acute Antibody-Mediated Rejection • C4d in PTCs by immunofluorescence or immunohistochemistry in antibody-mediated rejection • Neutrophils in PTCs & more prominent C4d positivity in some cases of ES in combined BMT-kidney transplant • No fibrinoid necrosis of arteries in ES

Acute T-Cell-Mediated Rejection • Endarteritis & tubulointerstitial infiltrates

Thrombotic Microangiopathy • Thrombi in small blood vessels, sometimes accompanied by endothelial swelling, edema, &/or fibrinoid necrosis • Schistocytes (fragmented red blood cells), particularly in walls of blood vessels • Electron microscopy shows subendothelial widening &/or lucencies

SELECTED REFERENCES 1. 2.

3.

4.

Shah NN et al: Procalcitonin and cytokine profiles in engraftment syndrome in pediatric stem cell transplantation. Pediatr Blood Cancer. 64(3), 2016 Kawai T et al: Long-term results in recipients of combined HLA-mismatched kidney and bone marrow transplantation without maintenance immunosuppression. Am J Transplant. 14(7):1599-611, 2014 Farris AB et al: Acute renal endothelial injury during marrow recovery in a cohort of combined kidney and bone marrow allografts. Am J Transplant. 11(7):1464-77, 2011 Troxell ML et al: Renal pathology in hematopoietic cell transplantation recipients. Mod Pathol. 21(4):396-406, 2008

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Engraftment Syndrome Erythrocyte Stasis in Peritubular Capillaries

Erythrocyte Stasis in Peritubular Capillaries

Focal Endarteritis During Engraftment Syndrome Episode

CD3(+) Cells in Glomeruli and Peritubular Capillaries

CD68(+) Cells in Peritubular Capillaries

CD34 Stains Peritubular Capillary Endothelium and Cells

(Left) Trichrome stain of a patient undergoing simultaneous bone marrow/kidney transplant in a protocol devised to induce tolerance shows widespread erythrocyte stasis ſt. (Right) Trichrome stain of a bone marrow/kidney allograft recipient in a tolerance protocol shows erythrocyte stasis with occasional cells that can be appreciated in PTCs by light microscopy ſt.

(Left) Focal endarteritis ﬈ is seen in a small artery from a bone marrow/kidney recipient in a tolerance protocol biopsied 10 days post transplant during an episode of engraftment syndrome (ES). It is unclear whether this represents cellular rejection or a manifestation of the endothelial injury from ES. (Right) CD3(+) cells in glomeruli ﬈, PTCs ﬊, and a focus of tubulitis ſt can be seen by immunohistochemistry in a bone marrow/kidney transplant recipient in a tolerance protocol.

(Left) CD68 stain shows numerous positive cells in PTCs ſt in a bone marrow/kidney transplant recipient in a protocol devised to induce tolerance. (Right) CD34 stain of a bone marrow/kidney transplant recipient in a protocol devised to induce tolerance shows CD34(+) cells in PTCs ﬈, possibly immature bone marrow cells, and patchy loss of CD34 staining of PTC endothelium ſt, suggesting endothelial damage.

172

Engraftment Syndrome Red Blood Cell Stasis in Peritubular Capillaries (Left) EM of a PTC in a bone marrow/kidney transplant recipient in a protocol devised to induce tolerance shows fibrin tactoids ﬈ and a reactive endothelium ﬈. (Right) EM of a simultaneous bone marrow/kidney transplant recipient in a protocol devised to induce tolerance shows a PTC engorged with erythrocytes with a denuded endothelium ﬈.

Erythrocytes in Glomerular Capillaries

Kidney Transplantation

Peritubular Capillary Fibrin

Leukocytes in Graft of Recipient Origin (Left) EM in a tolerance protocol patient status post simultaneous bone marrow/kidney transplant shows that glomerular capillaries are engorged by erythrocytes, and that there is focal thinning of the glomerular basement membrane ﬈. (Right) XY FISH in a female tolerance protocol recipient of a male renal and bone marrow allograft shows that CD45(+) cells in PTC are from the recipient ſt (green X chromosome, red Y chromosome, yellow CD45, blue DAPI). A mitotic figure can be seen ﬇.

Tubular Cells of Donor Origin

Recipient T Cells in Graft (Left) XY FISH in a female tolerance protocol recipient of a male renal and bone marrow allograft shows recipient CD45(+) cells ſt in PTCs. Donor tubular epithelial cells can be appreciated ﬇ (green X, red Y, yellow CD45, blue DAPI). (Right) XY FISH in an HLA-identical female recipient developed ES 28 days after receiving an HLA-identical male kidney and bone marrow transplant (X green, Y red, CD3 orange). About 75% of the T cells in the graft were of recipient origin ſt, consistent with TCMR to non-MHC antigens.

173

Kidney Transplantation

Kidney Diseases in Nonrenal Transplant Recipients KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Renal disease developing in native kidneys after transplantation of other organs or hematopoietic cells

• Varies with cause; small percentage biopsied (1-4%) ○ CNIT: Chronic, vascular, and tubulointerstitial – Nodular arteriolar hyalinosis, interstitial fibrosis, tubular atrophy, glomerulosclerosis ○ CNIT: Thrombotic microangiopathy – GBM duplication, thrombi endothelial injury ○ Membranous glomerulonephritis – GBM deposits and "spikes" along GBM ○ Minimal change disease – Foot process effacement ○ Drug toxicity – Crystals, oxalates, interstitial nephritis ○ Viral infection – Nuclear inclusions, positive viral antigens by IHC

ETIOLOGY/PATHOGENESIS • • • • • •

Calcineurin inhibitors (cyclosporine, tacrolimus) Antiviral drugs Polyomavirus Diabetic nephropathy Hypertensive nephrosclerosis Associated with graft-vs.-host disease in hematopoietic transplants

CLINICAL ISSUES • 16% of recipients of nonkidney organ transplants develop chronic renal failure in 10 years • Progressive renal failure • Proteinuria, nephrotic syndrome • Schistocytes, thrombocytopenia, elevated LDH

DIAGNOSTIC CHECKLIST • Cause for most cases assumed to be CNIT but without biopsy evidence

Peripheral Nodular Hyaline of CNIT

Striped Fibrosis Due to CNIT

Onion Skin Intimal Thickening of TMA

Subclinical TMA

(Left) Peripheral nodular hyaline in arterioles st is characteristic, although not pathognomonic, of calcineurin inhibitor toxicity (CNIT). This is a native kidney biopsy from a heart-lung transplant recipient. (Right) Fibrosis appears "striped" because it is along the medullary rays in the cortex ﬈, a watershed area prone to ischemia from many causes. This is a native kidney from a lung transplant recipient. (Courtesy S. Rosen, MD.)

(Left) Native kidney biopsy from a recipient of a heart transplant 13 years previously shows onion skin thickening of an arteriole st, a manifestation of thrombotic microangiopathy (TMA). (Right) Native kidney biopsy from a recipient of a liver transplant 5 years previously shows marked glomerular endothelial reaction manifested by loss of fenestrations ﬈ and irregularity along the GBM. A fibrin tactoid is in the lumen ﬉.

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Kidney Diseases in Nonrenal Transplant Recipients

Abbreviations • Calcineurin inhibitor (CNI)

Definitions • Renal disease developing in native kidneys after transplantation of other organs or hematopoietic cells

ETIOLOGY/PATHOGENESIS

○ Minimal change disease ○ HCV-related glomerular disease • Schistocytes, thrombocytopenia, elevated LDH ○ CNI toxicity ○ May also be subclinical thrombotic microangiopathy

Treatment • Varies with cause • Reduction or alternation of immunosuppressive therapy • Renal transplantation has been successful

Drug Toxicity

Prognosis

• CNIs (cyclosporine, tacrolimus) ○ Thrombotic microangiopathy ○ Interstitial fibrosis/tubular atrophy ○ Arteriolar hyalinosis ○ Focal segmental glomerulosclerosis ○ Acute tubular injury ○ Therapeutic levels usually much higher than kidney transplantation • Antiviral drugs • Hydroxyethyl starch nephrotoxicity

• Varies with cause • 4.5% develop ESRD overall

Infection • Polyomavirus (BK/JC) ○ Polyoma nephropathy occurs rarely in heart, liver, lung, or pancreas recipients • Epstein-Barr virus (posttransplant lymphoproliferative disease) • Adenovirus • Cytomegalovirus

Progression of Recipient Disease • • • • •

Diabetic nephropathy Hypertensive nephrosclerosis HCV-related glomerular disease Myeloma cast nephropathy Amyloidosis

Immunologic Reaction • Membranous glomerulonephritis ○ Associated with graft-vs.-host disease (GVHD) – Rare report with autologous stem cell transplantation • Minimal change disease ○ Associated with GVHD

CLINICAL ISSUES Epidemiology • 16.5% of recipients of nonkidney organ transplants develop chronic renal failure in 10 years (GFR < 30 mL/m1.73m²) ○ At 5 years: 18% of liver recipients, 16% of lung recipients, 11% of heart recipients

Presentation • Progressive renal failure ○ CNI toxicity ○ Hypertensive nephrosclerosis ○ Polyomavirus nephropathy • Proteinuria, nephrotic syndrome ○ Diabetic nephropathy ○ Focal segmental glomerulosclerosis ○ Membranous glomerulonephritis

Kidney Transplantation

TERMINOLOGY

MICROSCOPIC Histologic Features • CNI toxicity ○ Interstitial fibrosis, tubular atrophy, global or segmental glomerulosclerosis, nodular arteriolar hyalinosis ○ Acute tubular injury ○ Thrombotic microangiopathy – Acute □ Thrombi glomerular capillaries, arterioles □ Acute endothelial injury – Chronic □ Duplication of GBM □ Nodular arteriolar hyalinosis • Diabetic nephropathy ○ Arteriolar hyalinosis, nodular glomerulosclerosis • Hypertensive nephrosclerosis ○ Arterial intimal fibroelastosis, global or segmental glomerulosclerosis, interstitial fibrosis, tubular atrophy • Membranous glomerulonephritis ○ Thickened glomerular capillaries with deposits and "spikes" along GBM • HCV-related glomerular disease ○ Duplication of GBM, mesangial hypercellularity, pseudothrombi • Minimal change disease ○ Normal glomeruli, reabsorption droplets in tubules • Viral drug toxicity ○ Crystals in tubules and sometimes in glomerular capillaries ○ Oxalate deposition and pigmented casts • Drug reaction ○ Interstitial nephritis – Interstitial manifestation of GVHD in kidney not defined • Viral infection ○ Nuclear inclusions, hyperchromaticity, interstitial inflammation, granuloma (Adenovirus)

ANCILLARY TESTS Immunohistochemistry • Viral infection ○ Viral antigens (polyoma, Adenovirus, Cytomegalovirus)

Immunofluorescence • Membranous glomerulonephritis 175

Kidney Transplantation

Kidney Diseases in Nonrenal Transplant Recipients Kidney Disease in Recipients of Nonrenal Transplants Liver

Heart

Lung

Hematopoietic

210

28

49

49

Arteriosclerosis/hypertensive vascular disease

34%

71%

47%

8%

Calcineurin inhibitor toxicity/arteriolar hyalinosis

22%

25%

69%

6%

Thrombotic microangiopathy

9%

7%

24%

16%

Focal segmental glomerulosclerosis

19%

36%

24%

2%

MPGN

6%

4%

DM

19%

IgAN

4%

Minimal change disease

1%

16% (25% with tip lesion)

Membranous glomerulonephritis

4%

22%

Crescentic glomerulonephritis, NOS

0.5%

Amyloidosis

0.5%

Number Vascular Disease

Glomerular Disease

6% 7%

4%

Tubulointerstitial Disease Polyomavirus infection

2%

Oxalate deposition

10%

Pigmented casts

16%

Nephrocalcinosis ATN

21% 10%

50%

Hydroxyethyl starch nephrotoxicity

6%

4% 43%

27%

8%

Myeloma cast nephropathy

2%

Data are % of renal biopsies. More than 1 diagnosis in some biopsies; categories not uniformly identified or defined; criteria for biopsy not consistent. Combined data from Pillebout, O'Riordan, Schwarz, Lafaucheur, Kambham, Gutierrez, Chang, Kim, and Taheri and Colvin (unpublished).

○ IgG, C3, granular deposits along GBM ○ PLA2R(-) • HCV-related glomerular disease ○ IgM, IgG, C3, C1q granular deposits along GBM and in mesangium • Recurrent AL amyloid and cast nephropathy ○ Light chain restriction

SELECTED REFERENCES 1. 2. 3. 4.

Electron Microscopy • Minimal change disease ○ Effacement of podocyte foot processes • Diabetic nephropathy ○ Diffuse thickening of GBM • Membranous glomerulonephritis ○ Subepithelial deposits and GBM "spikes" • CNI toxicity (thrombotic microangiopathy) ○ Loss of fenestrations of glomerular endothelial cells ○ Duplication of GBM

5. 6.

7. 8.

9. 10.

DIAGNOSTIC CHECKLIST

11.

Pathologic Interpretation Pearls • Cause for renal failure often assumed to be CNIT without direct evidence or biopsy

12.

13.

176

Kuppachi S et al: BK polyoma virus infection and renal disease in non-renal solid organ transplantation. Clin Kidney J. 9(2):310-8, 2016 Wong L et al: Renal transplantation outcomes following heart and heartlung transplantation. Ir J Med Sci. ePub, 2016 Lachance K et al: Risk factors for chronic renal insufficiency following cardiac transplantation. Ann Transplant. 20:576-87, 2015 Mehta V et al: Adenovirus disease in six small bowel, kidney and heart transplant recipients; pathology and clinical outcome. Virchows Arch. 467(5):603-8, 2015 Terzi A et al: Clinicopathologic study of kidney biopsies in patients before or after liver transplant. Exp Clin Transplant. 12 Suppl 1:129-35, 2014 Kubal C et al: Chronic kidney disease after nonrenal solid organ transplantation: a histological assessment and utility of chronic allograft damage index scoring. Transplantation. 93(4):406-11, 2012 Kim JY et al: The variable pathology of kidney disease after liver transplantation. Transplantation. 89(2):215-21, 2010 Schwarz A et al: Biopsy-diagnosed renal disease in patients after transplantation of other organs and tissues. Am J Transplant. 10(9):2017-25, 2010 O'Riordan A et al: Renal biopsy in liver transplant recipients. Nephrol Dial Transplant. 24(7):2276-82, 2009 Lefaucheur C et al: Renal histopathological lesions after lung transplantation in patients with cystic fibrosis. Am J Transplant. 8(9):1901-10, 2008 Troxell ML et al: Renal pathology in hematopoietic cell transplantation recipients. Mod Pathol. 21(4):396-406, 2008 Chang A et al: Spectrum of renal pathology in hematopoietic cell transplantation: a series of 20 patients and review of the literature. Clin J Am Soc Nephrol. 2(5):1014-23, 2007 Pillebout E et al: Renal histopathological lesions after orthotopic liver transplantation (OLT). Am J Transplant. 5(5):1120-9, 2005

Kidney Diseases in Nonrenal Transplant Recipients Collapsing Focal Segmental Glomerulosclerosis Due to CNIT (Left) This native kidney biopsy from a recipient of a liver transplant 4 years previously, maintained on cyclosporine, shows patchy interstitial fibrosis, tubular atrophy, and global glomerulosclerosis. A small artery is normal. These findings are typical, but not diagnostic, of chronic CNIT. (Right) This native kidney from a heart-lung transplant recipient shows collapsing FSGS with reactive epithelial cells bridging between the GBM and Bowman capsule st. Marked arteriolar hyalinosis was evident.

Diabetic Glomerulopathy

Kidney Transplantation

Chronic CNIT

Immune Complex Glomerular Disease (Left) This is a native kidney biopsy from a recipient of a liver transplant 10 years previously for HCV. The biopsy also had immune complexes presumably related to the HCV, which are reported to exacerbate diabetic glomerulopathy. (Right) A native kidney from a recipient of liver transplant shows subepithelial ﬉, mesangial ﬈, and intramembranous ﬉ amorphous deposits, indicative of an immune complex glomerulopathy, probably related to HCV. Diabetic glomerulopathy was also present.

Acute TMA Due to CNIT

Subclinical TMA Due to CNIT (Left) This native kidney biopsy is from a heart transplant 6 days previously, maintained on tacrolimus. Cr rose to 4.2 mg/dL. Marked glomerular endothelial cell injury is present ﬈, manifested by swelling and loss of fenestrations. (Right) TMA may present without the usual signs, such as schistocytes and thrombocytopenia. This biopsy is from a recipient of a heart transplant 13 years ago with a Cr of 6 mg/dL. Marked glomerular endothelial injury is present with loss of fenestrations ﬉ and duplication of the GBM ﬈.

177

Kidney Transplantation

Graft-vs.-Host Glomerulopathies KEY FACTS

TERMINOLOGY • Graft-vs.-host disease (GVHD) glomerulopathy • Glomerular injury in setting of hematopoietic cell transplantation (HCT) and GVHD ○ Rare after autologous HCT

ETIOLOGY/PATHOGENESIS • HCT ○ Mostly allogeneic (80-100%) • Radiation &/or chemotherapy may be contributing factors

CLINICAL ISSUES • Proteinuria, nephrotic range ○ Onset associated with decreased immunosuppression • Drugs ○ Corticosteroids ○ Mycophenolate mofetil ○ Rituximab • Minimal change disease (MCD): ~ 90% complete remission

• Membranous glomerulonephritis (MGN): ~ 27% complete remission

MICROSCOPIC • 3 major patterns ○ MGN – PLA2R negative □ Rare positive case ○ Minimal change lesions ○ Focal segmental glomerulosclerosis • Concurrent interstitial nephritis, acute tubular injury, polyomavirus nephropathy, or thrombotic microangiopathy may be present

TOP DIFFERENTIAL DIAGNOSES • MGN, primary • Recurrent lymphoma • Thrombotic microangiopathy

Membranous Glomerulonephritis

IgG

Microspherular Electron-Dense Deposits

Minimal Change Disease

(Left) Jones methenamine silver demonstrates thick glomerular basement membrane with sparse silver staining due to the prominent extent of subepithelial immune complex deposition. (Right) IgG demonstrates strong granular to confluent staining of the capillary walls in this hematopoietic cell transplant patient with membranous glomerulonephritis (MGN) and graft-vs.-host disease (GVHD).

(Left) EM demonstrates subepithelial and intramembranous deposits with a microspherular substructure ﬉. A subset of MGN cases in the setting of GVHD may reveal this atypical finding. (Right) Widespread effacement of podocyte foot process ﬉ is shown in a patient 1 year after allogeneic BMT for acute myeloid leukemia. He developed nephrotic-range proteinuria while in remission from leukemia and had no other evidence of GVHD. The loss of endothelial fenestrations ﬈ suggests subclinical TMA.

178

Graft-vs.-Host Glomerulopathies

Abbreviations • Graft-vs.-host disease (GVHD) glomerulopathy

Definitions • Glomerular injury in setting of hematopoietic cell transplantation (HCT)

ETIOLOGY/PATHOGENESIS Hematopoietic Cell Transplantation • Mostly allogeneic (80-100%) • 3 patterns ○ Membranous glomerulonephritis (MGN) – Rare report in autologous HCT ○ Minimal change disease (MCD) ○ Focal segmental glomerulosclerosis • Radiation &/or chemotherapy may be contributing factors

Experimental Model • Chronic GVHD (parent to F1 bone marrow transplant) in mice leads to MGN due to antibodies to minor MHC antigens

CLINICAL ISSUES Presentation • Proteinuria, nephrotic range ○ Usual onset – MCD: ~ 8 months post transplant – MGN: ~ 14 months post transplant ○ Onset associated with decreased immunosuppression • Associated with GVHD ○ Skin, mucous membranes, GI tract, lungs ○ Acute GVHD – MCD: ~ 40% – MGN: ~ 80% ○ Chronic GVHD – MCD: ~ 50% – MGN: ~ 90%

Treatment • Drugs ○ Corticosteroids ○ Mycophenolate mofetil ○ Rituximab

Prognosis • MCD: ~ 90% complete remission • MGN: ~ 27% complete remission

– Segmental adhesions and sclerosis – Tip lesions described – Patchy tubular atrophy and fibrosis • Other diseases may be present ○ Interstitial nephritis, acute tubular injury, polyomavirus nephropathy, thrombotic microangiopathy, recurrent amyloidosis, myeloma cast nephropathy

ANCILLARY TESTS Immunofluorescence • MGN ○ Fine granular deposits diffusely along glomerular basement membrane for IgG and variably for other immunoglobulins and C3 ○ Kappa and lambda stain equally ○ No staining of deposits for phospholipase A2 receptor 1 • Minimal change lesion ○ No deposits • Focal segmental glomerulosclerosis ○ Segmental IgM, C3 in scarred glomeruli

Electron Microscopy • MGN ○ Subepithelial electron-dense deposits – Microspherule substructure may be present – Stage I or II (Ehrenreich-Churg) ○ Mesangial electron-dense deposits – Variably present • Minimal change lesion ○ Diffuse podocyte foot process effacement ○ No deposits • Focal segmental glomerulosclerosis ○ Same as MCD plus segmental adhesions • Endothelial tubuloreticular inclusions (rare)

DIFFERENTIAL DIAGNOSIS Membranous Glomerulonephritis, Primary • Absence of mesangial immune complexes

Recurrent Lymphoma • Manifested as MCD or MGN • Lack of GVHD history

Thrombotic Microangiopathy • Chronic phase shows double contours • Fibrin and nonspecific trapping of IgM/C3

SELECTED REFERENCES 1.

MICROSCOPIC Histologic Features • 3 major patterns ○ MGN (~ 60%) – Subepithelial spike formation by Jones silver stain ○ Minimal change lesion (~ 25%) – Podocyte hypertrophy – Tubular reabsorption droplets ○ Focal segmental glomerulosclerosis (~ 15%)

Kidney Transplantation

TERMINOLOGY

2.

3.

4.

5.

Brinkerhoff BT et al: Renal pathology in hematopoietic cell transplant recipients: a contemporary biopsy, nephrectomy, and autopsy series. Mod Pathol. 29(6):637-52, 2016 Hiramatsu R et al: Clinicopathological analysis of allogeneic hematopoietic stem cell transplantation-related membranous glomerulonephritis. Hum Pathol. 50:187-94, 2016 Byrne-Dugan CJ et al: Membranous nephropathy as a manifestation of graftversus-host disease: association with HLA antigen typing, phospholipase A2 receptor, and C4d. Am J Kidney Dis. 64(6):987-93, 2014 Mii A et al: Renal thrombotic microangiopathy associated with chronic graftversus-host disease after allogeneic hematopoietic stem cell transplantation. Pathol Int. 61(9):518-27, 2011 Brukamp K et al: Nephrotic syndrome after hematopoietic cell transplantation: do glomerular lesions represent renal graft-versus-host disease?. Clin J Am Soc Nephrol. 1(4):685-94, 2006

179

Kidney Transplantation

Calcineurin Inhibitor Toxicity KEY FACTS

TERMINOLOGY • Kidney dysfunction attributable to injury from calcineurin inhibitor immunosuppressive agents

ETIOLOGY/PATHOGENESIS • Transplanted and native kidneys affected by calcineurin inhibitor toxicity (CNIT) • CNIT is dose related with variable individual susceptibility • Functional CNIT: Reversible acute renal dysfunction associated with afferent arteriolar vasoconstriction • Structural CNIT: Tubular and vascular direct toxicity

CLINICAL ISSUES • Acute or chronic elevation of serum creatinine • Elevated blood or serum levels of CNI • Correlation of structural tissue injury and blood levels is not strong

MICROSCOPIC • Tubular toxicity

○ Acute: Acute tubular injury with focal isometric vacuolization of proximal tubular segments ○ Chronic: Striped interstitial fibrosis and tubular atrophy with microcalcifications • Vascular toxicity ○ Acute arteriolopathy: Smooth muscle loss and expansion of intima and media by loose matrix ○ Chronic arteriolopathy: Nodular medial hyalinization ○ Thrombotic microangiopathy: Acute and chronic

DIAGNOSTIC CHECKLIST • Exposure to CNI is sine qua non for diagnosis ○ Elevated blood levels of CNI and long-term exposure increase certainty of diagnosis • Absence of pathologic lesions does not exclude functional CNIT • Observation of combined tubulopathy and vasculopathy increases diagnostic certainty • No single histologic lesion is specific or pathognomonic

Isometric Vacuolization

Chronic Tubular Toxicity

Acute Vascular Toxicity

Chronic Vascular Toxicity

(Left) Straight portions of proximal tubules have isometric vacuolization ﬈ associated with elevated trough levels of calcineurin inhibitor (CNI). (Right) The striped pattern of fibrosis ﬈ in chronic tubular CNIT is probably a result of both chronic ischemia in "watershed" zones of the medullary rays, from hyaline arteriolosclerosis and direct tubular toxicity.

(Left) Thrombotic microangiopathy (TMA) in calcineurin inhibitor toxicity (CNIT) demonstrates mural fibrin deposition ﬊ with medial erythrocytolysis ﬈ and luminal stenosis. The lower arteriolar profile has prominent endothelium with preserved medial smooth muscle. (Right) Hyalinization increases from initial outer medial nodules to involve the media and intima, resulting in transmural hyalinization in renal allograft CNIT. Hyalinization retains a nodular pattern in outer media ﬈.

180

Calcineurin Inhibitor Toxicity

Abbreviations • Calcineurin inhibitor toxicity (CNIT)

○ Kidney transplants – TMA in 2-5% – Hyaline arteriolosclerosis in 60-70% at 2 years; > 90% at 10 years

Synonyms

Presentation

• Cyclosporine toxicity, cyclosporine A (CsA) toxicity, tacrolimus toxicity, FK506 toxicity

• Acute or chronic elevation of serum creatinine ○ CNIT may arise at any time after initiation of therapy • ↑ trough CNI blood levels may confirm diagnosis, but correlation of structural tissue injury and blood level is not strong • TMA may be systemic or localized to kidney (40%)

Definitions • Acute or chronic kidney dysfunction attributable to direct injury from calcineurin inhibitor immunosuppressive agents

ETIOLOGY/PATHOGENESIS Types of Calcineurin Inhibitor Toxicity • Functional CNIT: Reversible acute renal dysfunction associated with afferent arteriolar vasoconstriction • Structural CNIT: Characterized by cellular injury and matrix remodeling ○ Tubular CNIT: Acute tubular injury with vacuolization of epithelium ○ Vascular CNIT: Direct toxic injury to endothelium and smooth muscle of arterioles – Thrombotic microangiopathy (TMA) or chronic hyaline arteriolopathy – Glomerular endothelial injury also feature • Tubular and vascular toxicity typically coexist • Native and transplanted kidneys develop CNIT with similar histologic manifestations

Mechanisms • Histologic lesions are dose related ○ Acute CNIT with markedly elevated blood levels of CNI ○ Chronic CNIT with long-term exposure to CNI • Dose-independent susceptibility factors, e.g., genes for enzymes that metabolize arachidonic acid • Genetic defects in complement regulation in 30-65% of patients with CNI-TMA • CsA and tacrolimus bind intracellular receptors called immunophilins ○ Immunophilin/CNI complexes bind and inhibit calcineurin ○ Calcineurin is T-cell activator via nuclear factors of activated T cells (NFAT) ○ NFAT activates transcription of interleukin-2, interferonγ, and tumor necrosis factor-α ○ Endothelium: Increased thromboxane A2, endothelin-1, superoxide and peroxynitrite, decreased prostaglandin and prostacyclin, apoptosis, necrosis • Immunosuppressive potency and renal toxicity of CNI are pharmacologically inseparable • Renal toxic effects of CsA and tacrolimus are identical • CNIT affects endothelium, vascular smooth muscle, and tubular epithelium ○ Tubular epithelium: Vacuolization, megamitochondria, calcification, necrosis ○ Smooth muscle: Vacuolization, necrosis, apoptosis, hyalinization

CLINICAL ISSUES Epidemiology

Kidney Transplantation

TERMINOLOGY

Treatment • Dose reduction or cessation of CNI therapy • Complement inhibitors explored in TMA

Prognosis • Acute CNIT typically reversible and associated with resolution of histologic changes • Chronic CNIT less likely to be reversible ○ Resolution of arteriolopathy rarely reported

MICROSCOPIC Histologic Features • Functional CNIT ○ No morphologic tissue injury by definition • Tubular CNIT ○ Acute – Focal proximal tubular epithelial isometric vacuolization in straight > convoluted segments – Vacuolar changes may be accompanied by acute tubular injury ± dystrophic microcalcification – Large eosinophilic cytoplasmic granules are megamitochondria (CsA toxicity) or lysosomes (tacrolimus toxicity) ○ Chronic – Striped fibrosis characterized by radial fibrosis of cortical medullary rays with intervening nonscarred parenchyma ± tubular microcalcification □ Result of chronic ischemia from arteriolopathy and direct tubular toxicity • Vascular CNIT ○ Acute and chronic vasculopathy may be present in same biopsy ○ Acute arteriolopathy: Focal lesion – Loss of definition of smooth muscle cells, cytoplasmic vacuolization, and dropout – Clear or basophilic medial or intimal loose matrix accumulation with separation of myocytes – Intimal or medial platelet insudates [CD61(+)] ○ TMA – Arteriolar thrombi, intimal and medial fibrinoid change ± erythrocytolysis, platelets [CD61(+)] – Obliterative arteriolopathy has stenosis, intimal and medial concentric hypercellularity ("onion skinning") – Arteries may have intimal myxoid thickening ○ Chronic arteriolopathy – Early lesions have hyaline replacement of individual outer medial smooth muscle cells

• Incidence 181

Kidney Transplantation

Calcineurin Inhibitor Toxicity – Nodular hyalinization of outer media imparts eosinophilic, PAS(+) beaded necklace appearance – Over months to years, entire vessel wall is hyalinized – Hyalinization mainly affects afferent arterioles □ Can involve vasa recta and small arteries – Frequency of arteriolar hyalinosis similar with cyclosporine and tacrolimus • Glomerulopathy and CNIT ○ Acute TMA – Capillary thrombi; glomerular hilar thrombi ("pouch lesions") – Capillary double contours and mesangiolysis ○ Chronic TMA and other chronic lesions – Capillary basement membrane double contours – Ischemic collapse, obsolescence, and focal segmental glomerular sclerosis • Immunohistology ○ Acute arteriolopathy and TMA – Immunofluorescence: Arteriolar and glomerular IgM, C3, and fibrinogen – Immunoperoxidase staining: CD61 or CD62 (+) platelet deposits in arterioles and glomeruli ○ Chronic arteriolopathy – IgM, C3, and C1q in hyaline deposits; no platelets • Electron microscopy ○ Tubules: Dilated endoplasmic reticulum; multiple large lysosomes, megamitochondria, endocytotic vesicles ○ Arterioles and glomeruli – Endothelial swelling, cytoplasmic vacuolization, detachment from basal lamina and apoptosis or necrosis □ Capillary double contours and interposition – Myocyte vacuoles, disruption of myofibrils, detachment from basal lamina, apoptosis – Electron-dense hyaline material replaces smooth muscle

DIFFERENTIAL DIAGNOSIS Tubulopathy • Osmotic tubulopathy associated with exposure to parenteral carbohydrates, intravenous immunoglobulin, or radiocontrast agents ○ Diffuse cytoplasmic vacuolization, swollen epithelium, preserved brush border – Vacuoles are endosomes and phagolysosomes • Ischemic acute tubular injury ○ Coarse irregular vacuoles • Tubulopathy associated with lipiduria in nephrotic syndrome ○ Vacuoles are isometric, focal, and contain lipid, ± interstitial foam cells ○ PAS(+) protein droplets in tubules; stain for Ig and albumin by immunofluorescence ○ Causal glomerulopathy is typically identified

Vasculopathy • TMA ○ Acute antibody-mediated rejection – Transplant glomerulitis, peritubular capillaritis, each with neutrophils, and C4d (+) 182

– Donor-specific antibodies ○ Recurrent hemolytic uremic syndrome ○ Antiphospholipid nephropathy: Antiphospholipid or anticardiolipin antibodies ○ Malignant hypertension – May be due to complement dysregulation • Acute arteriolopathy ○ Indistinguishable changes seen in severe hypertensive arteriolosclerosis • Hyaline arteriolosclerosis of diabetic nephropathy and hypertension ○ Initial lesions classically intimal rather than medial ○ Peripheral nodular medial hyalinization may be seen rarely ○ Transmural hyalinization in advanced disease, making determination of etiology difficult ○ Hyaline deposits in afferent and efferent vessels in diabetic arteriolopathy • Amyloid vasculopathy deposits Congo red (+) and not typically nodular

Focal Segmental Glomerulosclerosis • Widespread effacement of foot processes in primary focal segmental glomerulosclerosis (FSGS) • Less conspicuous arteriolar hyalinosis • CNIT may have collapsing variant of FSGS

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Absence of pathologic lesions in kidney biopsy does not exclude CNIT • Exposure to CNI inhibitors is sine qua non for CNIT • Combined tubulopathy and vasculopathy increases diagnostic certainty

Pathologic Interpretation Pearls • Acute CNIT: Tubulopathy with isometric vacuolization, acute tubular injury, acute arteriolopathy, and TMA • Chronic CNIT: Nodular arteriolar hyalinization, striped fibrosis, calcifications, global and segmental glomerular sclerosis • No single histologic lesion specific or pathognomic • Lack of arteriolar hyalinosis 3 months to 3 years after transplantation associated with ↓ 5-year graft survival ○ May reflect decreased CNI dosage or nonadherence ○ Increased susceptibility to antibody-mediated rejection

SELECTED REFERENCES 1. 2. 3. 4.

5.

Einecke G et al: Hyalinosis lesions in renal transplant biopsies: timedependent complexity of interpretation. Am J Transplant. ePub, 2016 Jodele S et al: The genetic fingerprint of susceptibility for transplantassociated thrombotic microangiopathy. Blood. 127(8):989-96, 2016 Bröcker V et al: Arteriolar lesions in renal transplant biopsies: prevalence, progression, and clinical significance. Am J Pathol. 180(5):1852-62, 2012 Snanoudj R et al: Specificity of histological markers of long-term CNI nephrotoxicity in kidney-transplant recipients under low-dose cyclosporine therapy. Am J Transplant. 11(12):2635-46, 2011 Mihatsch MJ et al: Histopathology of cyclosporine nephrotoxicity. Transplant Proc. 20(3 Suppl 3):759-71, 1988

Calcineurin Inhibitor Toxicity

Acute Vascular Toxicity (Left) The profile of a normalappearing arteriole is seen with PAS highlighting the basal lamina of the medial smooth muscle cells ﬈. The endothelial basal lamina is also evident ﬊. (Right) In early vascular CNIT, the medial smooth muscle cells lose their discrete definition ﬈ and may acquire PAS(+) granules ﬇. The discrete, well-delineated appearance of outer medial hyalin nodules ﬊ indicates hyaline replacement of myocytes within the confines of the basal lamina.

Chronic Vascular Toxicity

Kidney Transplantation

Normal Arteriole

Hyaline Replacing Myocyte (Left) Nodular outer medial hyalinization ﬈ with a discrete beaded appearance is evident in this biopsy from a patient with chronic CNIT. Prominent intimal hyalinization with thickening of the endothelial basal lamina is also evident ﬊. (Right) EM shows nodular hyaline arteriolosclerosis in a kidney transplant with CNIT. Upper myocyte shows shrinkage & detachment from basal lamina ﬈. Rounded amorphous hyaline material ﬇ fills the basal lamina, replacing the myocyte and compressing adjacent cells ſt.

Arteriolar Hyalinosis Vasa Recta

Peripheral Nodular Hyalinosis (Left) Vasa recta are also prone to chronic CNIT. This is an example of extensive transmural hyalinization of the arteriolar vasa in a renal transplant with chronic CNIT ﬈. Venules are unaffected ﬊. (Right) Multiple peripheral nodular hyaline deposits are present in this afferent arteriole from a donor biopsy st. Occasionally, this pattern of hyalinosis can be seen without a history calcineurin inhibitor exposure.

183

Kidney Transplantation

Calcineurin Inhibitor Toxicity

Myxoid Matrix

CD61 Deposition

Obliterative Arteriolopathy

CD61 Deposits Suggest TMA

Glomerular Thrombi

Platelet CD61 in Thrombi

(Left) Myxoid matrix accumulation is shown in the media and intima of an arteriole from a kidney transplant with CNIT. There is segmental absence of smooth muscle cell nuclei ﬈. (Right) Immunohistochemical staining for CD61 may reveal unexpected platelet insudation in the intima or media of arterioles with myxoid changes by light microscopy. The tissue is a biopsy from a renal transplant with CNIT.

(Left) A kidney transplant biopsy with severe obliterative arteriolopathy from CNIT is shown. The arterioles have luminal obliteration, loss of smooth muscle, and matrix accumulation ﬈ and were observed in the context of greatly elevated blood levels of tacrolimus. (Right) Obliterative arteriolopathy attributable to CNIT may have unrecognized mural platelet deposits. Immunostaining for CD61 reveals granular platelets and platelet microparticles in the intima and media.

(Left) This glomerulus has segmental occlusive glomerular capillary fibrinplatelet thrombi from a kidney transplant with CNIT and TMA. Pericapsulitis is nonspecific. (Right) Immunohistochemical staining for CD61, a marker of platelets, reveals occlusive glomerular capillary thrombi in a renal allograft with CNIT and TMA.

184

Calcineurin Inhibitor Toxicity

Endothelial Injury in CNIT (Left) Glomeruli in chronic TMA due to CNIT have segmental or global double contours of the GBM ﬈. These lesions represent repair of endothelial injury and must be distinguished from transplant glomerulopathy. Thrombi may be absent from glomeruli with these lesions. (Right) EM of a native kidney with TMA due to CNI reveals marked endothelial injury manifested by loss of fenestrations ﬉ in this glomerular capillary from a liver transplant recipient.

Focal Segmental Glomerulosclerosis

Kidney Transplantation

Chronic Vascular Toxicity

Collapsing Glomerulopathy (Left) Perihilar focal segmental glomerulosclerosis ﬊ associated with arteriolar hyalinization ﬈ in a renal allograft is shown in a patient with exposure to CNI for more than 7 years. (Right) Collapsing glomerulopathy is shown in a native kidney of a heart-lung transplant recipient on CNI for 10 years. Severe arteriolar hyalinosis due to CNI was also present. Severe microvascular disease is a known cause of collapsing glomerulopathy.

Acute Tubular Toxicity

Chronic Tubular Toxicity: Striped Fibrosis (Left) Megamitochondria appear as irregularly shaped eosinophilic globules ﬊ in the epithelial cytoplasm in tubular CNIT. (Right) The pattern of striped fibrosis is due to ischemia in the watershed areas of the cortex and the medullary rays related to CNIT, here illustrated in a trichrome stain of a native kidney from a lung transplant recipient. (Courtesy S. Rosen, MD.)

185

Kidney Transplantation

mTOR Inhibitor Toxicity KEY FACTS

ETIOLOGY/PATHOGENESIS • Mammalian target of rapamycin (mTOR) drugs ○ Inhibit tubular epithelial cell proliferation and apoptosis in setting of acute injury ○ Decreases VEGF synthesis, related to development of focal segmental glomerulosclerosis (FSGS)

CLINICAL ISSUES • Acute renal failure, delayed graft function (acute toxicity) • Proteinuria (chronic toxicity)

MICROSCOPIC • Acute mTOR inhibitor toxicity ○ Severe acute tubular injury ○ Epithelial cell necrosis ○ Atypical, eosinophilic PAS(-) casts reminiscent of myeloma cast nephropathy – Stain for cytokeratin by immunohistochemistry

○ Myoglobin-appearing casts; stain for myoglobin by immunohistochemistry ○ Thrombotic microangiopathy • Chronic mTOR inhibitor toxicity ○ FSGS ○ Abnormal podocyte phenotype, suggestive of podocyte dedifferentiation

TOP DIFFERENTIAL DIAGNOSES • Acute mTOR inhibitor toxicity ○ Severe acute tubular necrosis ○ Acute antibody-mediated rejection ○ Light chain (myeloma) cast nephropathy ○ Rhabdomyolysis • Chronic mTOR inhibitor toxicity ○ FSGS due to other factors

Acute Tubular Injury

Focal Segmental Glomerulosclerosis

Thrombus

Myoglobin

(Left) PAS shows acute rapamycin toxicity with PAS(-) casts ﬊ and severe acute tubular injury. The material consists of cytoplasmic debris from damaged tubular epithelial cells. (Right) De novo focal segmental glomerulosclerosis ﬈ due to chronic mammalian target of rapamycin (mTOR) inhibitor toxicity is present in a 58-yearold man with 425 mg/day of proteinuria 2.5 years after transplantation. He had never been treated with calcineurin inhibitors.

(Left) Acute mTOR inhibitor toxicity shows a glomerular thrombus ﬊ in a renal transplant patient on sirolimus and tacrolimus. Either sirolimus or tacrolimus by itself may cause thrombotic microangiopathy (TMA); there is greater risk of TMA in combined sirolimus and tacrolimus therapy than in sirolimus without a calcineurin inhibitor. (Right) Tubular casts can stain for myoglobin by IHC in mTOR inhibitor toxicity, similar to those in rhabdomyolysis. This is presumptive evidence of muscle injury.

186

mTOR Inhibitor Toxicity

Abbreviations • Mammalian target of rapamycin (mTOR)

Definitions • mTOR inhibitors used in renal transplantation ○ Rapamycin (sirolimus) ○ Everolimus – Similar structure to sirolimus; shorter half-life

ETIOLOGY/PATHOGENESIS mTOR Inhibitor (Rapamycin/Sirolimus): Drug Effects • Sirolimus binds FK506 binding protein-12 to form sirolimus effector protein (SEP) complex • SEP complex inhibits mTOR pathway ○ Sirolimus blocks cytokine-mediated signal transduction, affecting T-cell cycle progression ○ Decreases lymphocyte proliferation • mTOR expressed in kidney, e.g., tubular epithelial cells ○ Repair response to acute tubular injury requires tubular epithelial cell turnover and proliferation – Sirolimus inhibits tubular epithelial cell proliferation and apoptosis • Decreases VEGF synthesis ○ Mechanism of thrombotic microangiopathy (TMA)

– Epithelial simplification, loss of tubular brush border – Vacuolization, shedding – Epithelial cell necrosis – Tubular dilation ○ Casts – Atypical, eosinophilic PAS(-) casts □ Stain for cytokeratin by immunohistochemistry – Irregular, sharply demarcated cast edges – Casts may appear "fractured" with surrounding cellular reaction □ Reminiscent of myeloma cast nephropathy – Myoglobin-appearing casts; stain for myoglobin by immunohistochemistry ○ Histologic resolution upon sirolimus removal ○ TMA – Increased risk in patients on both sirolimus and cyclosporine – Some associated with lab evidence of acute TMA: Thrombocytopenia, anemia, low haptoglobin levels • Chronic mTOR inhibitor toxicity ○ FSGS – May have collapsing pattern □ May be due to unsampled TMA – Some have proteinuria without FSGS

DIFFERENTIAL DIAGNOSIS

mTOR Inhibitor Effect on Podocyte

Severe Acute Tubular Necrosis

• Focal segmental glomerulosclerosis (FSGS) lesions show abnormal podocyte phenotype, suggestive of podocyte dedifferentiation ○ Pax-2 and cytokeratin expression in proliferating podocytes ○ Loss of synaptopodin and VEGF expression ○ Downregulated nephrin expression in podocytes – Similar pattern in sirolimus-related FSGS and other types of FSGS • In vitro decreased VEGF synthesis and Akt phosphorylation by podocytes in presence of sirolimus and decreased synthesis of WT1, protein required for podocyte integrity

• Due to causes other than mTOR inhibitor

CLINICAL ISSUES Presentation • Delayed graft function ○ More common in mTOR inhibitor treatment (25%) than without (9%) ○ Correlates with mTOR inhibitor dose • Acute renal failure (acute mTOR inhibitor toxicity) • Proteinuria (chronic mTOR inhibitor toxicity) • Toxicity can occur in native kidneys

Treatment • Drugs ○ Discontinuation or reduced dose of mTOR inhibitor ○ Initiation of alternative immunosuppressive therapy

MICROSCOPIC

Rhabdomyolysis • Myoglobin casts present in both rhabdomyolysis and mTOR inhibitor toxicity

Acute Antibody-Mediated Rejection • C4d(+) peritubular capillaries • May show acute tubular necrosis &/or TMA

Light Chain (Myeloma) Cast Nephropathy • Monotypic light chain immunolocalization • Serum or urine paraprotein usually detectable • Unusual in renal allograft

Focal Segmental Glomerulosclerosis • Recurrent or de novo in allograft • FSGS pattern may be result of chronic calcineurin inhibitor (CNI) toxicity ○ Arteriolar hyalinosis also present in chronic CNI toxicity

SELECTED REFERENCES 1. 2.

3. 4.

5.

Histologic Features • Acute mTOR inhibitor toxicity ○ Severe acute tubular injury

Kidney Transplantation

TERMINOLOGY

6.

Koppelstaetter C et al: Effect of cyclosporine, tacrolimus and sirolimus on cellular senescence in renal epithelial cells. Toxicol In Vitro. 48:86-92, 2018 Vollenbröker B et al: mTOR regulates expression of slit diaphragm proteins and cytoskeleton structure in podocytes. Am J Physiol Renal Physiol. 296(2):F418-26, 2009 Letavernier E et al: High sirolimus levels may induce focal segmental glomerulosclerosis de novo. Clin J Am Soc Nephrol. 2(2):326-33, 2007 Pelletier R et al: Acute renal failure following kidney transplantation associated with myoglobinuria in patients treated with rapamycin. Transplantation. 82(5):645-50, 2006 Sartelet H et al: Sirolimus-induced thrombotic microangiopathy is associated with decreased expression of vascular endothelial growth factor in kidneys. Am J Transplant. 5(10):2441-7, 2005 Smith KD et al: Delayed graft function and cast nephropathy associated with tacrolimus plus rapamycin use. J Am Soc Nephrol. 14(4):1037-45, 2003

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Acute Pyelonephritis KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Acute renal parenchymal inflammation due to bacterial or fungal infection

• • • • • •

ETIOLOGY/PATHOGENESIS • Retrograde infection via urethra, bladder, and ureters ○ Escherichia coli common organism • Risk factors include immunosuppression, advanced age, diabetes, female gender, ileal loop, urinary reflux

CLINICAL ISSUES • • • •

Fever, chills, graft tenderness Leukocyte casts Renal failure Early acute pyelonephritis (APN) associated with increased late graft loss • May precipitate acute rejection episode • Some persistent cases of allograft APN respond to combination of antimicrobial therapy and steroids

Neutrophilic infiltrate and casts in tubules Abscesses and papillary necrosis may arise Early infection in medulla and collecting ducts Pelvic inflammation in ascending infection Acute tubulointerstitial rejection may coexist Glomeruli and blood vessels relatively spared

ANCILLARY TESTS • C4d binds bacterial surfaces in APN via activation of lectin pathway in ring pattern

TOP DIFFERENTIAL DIAGNOSES • Concurrent pyelonephritis and acute T-cell-mediated rejection • Acute T-cell-mediated rejection • Acute antibody-mediated rejection • Drug-induced interstitial nephritis • Ischemic or toxic tubular injury

Gross Appearance of Papillary Necrosis

Necrotic Papilla in Acute Pyelonephritis

Neutrophil Casts in Transplant Kidney

Bacterial Colonies in Acute Pyelonephritis

(Left) Bivalved transplant kidney nephrectomy with severe acute pyelonephritis and papillary necrosis is shown. The necrotic papilla is sharply demarcated by a hemorrhagic zone ﬉. (Right) This transplant kidney has severe acute pyelonephritis and papillary necrosis. The necrotic papilla is sharply demarcated by a hemorrhagic zone ﬉. The necrotic area shows no residual viable cells ﬈.

(Left) Light microscopy shows acute pyelonephritis in this renal allograft. Abundant neutrophils are present in the edematous interstitium ﬈ and prominently as tubular casts ﬉. (Right) Hematoxylin & eosin at high power reveals a cluster of bacteria in the medulla of a transplant kidney with severe pyelonephritis. In contrast to calcium deposits, bacteria are more uniform and are typically smaller.

188

Acute Pyelonephritis

Abbreviations • Acute pyelonephritis (APN)

Synonyms • Acute bacterial nephritis (used when no pyelitis) • Upper urinary tract infection (UTI)

Definitions • Acute bacterial infection of kidney parenchyma

ETIOLOGY/PATHOGENESIS Ascending Infection • Retrograde infection via urethra, bladder, and ureters • Most common route of infection (95%) • Usually associated with predisposing factor ○ Renal transplantation immunosuppression – APN most common bacterial complication – Cyclosporine A may compromise host defenses against Escherichia coli ○ Obstruction ○ Stent ○ Reflux nephropathy ○ Diabetes • Gram-negative bacteria from gastrointestinal tract (fecal flora) most common ○ E. coli most common organism – Uropathogenic and virulence factors include □ Fimbriae/pili and serotypes O, K, and H ○ P fimbriae (mannose resistant) – Attach to digalactoside residue on urothelial cells and facilitate persistent infection – Enhance host innate inflammatory response by interaction with toll-like receptor 4 (TLR4), resulting in IL-6 and IL-8 production □ α-intercalated cells of collecting ducts may defend against E. coli by acidifying urine and secreting bacteriostatic protein lipocalin 2 (NGAL) ○ Type 1 fimbriae (mannose sensitive) – Binds Tamm-Horsfall protein ○ Proteus ○ Klebsiella ○ Enterobacter ○ Pseudomonas ○ Streptococcus faecalis

Hematogenous Infection • In septicemia or bacterial endocarditis ○ Staphylococcus aureus common pathogen ○ Fungal organisms in immunocompromised hosts

Asymptomatic Bacteriuria • Bacterial counts of > 100,000 colony forming units/mL of urine in asymptomatic patients

CLINICAL ISSUES Site • Pelvic urothelium (pyelitis) • Renal tubules and interstitium (pyelonephritis)

• Renal cortical abscesses (bacterial nephritis)

Presentation • • • • • •

May be subclinical Fever and chills Nausea and vomiting Flank pain Costovertebral angle tenderness Acute renal failure ○ Transplant kidney ○ Emphysematous pyelonephritis

Kidney Transplantation

TERMINOLOGY

Laboratory Tests • Urine microscopy ○ Pyuria (white blood cells in urine) ○ Leukocyte casts ○ Gram stain may be positive • Urine cultures ○ Midstream urine minimizes contamination

Treatment • Drugs ○ Antimicrobials – Empiric therapy with gram-negative coverage until culture results obtained ○ Some persistent cases of allograft pyelonephritis respond to combination of antimicrobial therapy and steroids – Possibly representing concurrent APN and acute rejection

Prognosis • Acute uncomplicated APN responds to antimicrobial therapy in > 90% of cases ○ Lack of response in – Drug-resistant bacterial strains – Presence of anatomic abnormality (reflux, etc.) – Urinary tract obstruction • Recurrent pyelonephritis can lead to chronic pyelonephritis and chronic kidney disease • APN or asymptomatic bacteruria increases risk of acute rejection • APN in first 6 months post transplant associated with increased graft dysfunction at 2 years (43% vs. 18% in Japanese study) • APN < 30 days post transplant with lower long-term graft and patient survival (Brazilian study)

MACROSCOPIC General Features • Enlarged and edematous kidney • Ascending pyelonephritis ○ Characteristic straight yellow streaks seen in medulla – Correspond to collecting ducts filled with pus ○ Pyelitis usual ○ Scarred areas represent residua of prior episodes – Cortical scars in ascending APN overlie pelvic calyces ○ Inflammation may extend into perirenal fat with perinephric abscess ○ Calculi or strictures may be seen in pelvis or ureters • Emphysematous pyelonephritis 189

Kidney Transplantation

Acute Pyelonephritis ○ Empty (gas-filled) rounded spaces in cortex and perirenal fat ○ Dissection planes under capsule have abscesses

MICROSCOPIC Histologic Features • Patchy neutrophilic infiltrate in tubules and interstitium ○ Neutrophil casts in tubular lumina, sometimes with associated bacteria ○ Neutrophilic tubulitis ○ Abscesses with tubular destruction in severe cases • Lymphocytes, plasma cells, eosinophils, and macrophages seen within days • Acute tubular injury ○ Loss of proximal tubular brush border ○ Simplified tubular epithelium ○ Sloughed epithelial cells in tubular lumina • Tubular basement membranes disrupted • Papillary necrosis in severe cases ○ Usually tip of papilla unlike analgesic nephropathy

Variants • Ascending infection ○ Acute inflammation of pelvis usual ○ Early infection in medulla and collecting ducts ○ Organisms invade via fornix of calyx and intrarenal reflux through collecting ducts • Hematogenous infection ○ Randomly scattered abscesses in cortex ○ Occasionally, septic emboli in glomeruli containing organisms in endocarditis ○ Little or no inflammation of pelvis • Emphysematous pyelonephritis ○ Round empty spaces and planes of dissection in tissue due to gas-forming organisms • Acute lobar nephronia ○ Variant that suggests mass lesions in imaging studies ○ Focal areas of intense inflammation and edema in cortex

ANCILLARY TESTS Histochemistry • Tissue Gram stain (Brown-Brenn) ○ Detects gram-positive & -negative organisms ○ Bacteria typically not conspicuous in tubules but can be detected in abscesses • Jones methenamine silver ○ Positive in tubules and abscesses if fungal organisms present

Immunohistochemistry • C4d binds bacterial surfaces in APN via lectin pathway activation in ring pattern

DIFFERENTIAL DIAGNOSIS Drug-Induced Interstitial Nephritis • Clinical history of drug exposure ○ Neutrophilic infiltrate less prominent ○ Urine and blood cultures are negative 190

Acute T-Cell-Mediated Rejection • Allograft pyelonephritis may be refractory to antimicrobial therapy ○ May represent combination of acute allograft pyelonephritis and acute rejection (type I) – Gene expression analysis of some allograft pyelonephritis resembles acute rejection compared to native pyelonephritis ○ Bacterial infection may induce loss of tolerance to allograft

Acute Antibody-Mediated Rejection • Neutrophils in tubules sometimes mimic APN • C4d in peritubular capillaries • Abscesses not seen

Ischemic or Toxic Tubular Injury • Less neutrophilic infiltrate • No abscesses • Tubular injury in area without inflammation

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Abscess formation • Papillary necrosis • Histological evidence of chronic pyelonephritis ○ Numerous hyaline casts in atrophic tubules reminiscent of thyroidization ○ Can indicate prior episodes of APN or presence of urinary outflow obstruction

Pathologic Interpretation Pearls • Fungal infections can be missed (carefully review PAS and silver stains) • Persistent infection may have abundant eosinophils

SELECTED REFERENCES 1.

Alhajjaj FS et al: Emphysematous pyelonephritis in renal allograft - a case report. Int J Health Sci (Qassim). 10(2):311-3, 2016 2. Kroth LV et al: Acute graft pyelonephritis occurring up to 30 days after kidney transplantation: epidemiology, risk factors, and survival. Transplant Proc. 48(7):2298-2300, 2016 3. Oghumu S et al: Differential gene expression pattern in biopsies with renal allograft pyelonephritis and allograft rejection. Clin Transplant. 30(9):111533, 2016 4. Shin DH et al: Early-onset graft pyelonephritis is predictive of long-term outcome of renal allografts. Tohoku J Exp Med. 236(3):175-83, 2015 5. Singh R et al: Asymptomatic bacteriuria and urinary tract infections among renal allograft recipients. Curr Opin Infect Dis. 28(1):112-6, 2015 6. Kumar S et al: Acute pyelonephritis in diabetes mellitus: Single center experience. Indian J Nephrol. 24(6):367-71, 2014 7. Oghumu S et al: Acute pyelonephritis in renal allografts: a new role for microRNAs? Transplantation. 97(5):559-68, 2014 8. Tourneur E et al: Cyclosporine A impairs nucleotide binding oligomerization domain (Nod1)-mediated innate antibacterial renal defenses in mice and human transplant recipients. PLoS Pathog. 9(1):e1003152, 2013 9. Schmidt S et al: Emphysematous pyelonephritis in a kidney allograft. Am J Kidney Dis. 53:895-7, 2009 10. Joss N et al: Lobar nephronia in a transplanted kidney. Clin Nephrol. 64:3114, 2005

Acute Pyelonephritis

Neutrophil Casts in Medulla (Left) Hematoxylin & eosin shows pelvic urothelium infiltrated by neutrophils and lymphocytes ﬉, compatible with acute pyelonephritis. Neutrophil exudate is seen in the lumen along with bacteria ﬊. (Right) Periodic acid-Schiff shows neutrophil casts ﬇ within the collecting ducts of the medulla. Mild interstitial edema is seen. Early acute pyelonephritis due to ascending infection can be localized to the medulla only.

Neutrophil Casts in Acute Pyelonephritis

Kidney Transplantation

Acute Pyelitis With Bacteria

Neutrophil Casts and Tubular Epithelial Reactive Changes (Left) Periodic acid-Schiff of a kidney biopsy shows neutrophil casts within the cortical tubules ﬊, compatible with acute bacterial pyelonephritis. Interstitial neutrophils were seen elsewhere. (Right) A cortical collecting duct shows a neutrophil cast ﬉ filling the lumen. The tubular epithelium is attenuated and basophilic, indicating injury and reactive changes, respectively. The only tubules with branches in the kidney are the collecting ducts.

Neutrophil Casts in Transplant Kidney

C4d Immunostain in Acute Pyelonephritis (Left) A renal transplant biopsy shows intense neutrophilic infiltrate forming a tubular cast. The patient had a positive urine culture. This pattern strongly favors acute pyelonephritis over rejection, but neutrophils in tubules are not uncommon in acute antibody-mediated rejection. (Right) C4d IHC of acute pyelonephritis shows a cluster of positive-staining cocci in a tubular lumen ﬉. C4d is activated by bacterial surface carbohydrates via the lectin pathway (mannose-binding lectin, ficolin).

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Kidney Transplantation

Polyomavirus Nephritis KEY FACTS

TERMINOLOGY

• Tubular basement membrane immune complex deposition

• Polyomavirus infection in kidney allografts or native kidneys of immunosuppressed or immunocompromised patients

ANCILLARY TESTS

ETIOLOGY/PATHOGENESIS • BK polyomavirus causes ~ 85% of PVN • JC virus causes ~ 15% of PVN

CLINICAL ISSUES • • • • •

Acute renal failure ~ 5% prevalence in kidney transplant patients Association with ureteral obstruction Treated by reduction/change in immunosuppressive drugs 7-100% 3-year graft loss, depending on pathologic stage

MICROSCOPIC • Interstitial inflammation, mononuclear • Tubulitis • Nuclear inclusions

• Immunohistochemistry (IHC) for polyoma large T antigen • Electron microscopy for viral particles • Plasma PCR screen for viral load > 10⁴/mL

TOP DIFFERENTIAL DIAGNOSES • • • •

Acute tubulointerstitial (type I) rejection Adenovirus nephritis Acute tubular injury/necrosis Acute interstitial nephritis

DIAGNOSTIC CHECKLIST • Concurrent polyomavirus nephritis and acute rejection occurs, but infrequently ○ Endarteritis or C4d in peritubular capillaries indicates rejection also present • When borderline inflammation present, perform SV40 IHC, even in absence of viral nuclear changes

Zonal Distribution of Inflammation

Intranuclear Inclusion

Immunohistochemistry for SV40 Large T Antigen

Polyomavirus Virions

(Left) Periodic acid-Schiff demonstrates dense zonal or regional interstitial inflammation, which emphasizes the importance of sufficiently sampling an allograft. The diagnosis of polyomavirus nephritis (PVN) would be missed if only the lower 1/2 of this renal cortex were biopsied. The virus is typically found in the areas of inflammation. (Right) Hematoxylin & eosin shows an intranuclear inclusion ﬈ with a ground-glass appearance in this distal tubule. The adjacent interstitium shows edema and scattered lymphocytes.

(Left) Immunohistochemistry for polyoma large T antigen (SV40) stains many tubular epithelial cell nuclei ﬈ accompanied by prominent interstitial inflammation. This is diagnostic of polyomavirus infection but does not distinguish the specific viral agent. (Right) High-power electron micrograph of a tubular epithelial cell nucleus shows a cluster of polyoma virions that measure ~ 40 nm. These are significantly smaller than adenovirus or herpesviruses.

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Polyomavirus Nephritis

Abbreviations • Polyomavirus nephritis (PVN)

Synonyms • Polyomavirus nephropathy • BK virus nephropathy

Definitions • Polyomavirus infection of kidney, usually in immunocompromised host

ETIOLOGY/PATHOGENESIS Infectious Agents • Human polyomavirus ○ BK virus – Tropism for genitourinary tract epithelium – High seroprevalence in adults (80%) – Pathogenic only in immunocompromised patients – Causes ~ 85% of PVN ○ John Cunningham (JC) virus – Causes ~ 15% of PVN, usually milder than BK – Causes progressive multifocal leukoencephalopathy ○ Simian virus 40 (SV40), Merkel cell polyomavirus – Rarely, if ever, cause interstitial nephritis in humans

Pathogenesis • Renal allograft ○ Reactivation of latent virus from donor organ ○ Renal injury promotes viral replication ○ Rejection contributes to pathogenesis • Native kidney ○ AIDS, genetic immunodeficiency, immunosuppression ○ Uncommon in stem cell or solid organ (other than kidney) transplantation

CLINICAL ISSUES Epidemiology • Incidence ○ ~ 5% in kidney transplant patients on tacrolimus and mycophenolate mofetil (MMF) • Risk factors ○ Tacrolimus/MMF vs. cyclosporine/MMF (odds ratio = 3) ○ Prior rejection episode, older age, male gender

Presentation • Acute renal failure • Hemorrhagic cystitis • Ureteral stenosis ○ Occurs in 5-10% of BK PVN • Late complication: High-grade urothelial malignancies with expression of viral proteins (large T)

Laboratory Tests • Plasma PCR ○ > 10⁴ virions/mL highly specific for PVN (98%) but not sensitive (64%) – ~ 30% of PVN have plasma levels < 10⁴/mL ○ Rare PVN in absence of BK viremia

– Typical of JC virus PVN • Urine ○ Decoy cells by cytology – Not specific for PVN but indicate polyoma infection of urinary tract ○ Urine PCR less specific for PVN ○ Viral aggregates ("Haufen") by negative-staining electron microscopy (EM) – High sensitivity and specificity for PVN (> 98%)

Kidney Transplantation

TERMINOLOGY

Treatment • Reduce tacrolimus/mycophenolate/switch to low-dose cyclosporine • Current antivirals not highly effective ○ Cidofovir ○ Leflunomide • Retransplant generally successful

Prognosis • Graft loss depends on stage at diagnosis (13-100%) ○ Poorer prognosis with interstitial fibrosis and tubular atrophy ○ Rare graft loss with JC • Rejection episodes follow in 8-12% • Residual impairment of renal function common • BK virus possibly oncogenic ○ Minority of urothelial tumors in renal allografts express large T antigen

MICROSCOPIC Histologic Features • Interstitial mononuclear inflammation ○ Lymphocytes and eosinophils ○ Plasma cells usually prominent ○ Often associated with viral-infected epithelial cells • Intranuclear inclusions in tubular epithelium ○ Ground-glass nuclear appearance ○ Nuclear enlargement and hyperchromatism ○ Nuclear inclusions may be present in sloughed cells in tubular lumina ○ Inclusions may not be evident in early PVN • Tubulitis and tubular injury ○ Plasma cells occasionally in tubules ○ Apoptosis common • Distal nephron involved more than proximal nephron ○ May involve only renal medulla in early stages, especially collecting ducts ○ Advanced stages involve parietal epithelial cells of glomeruli – Can mimic cellular crescent • Late changes ○ Interstitial fibrosis and tubular atrophy ○ Extent of tubulointerstitial scarring often correlates with duration of viral infection ○ Correlates with graft survival ○ Dedifferentiated pattern of tubular epithelial cells; appear spindled, possibly reflecting epithelialmesenchymal transition • High-grade urothelial and renal cell carcinoma reported ○ Expresses large T antigen in all tumor cells (not VP1) 193

Kidney Transplantation

Polyomavirus Nephritis Classifications for Polyomavirus Nephritis Stage

University of Maryland (2004)

3-Year Graft Loss (Maryland)

Banff Working Group (2009)

3-Year Graft Loss (Banff)

A (early)

Virus-infected cells with no or minimal interstitial inflammation or tubular atrophy

13%

Virus-infected cells with no or minimal tubular injury

7%

B (active)

Virus-infected cells with interstitial B1: 40%; B2: 60%; inflammation/tubular atrophy involving < 25% B3: 77% (B1), 26-50% (B2), or > 50% (B3) of cortex

Tubular epithelial cell necrosis or lysis with denudation of basement membrane spanning > 2 cells

50%

C (inactive/late)

Rare cytopathic effect with extensive interstitial inflammation/tubular atrophy

> 50% interstitial fibrosis with any degree of tubular injury

100%

100%

ANCILLARY TESTS Immunohistochemistry • Polyomavirus large T antigen IHC diagnostic ○ Protein of early phase of polyomavirus infection ○ Antibody to SV40 large T antigen detects BK and JC viruses ○ Strong nuclear staining of epithelial cells – Tubular epithelial cells, often mostly distal tubules and collecting ducts – Commonly clustered positive cells – Infrequent glomerular parietal epithelial cells ○ Best results reported with PAb416 at concentration > 1/100 and polymer-based detection

Immunofluorescence • Granular staining of tubular basement membranes (TBMs) for IgG, C3, and C4d in ~ 50% of PVN ○ Viral antigens reported in deposits in 1 of 2 studies (not large T or VP1) ○ May persist despite disappearance of polyomavirus by IHC or EM ○ Significance unknown, associated with higher creatinine • C4d in atrophic TBMs may mimic peritubular capillary staining in antibody-mediated rejection

Electron Microscopy • Polyomavirus particles present within epithelial cells ○ ~ 40 nm viral particles in paracrystalline arrays or loose clusters ○ Nuclear and occasionally cytoplasmic location • Discrete electron-dense deposits within TBMs ○ May be present in atrophic tubules ○ Use immunofluorescence to confirm that electron-dense deposits represent immune complexes

DIFFERENTIAL DIAGNOSIS

Acute Tubular Injury/Necrosis • Reactive atypia of tubular epithelial cells mimics viral nuclear changes • IHC negative for polyoma large T antigen (SV40)

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Stage of disease correlates with outcome • Viral load scores (Banff) as % of IHC-positive tubular nuclei ○ pv1 < 1% ○ pv2 = 1-10% ○ pv3 > 10%

Pathologic Interpretation Pearls • Predominant medullary inflammation raises PVN suspicion • When borderline inflammatory infiltrate present, large T antigen IHC is indicated ○ Early PVN may lack viral cytopathic changes ○ Large T antigen in 1 tubular epithelial cell nucleus is diagnostic of infection • Prominent interstitial inflammation with sparse large T antigen expression suggests concurrent acute rejection • Concurrent PVN and acute rejection occurs, but infrequently • TBM immune complexes may persist after viral stains become negative

SELECTED REFERENCES 1. 2.

3.

4.

Acute Tubulointerstitial (Type I) Rejection • Prominent interstitial inflammation with tubulitis unassociated with large T antigen ○ Rare cases of concurrent acute rejection and PVN

5.

6.

Adenovirus Tubulointerstitial Nephritis • Interstitial hemorrhage, necrosis, and granulomas • Positive adenovirus IHC, negative polyoma IHC

Acute Interstitial Nephritis • IHC negative for polyoma large T antigen (SV40)

194

7. 8.

Castro T et al: Polyomavirus BK and JC in individuals with chronic kidney failure, kidney transplantation, and healthy controls. J Clin Virol. 89:5-9, 2017 Simard-Meilleur MC et al: Stabilization of renal function after the first year of follow-up in kidney transplant recipients treated for significant BK polyomavirus infection or BK polyomavirus-associated nephropathy. Transpl Infect Dis. 19(3), 2017 Adam B et al: Banff Initiative for Quality Assurance in Transplantation (BIFQUIT): reproducibility of polyomavirus immunohistochemistry in kidney allografts. Am J Transplant. 14(9):2137-47, 2014 Singh HK et al: Polyomavirus nephropathy: quantitative urinary polyomavirus-Haufen testing accurately predicts the degree of intrarenal viral disease. Transplantation. 99(3):609-15, 2014 Hirsch HH et al: Polyomavirus BK replication in de novo kidney transplant patients receiving tacrolimus or cyclosporine: a prospective, randomized, multicenter study. Am J Transplant. 13(1):136-45, 2013 McDaid J et al: Transitional cell carcinoma arising within a pediatric donor renal transplant in association with BK nephropathy. Transplantation. 95(5):e28-30, 2013 Menter T et al: Pathology of resolving polyomavirus-associated nephropathy. Am J Transplant. 13(6):1474-83, 2013 Bracamonte E et al: Tubular basement membrane immune deposits in association with BK polyomavirus nephropathy. Am J Transplant. 7(6):155260, 2007

Polyomavirus Nephritis

Polyomavirus Large T Antigen (Left) Light microscopy of PVN shows widespread tubular changes with large, atypical epithelial cells with spindle shapes and enlarged nuclei ﬈. The inflammation is confined to areas with viral cytopathic changes. (Right) Immunohistochemistry for polyoma large T antigen (SV40) shows widespread infection of tubules with a characteristic clustering of positive cells within individual tubules. The inflammatory infiltrate is typically in the same area as the virus, suggesting that the virus causes the inflammation.

Intranuclear Inclusion

Kidney Transplantation

Prominent Interstitial Inflammation

Viral Cytopathic Effect (Left) In this biopsy from a patient with an advanced stage of PVN, tubular atrophy, interstitial fibrosis, and a focal mononuclear infiltrate are present, which are all nonspecific findings. Only 1 nuclear inclusion was found as a clue to the etiology ﬉. (Right) Periodic acid-Schiff shows severe interstitial inflammation and a typical, lavender, homogeneous intranuclear inclusion ﬈ of polyomavirus. Lymphocytes and plasma cells are in the interstitial infiltrate.

Ground-Glass Intranuclear Inclusions

Intranuclear Inclusions (Left) Periodic acid-Schiff reveals several tubular epithelial cells with groundglass intranuclear inclusions ﬈ and nucleoli displaced against the nuclear membrane. Tubulitis ﬊ [or lymphocytes between tubular epithelial cells and tubular basement membrane (TBM)] is a common finding that mimics acute rejection when viral cytopathic changes are not prominent. (Right) Spindleshaped (dedifferentiated) tubular cells show viral inclusions in nuclei ﬉.

195

Kidney Transplantation

Polyomavirus Nephritis

Viral Cytopathic Effect

Plasma Cell Tubulitis

Collecting Duct Cast

Virion Shedding in Tubule

Decoy Cells

"Haufen" Viral Particles in Urine

(Left) Hematoxylin & eosin shows several nuclei ﬈ with characteristic cytopathic effect of polyomavirus infection. Nucleoli are often pushed to the nuclear membrane. These features correlate with electron microscopic (EM) findings. (Right) This tubule has largely dedifferentiated or missing epithelium. A plasma cell is present in the tubule ﬈, a distinctive finding in PVN. Many plasma cells ﬉ are in the interstitium, which is also typical of PVN.

(Left) Antibody to SV40 large T antigen shows prominent staining of debris in a collecting duct cast. When shed in the urine, these are the origin of the "Haufen" detected by EM. (Courtesy V. Nickeleit, MD.) (Right) EM of a tubular epithelial cell shows shedding of polyomavirus into the lumen ﬈ and formation of cast-like aggregates ﬉, which can be detected in the urine as "Haufen" by negativestaining EM.

(Left) Composite image of urine cytology from patients with PVN shows decoy cells with inclusions. These cells, which resemble malignant cells, are not diagnostic of PVN but only of polyoma infection of the urinary tract, which may be asymptomatic. (Right) Urine sediment examined by negative-staining EM shows an aggregate of virions of ~ 40 nm in diameter, typical of polyomavirus. These aggregates are found almost exclusively in patients with PVN. (Courtesy V. Nickeleit, MD.)

196

Polyomavirus Nephritis

Large T Antigen in Parietal Epithelium (Left) Hematoxylin & eosin shows several parietal epithelial cells with enlarged and smudged nuclei ſt and scattered lymphocytes between the Bowman capsule and epithelial cells (or "capsulitis"), which is an uncommon histologic feature of severe PVN. (Right) SV40 large T IHC shows several parietal epithelial cells ﬈ infected by polyomavirus without nuclear enlargement. Scattered interstitial inflammation ﬊ is present adjacent to this glomerulus.

Pseudocrescent

Kidney Transplantation

Parietal Epithelial Cells With Inclusions

Severe Tubular Atrophy and Loss (Left) Hematoxylin & eosin shows prominence of the parietal epithelial cells ﬈, which may be seen in a small subset of PVN and may occasionally mimic cellular crescents. Glomerular fibrinoid necrosis is absent. (Right) An advanced stage of PVN may show only nonspecific tubular loss, atrophy, and interstitial fibrosis, a.k.a. chronic allograft nephropathy. Without a prior diagnosis of PVN, the cause would be unknown in this patient.

Index Case From Patient B.K.

Urothelial Viral Infection (Left) This is the original case of polyomavirus infection from an allograft, reported by S.D. Gardner (St. Mary's Hospital, London). The ureter shows intense inflammation and ulceration. The novel polyomavirus was then named after the patient's initials, B.K. (Courtesy E. Ramos, MD.) (Right) Ureter from an allograft nephrectomy with PVN shows numerous positive urothelial cells ﬉ for SV40 large T antigen. The mucosa shows marked inflammation with scattered lymphocytes invading the epithelium ﬈.

197

Kidney Transplantation

Polyomavirus Nephritis

IgG Deposits in TBMs

Granular C4d Deposits in TBMs

JC Polyomavirus Nephropathy

JC Polyomavirus Nephropathy

High-Grade Transitional Cell Carcinoma

High-Grade Urothelial Carcinoma Expressing Polyomavirus Large T Antigen

(Left) Immunofluorescence (IF) in a case of PVN shows intense granular deposition of IgG in some but not all TBMs. Granular IgG in the TBM is indicative of immune complex deposition and is not a nonspecific finding. C3 is similarly deposited but is not specific since C3 is commonly detected in the TBM. (Right) Immunohistochemistry for C4d shows granular deposits along the TBM of a subset of the tubules. The peritubular capillaries are negative.

(Left) John Cunningham (JC) polyomavirus infections have similar, although often milder, histologic changes. Viral cytopathic effect is seen in 1 nucleus ſt. JC virus is suspected when the SV40 large T antigen stain is positive, but the blood PCR for BK virus is negative, since only the latter is specific for BK. (Courtesy R.N. Smith, MD.) (Right) JC virus large T antigen crossreacts with the usual antibody to SV40 large T antigen as shown in the medulla of this case of JC polyomavirus nephropathy. (Courtesy R.N. Smith, MD.)

(Left) High-grade urothelial and renal cell carcinomas have been reported in renal transplants that express the large T antigen in all tumor cells (but not the normal parenchyma). This high-grade papillary pelvic tumor was detected 5 years after an episode of PVN. (Courtesy E. Farkash, MD.) (Right) Approximately 20% of the urothelial tumors arising in renal transplant recipients express polyomavirus large T antigen. Large T antigens of other polyomaviruses are known to be oncogenic. (Courtesy E. Farkash, MD.)

198

Polyomavirus Nephritis

TBM Deposits (Left) Low-power EM of a cross section of a tubule reveals widespread, electrondense, amorphous deposits in the TBM ﬉. (Right) EM of the TBM deposits at high power shows that the deposits are amorphous and contain scattered membranous debris. No viral particles are evident. SV40 large T antigen has been reported in these deposits using indirect IF microscopy.

Intranuclear Virions

Kidney Transplantation

TBM Deposits

Intranuclear Virions (Left) EM of a severely altered tubule shows large aggregates of polyoma virions in the tubular epithelial nuclei st as well as an intratubular plasma cell ﬈ and lymphocyte ﬉. The TBM is unremarkable ﬈. (Right) EM demonstrates numerous individual viral particles ﬈ within the nucleus ﬊ of a tubular epithelial cell. Nucleoli ﬉ are pushed aside against the nuclear membrane. This can also be appreciated on light microscopy.

Cytoplasmic Virions

BK Polyoma Virions (Left) EM shows an aggregate of virions within the cytoplasm next to the nucleus of this infected tubular epithelial cell. The viral particles can be closely grouped together or occasionally arranged in a paracrystalline array (not shown). (Right) EM of BK polyomavirus shows viral particles in the nucleus of ~ 40 nm. Polyomavirus is substantially smaller than adenovirus (75-80 nm) or herpes group viruses (150-200 nm).

199

Kidney Transplantation

Adenovirus, Kidney KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Synonym ○ Adenovirus (AdV) nephritis

• • • •

ETIOLOGY/PATHOGENESIS • Nonenveloped, double-stranded DNA virus

Granulomatous inflammation Necrosis of tubules Interstitial hemorrhage Smudgy, basophilic, intranuclear inclusions in tubular cells

ANCILLARY TESTS

CLINICAL ISSUES • Fever • Graft tenderness • Hemorrhagic cystitis usual with kidney involvement ○ Gross hematuria • Acute renal failure • Blood real-time polymerase chain reaction for diagnosis and surveillance • Cidofovir, ribavirin, intravenous immunoglobulin • Recovery common if localized • > 60% fatal if disseminated

• Positive for AdV by immunohistochemistry • EM shows characteristic 60- to 80-nm virions

TOP DIFFERENTIAL DIAGNOSES • Acute T-cell-mediated rejection • Polyomavirus nephritis • Drug-induced acute interstitial nephritis

DIAGNOSTIC CHECKLIST • Coinfection with other organisms common • Antiviral antibody panel valuable in differential diagnosis (AdV, SV40, CMV)

Necrotizing Granulomata

Adenovirus Cytopathic Effect

Adenovirus Immunohistochemistry

Adenovirus Virions

(Left) Necrotizing granulomata are typical of adenovirus infection with neutrophils, plasma cells, and lymphocytes in this renal allograft. Viral cytopathic effect ﬈ and tubulitis ﬊ are noted. (Courtesy L. NovoaTakara, MD.) (Right) Hematoxylin and eosin shows viral cytopathic effect ﬈ in several tubular epithelial cell nuclei within a tubule with tubulitis ﬉ and necrosis ﬊, which is surrounded by abundant interstitial inflammation and a hint of palisading macrophages.

(Left) Adenovirus immunohistochemistry shows both strong staining ﬈ and faint staining ﬊ in the nuclei of several tubular epithelial cells in a renal allograft biopsy. Note the scattered interstitial inflammatory cells and focal tubulitis ﬉. (Right) Electron microscopy shows adenovirus virions ſt in a paracrystalline array within an infected epithelial cell nucleus. The individual virions measure ~ 80 nm in diameter, which is 2x the size of human polyomavirus.

200

Adenovirus, Kidney

Infectious Agents • Adenovirus (AdV) ○ Nonenveloped, double-stranded DNA virus ○ Possible routes of infection – Reactivation of endogenous latent infection – Transplanted organ or tissue • Other organs affected: Bladder, lung, liver, GI tract

CLINICAL ISSUES Epidemiology • Incidence ○ Rare in kidney transplant recipients (< 1%) – Onset typically in first 3 months post transplant ○ AdV more common in stem cell recipients (3-7%) – Native kidney occasionally involved • Age ○ Children more susceptible (< 5 years)

Presentation • Fever • Hemorrhagic cystitis ○ Gross hematuria ○ AdV rarely causes renal infection in absence of cystitis • Acute kidney injury • Graft tenderness • Bladder and ureter involvement may cause obstructive uropathy

○ Granulomas associated with viral-infected tubular epithelial cells and tubular destruction – Peritubular location ○ Viral cytopathic effect in tubular epithelial cells – Smudgy basophilic intranuclear inclusions □ Detached infected cells within tubular lumina □ Distal tubules infected more than proximal tubules • Glomeruli ○ Glomerular (visceral and parietal) epithelial cells may be infected • Vessels: No specific finding

ANCILLARY TESTS Immunohistochemistry • Nuclear and cytoplasmic staining for AdV

Immunofluorescence • No deposits along the tubular basement membranes

Electron Microscopy • Viral particles 60-80 nm in diameter in tubular nuclei

DIFFERENTIAL DIAGNOSIS Acute T-Cell-Mediated Rejection • Reactive atypia of tubular nuclei may mimic viral inclusions • Granulomas sometimes seen associated with tubular destruction in acute T-cell-mediated rejection • Less severe hemorrhage and tubular necrosis than AdV • AdV lacks endarteritis [and C4d(+)]

Laboratory Tests

Polyomavirus Nephritis

• Real-time polymerase chain reaction ○ AdV in blood precedes symptoms by > 3 weeks • Shell vial assay (culture) • Enzyme immunoassay for AdV antigen in blood

• Positive SV40 immunohistochemistry • Less hemorrhage and tubular necrosis than AdV • More plasma cells and less granulomatous inflammation

Treatment

• Hemorrhage and necrosis minimal • No viral antigen present

• Drugs ○ Cidofovir, ribavirin; valganciclovir or ganciclovir ○ Intravenous immunoglobulin • Reduction of immunosuppressive agents

Prognosis • Disseminated disease often fatal (> 60%) • Recovery common if localized

MACROSCOPIC General Features • White-yellowish streaks with hemorrhagic rim primarily in medulla • Hemorrhagic mucosal surface in renal pelvis and ureters

MICROSCOPIC

Drug-Induced Acute Interstitial Nephritis

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Necrotizing granuloma is distinctive feature • Panel of antiviral antibodies valuable in differential diagnosis (AdV, SV40, CMV, HSV) • Coinfection with other fungal or viral organisms may occur

SELECTED REFERENCES 1.

2.

3.

Histologic Features • Tubules and interstitium ○ Acute tubular injury and interstitial nephritis – Focal necrosis of tubules – Interstitial hemorrhage and edema

Kidney Transplantation

ETIOLOGY/PATHOGENESIS

4.

5.

Nanmoku K et al: Clinical characteristics and outcomes of adenovirus infection of the urinary tract after renal transplantation. Transpl Infect Dis. 18(2):234-9, 2016 Klein J et al: Late presentation of adenovirus-induced hemorrhagic cystitis and ureteral obstruction in a kidney-pancreas transplant recipient. Proc (Bayl Univ Med Cent). 28(4):488-91, 2015 Mehta V et al: Adenovirus disease in six small bowel, kidney and heart transplant recipients; pathology and clinical outcome. Virchows Arch. 467(5):603-8, 2015 Lachiewicz AM et al: Adenovirus causing fever, upper respiratory infection, and allograft nephritis complicated by persistent asymptomatic viremia. Transpl Infect Dis. 16(4):648-52, 2014 Keddis M et al: Adenovirus-induced interstitial nephritis following umbilical cord blood transplant for chronic lymphocytic leukemia. Am J Kidney Dis. 59(6):886-90, 2012

201

Kidney Transplantation

Adenovirus, Kidney

Adenovirus Nephritis

Medullary Interstitial Inflammation

Granulomas

Peritubular Multinucleated Giant Cells

Adenovirus Smudged Nuclei

Peritubular Multinucleated Giant Cells

(Left) Hematoxylin and eosin demonstrates severe interstitial inflammation, granulomas ﬈, tubular necrosis ﬊, and interstitial edema in this renal allograft with adenovirus infection. (Right) Periodic acid-Schiff reveals prominent interstitial inflammation and frequent tubulitis in the renal medulla, which is common for adenovirus but atypical for acute rejection. Patchy involvement of the renal cortex (not shown) is also present.

(Left) Granulomas of epithelioid macrophages and tubular destruction are typical of adenovirus infection in the kidney. Granulomas are not common in polyomavirus or cytomegalovirus infections. (Right) Hematoxylin and eosin reveals 2 giant cells ﬈ surrounding a severely injured tubule with necrosis ﬊. Interstitial inflammatory cells consist of epithelioid macrophages, lymphocytes, plasma cells, and rare eosinophils.

(Left) Periodic acid-Schiff shows enlarged, tubular epithelial cell nuclei with a smudged appearance ﬈, which resemble polyomavirus, but the presence of tubular degeneration/necrosis ﬊ and granulomatous inflammation favors adenovirus nephritis. (Right) Periodic acid-Schiff highlights multinucleated giant cells ﬈ closely associated with an injured tubule, which contains necrotic and sloughed epithelial cells that are intermixed with lymphocytes in a renal allograft infected with adenovirus.

202

Adenovirus, Kidney

Adenovirus Immunohistochemistry (Left) In situ hybridization for adenovirus highlights a few tubular epithelial cell nuclei ﬈ and cell cytoplasm ﬊ within this tubule with extensive injury and necrosis. (Right) Adenovirus antigens are detected primarily in the nucleus of tubular epithelial cells in the kidney ﬈. Positive cells are typically sparse. A granuloma is seen nearby ﬉.

Adenovirus Virions

Kidney Transplantation

Adenovirus In Situ Hybridization

Adenovirus Virions (Left) Electron microscopy shows numerous adenovirus viral particles forming a paracrystalline array ſt within the nucleus of this infected epithelial cell. (Right) Electron microscopy reveals numerous adenovirus virions ﬊ that have displaced the nuclear chromatin ﬈ against the nuclear membrane within this infected epithelial cell nucleus. The nucleus corresponds with a nuclear inclusion that is observed by light microscopy.

Adenovirus Virions

Adenovirus Dimensions (Left) Electron microscopy demonstrates numerous individual adenovirus virions ſt within this nucleus, which has displaced the nuclear chromatin ﬊ against the nuclear membrane ﬈. (Right) Adenovirus virions are 60-80 nm in diameter, as seen here in a nucleus of an infected tubular epithelial cell. The diameter is larger than the ~ 30- to 45-nm polyomavirus and smaller than cytomegalovirus (150-200 nm).

203

Kidney Transplantation

Cytomegalovirus Infection KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Cytomegalovirus (CMV) infection in kidneys, usually associated with systemic CMV in immunocompromised patients

• "Owl-eye" nuclear inclusions ○ Most prominent in tubular epithelium ○ Glomerular capillary &/or peritubular capillary endothelial cells • Interstitial inflammation, mononuclear • Acute glomerulonephritis (rare)

ETIOLOGY/PATHOGENESIS • Most individuals infected by CMV before adulthood • Immunocompromised patients at risk ○ Neonatal CMV ○ Transplant CMV

CLINICAL ISSUES • Presentation ○ Renal dysfunction ○ Flu-like symptoms • Antiviral agents ○ Ganciclovir or valganciclovir ○ CMV immune globulin • Reduce or alter immunosuppressive agents

TOP DIFFERENTIAL DIAGNOSES • Polyomavirus nephropathy • Adenovirus nephritis • Acute T-cell-mediated rejection

DIAGNOSTIC CHECKLIST • CMV intranuclear inclusions present in predominantly endothelial cells or epithelial cells • Coinfection with other fungal or viral organisms may occur • Features of foscarnet toxicity may be present in patients treated for CMV infection

Glomerular CMV

Peritubular Capillary CMV

CMV Immunohistochemistry

CMV Virions

(Left) Hematoxylin and eosin shows characteristic intranuclear ("owl-eye") inclusions ﬈ of cytomegalovirus (CMV) infection within the glomerular endothelial cells. (Right) Hematoxylin and eosin shows CMV intranuclear inclusions ﬈ in endothelial cells of a peritubular capillary with basophilic cytoplasmic changes ﬊ that are also present in a tubular epithelial cell ﬉.

(Left) CMV immunohistochemistry demonstrates strong nuclear staining ﬈ and a blush of cytoplasmic staining within several glomerular endothelial cells. (Right) Electron microscopy shows individual viral particles ﬈ measuring ~ 150-200 nm in diameter with dense central cores surrounded by a thick capsule, which is characteristic of cytomegalovirus. (Courtesy J. Taxy, MD.)

204

Cytomegalovirus Infection

CLINICAL ISSUES

Abbreviations

Epidemiology

• Cytomegalovirus (CMV)

• Incidence ○ Neonatal CMV – Most common neonatal infection – 0.2-2% of live births in USA – 9.4 per 100,000 infants ages 1-4 years in Australia ○ Transplant CMV – ~ 20% incidence of CMV disease with ganciclovir prophylaxis – ~ 45% incidence without prophylaxis – Frequency of CMV infection in renal transplant biopsies < 1% • Age ○ Neonatal, intrauterine ○ Immunocompromised adults • Sex ○ Male predilection • Ethnicity ○ No ethnic predilection

Synonyms • CMV tubulointerstitial nephritis (TIN) • CMV glomerulopathy • CMV nephropathy

Definitions • Direct CMV infection of kidneys, usually associated with systemic CMV involvement and immunocompromise ○ May promote indirect kidney injury, particularly in renal transplants, including acute allograft glomerulopathy • Causes benign, self-limited mononucleosis syndrome in normal individuals

ETIOLOGY/PATHOGENESIS Infectious Agents • CMV ○ Herpesviridae – β-subfamily – Double-stranded DNA virus ○ a.k.a. human herpesvirus-5 (HHV-5)

Risk Factors • Immunocompromised patients at risk for systemic CMV ○ Transplant recipients on immunosuppression – Transplant CMV from donor organ or reactivation in recipient – Matching CMV serologic status in renal transplant patients has minimized incidence of CMV TIN ○ Infants – Neonatal CMV infection from maternal transmission ○ HIV-infected patients

Site of Infection • Epithelium, endothelium, monocytes • Renal involvement almost always associated with systemic infection ○ Lungs ○ Liver ○ GI tract ○ Pancreas ○ Adrenal glands ○ Epididymitis ○ Bone marrow ○ Retina

Latent Virus • Most individuals infected before adulthood ○ Benign self-limited disease in normal individuals ○ Seroprevalence (90%) ○ Virus remains present in latent state lifelong

Effects on Immune System • Increased IL-6 and IL-10, decreased Th1 cytokines (γinterferon) • Decreased expression of HLA antigens

Kidney Transplantation

TERMINOLOGY

Presentation • • • • •

Fever Malaise Leukopenia Acute kidney injury Proteinuria

Laboratory Tests • CMV IgM antibodies ○ Suggest recent or active infection ○ False-positives due to rheumatoid factor • CMV IgG antibodies • CMV antigen test ○ Indirect IF test to detect pp65 protein of CMV in peripheral blood leukocytes • CMV polymerase chain reaction • Viral culture ○ Shell vial assay

Treatment • Drugs ○ Ganciclovir or valganciclovir – Prophylaxis – Intravenous therapy ○ Foscarnet – Side effects include crystal formation leading to glomerulopathy – Multinucleation of tubular epithelial cell nuclei may persist after foscarnet therapy ○ Cidofovir ○ CMV intravenous immune globulin • Reduce or alter immunosuppressive agents • Vaccination to prevent maternal transmission

Prognosis • Neonatal CMV ○ 30% mortality among symptomatic infants ○ Survivors commonly have neurologic deficits • CMV disease in transplant recipient 205

Kidney Transplantation

Cytomegalovirus Infection Immunohistochemistry Antibody

Reactivity

Staining Pattern

Comment

CMV

Positive

Nuclear & cytoplasmic

Epithelial or endothelial cells

SV40

Negative

Not applicable

Adenovirus

Negative

Not applicable

EBV-LMP

Negative

Not applicable

HSV1/2

Negative

Not applicable

○ Increased graft loss in past (10-20%) ○ Less adverse effect of CMV in patients on current immunosuppressive protocols

MICROSCOPIC Histologic Features • Pattern I: Large intranuclear inclusions in tubular epithelial cells with interstitial nephritis ○ Variable interstitial inflammation ○ Occasional granulomatous inflammation ○ Rare or no intranuclear inclusions in endothelial cells ○ Monocyte inclusions in interstitial infiltrate • Pattern II: Large eosinophilic intranuclear inclusions in endothelial cells ○ Glomerular and peritubular capillary endothelial cells may be infected ○ When endothelial cells are predominant cell infected by CMV, epithelial cells tend to be spared ○ Interstitial inflammation not prominent in cases with primarily endothelial cell infection • Pattern III: Acute glomerulonephritis (rare) ○ Endocapillary hypercellularity ○ Inclusions in glomerular endothelial cells or in circulating monocytes ○ Crescents may be present ○ Scant deposits by EM

ANCILLARY TESTS In Situ Hybridization • CMV positive

Electron Microscopy • Virions in nucleus and cytoplasm • 150-200 nm in diameter • Dense core surrounded by thick capsule

DIFFERENTIAL DIAGNOSIS

• Viral cytopathic effect in tubular epithelial cells • Immunohistochemical confirmation of adenovirus infection

Acute T-Cell-Mediated Rejection • Prominent interstitial inflammation with tubulitis • No viral cytopathic effect • Endarteritis helpful if present

Acute Allograft Glomerulopathy (Form of Rejection) • • • • • • •

Marked glomerular cell endothelial swelling and activation Mesangiolysis: Webs of PAS(+) material CD8 T cells in glomeruli No inclusions or viral antigens in glomeruli May be indirect effect of CMV in some patients Endarteritis common C4d(-)

Acute Glomerulonephritis • No CMV inclusions or antigens • Deposits in GBM

Immunotactoid Glomerulopathy • Microtubular deposits composed of immunoglobulins (often monoclonal) • Few de novo cases in allografts associated with CMV infection

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Inclusions readily evident at low magnification • CMV intranuclear inclusions present in predominantly endothelial cells or epithelial cells • Coinfection with other fungal or viral organisms may occur • Features of foscarnet toxicity may be present in patients treated for CMV infection

SELECTED REFERENCES 1.

Polyomavirus Nephritis • Intranuclear inclusions with ground-glass appearance in tubular epithelial cells • Immunohistochemical for SV40 large T antigen (+) • Prominent interstitial plasmacytic inflammation with tubulitis • No endothelial inclusions

2.

3.

4. 5.

Adenovirus Nephritis • Prominent interstitial inflammation and tubular necrosis • Granulomatous inflammation

206

6.

Redondo-Pachón D et al: Adaptive NKG2C+ NK cell response and the risk of cytomegalovirus infection in kidney transplant recipients. J Immunol. 198(1):94-101, 2017 Feng S et al: Incidence and risk factors for cytomegalovirus infection in patients with kidney transplantation: a single-center experience. Transplant Proc. 48(8):2695-2699, 2016 Vichot AA et al: Cytomegalovirus glomerulopathy and cytomegalovirus interstitial nephritis on sequential transplant kidney biopsies. Am J Kidney Dis. 63(3):536-9, 2014 Rane S et al: Spectrum of cytomegalovirus-induced renal pathology in renal allograft recipients. Transplant Proc. 44(3):713-6, 2012 Agrawal V et al: Polyomavirus nephropathy and cytomegalovirus nephritis in renal allograft recipients. Indian J Pathol Microbiol. 53(4):672-5, 2010 Humar A et al: An assessment of herpesvirus co-infections in patients with CMV disease: correlation with clinical and virologic outcomes. Am J Transplant. 9(2):374-81, 2009

Cytomegalovirus Infection

Tubular Epithelial Cells (Left) Hematoxylin and eosin of a renal transplant biopsy shows a moderately intense mononuclear infiltrate, edema, and tubular injury, resembling acute T-cellmediated (cellular) rejection. Inclusions typical of CMV, however, are seen in the tubules, even at low power ﬉. (Right) Hematoxylin and eosin of an autopsy from an infant who died from systemic CMV disease shows frequent large eosinophilic inclusions in the tubular epithelial cells, which have a hobnail pattern.

Peritubular Capillary CMV

Kidney Transplantation

Proximal Tubules

CMV Intranuclear Inclusions (Left) CMV immunohistochemistry shows strong nuclear ﬈ and some cytoplasmic ﬊ staining of many peritubular capillary endothelial cells ﬉ with marked nuclear enlargement. No CMV staining of the tubular epithelial cells is noted in this photomicrograph. (Right) Hematoxylin and eosin shows numerous CMV intranuclear inclusions ﬈ within endothelial cells in small vessels within the renal sinus of this transplant nephrectomy specimen.

Acute Allograft Glomerulopathy

CMV Virions (Left) Acute allograft glomerulopathy, a form of acute cellular rejection, in a renal transplant recipient with systemic CMV shows endothelial swelling and scattered mononuclear cells in capillary loops. The PAS(+) webs are indicative of mesangiolysis ﬉. No CMV inclusions were present. C4d was negative. (Right) This electron micrograph reveals several clusters of CMV virions that are contained within an electron-lucent membrane ﬈ in the nucleus of this infected epithelial cell. (Courtesy of J. Taxy, MD.)

207

Kidney Transplantation

Histoplasmosis KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Synonyms ○ Ohio Valley disease ○ Darling disease

• Oval to round fungal organisms, 2-4 μm in diameter • Granulomatous interstitial nephritis ○ Noncaseating granulomas ○ Focal necrosis, medulla ○ Prominent interstitial inflammation • Cortical necrosis • Thrombotic microangiopathy • Mesangial proliferative glomerulonephritis (rare)

ETIOLOGY/PATHOGENESIS • Histoplasma capsulatum ○ Dimorphic fungus ○ Endemic to Mississippi and Ohio River Valleys ○ Present in soil ○ Inhalation of airborne conidia (spores)

TOP DIFFERENTIAL DIAGNOSES • • • • • • •

CLINICAL ISSUES • M:F = 4:1

MACROSCOPIC • Discrete nodular masses • Papillary necrosis • Diffuse inflammation and necrosis

Blastomycosis Cryptococcosis Candidiasis Tuberculosis Coccidioidomycosis Sarcoidosis Drug-induced acute interstitial nephritis

Cortical Necrosis

Gomori Methenamine Silver

Histoplasma capsulatum

Histoplasma and Red Blood Cells

(Left) Hematoxylin and eosin shows extensive cortical necrosis at autopsy in a kidney with numerous round Histoplasma organisms due to disseminated disease in a patient with AIDS. A remnant of a tubule is noted ﬈. (Right) Gomori methenamine silver stain highlights numerous fungal organisms from a patient with disseminated histoplasmosis within a necrotic area of the renal cortex.

(Left) Periodic acid-Schiff stain shows numerous clusters of round organisms ﬈ characteristic of Histoplasma capsulatum within the renal tubules. (Right) Gomori methenamine silver stain confirms scattered Histoplasma organisms ﬈, which are smaller in diameter than the red blood cells (6-8 μm in diameter) ﬊ within an adjacent peritubular capillary.

208

Histoplasmosis

MICROSCOPIC

Environmental Exposure

Histologic Features

• Present in soil ○ Bird or bat droppings • Inhalation of airborne conidia (spores)

• Fungi are oval to round, 2-4 μm in diameter • Granulomatous interstitial nephritis ○ Noncaseating granulomas ○ Occasional intratubular granulomas ○ Organisms in macrophages ○ Absence of granulomatous response in severely immunosuppressed patients ○ Focal necrosis, medulla ○ Prominent interstitial inflammation • Cortical necrosis • Thrombotic microangiopathy ○ Glomerular capillary thrombi with entrapped fungal organisms may be observed • Mesangial proliferative glomerulonephritis ○ Rare association with disseminated histoplasmosis ○ H. capsulatum antigen detected in mesangial areas

Infectious Agents • Histoplasma capsulatum ○ Dimorphic fungus (yeast in body, mycelia in soil) ○ Endemic to Mississippi and Ohio River Valleys ○ South and Central America, Africa, Australia, and East Asia

CLINICAL ISSUES Epidemiology • Incidence ○ Kidney involved in 40% with disseminated disease ○ AIDS patients in endemic areas at risk (10-25%) • Sex ○ M:F = 4:1 – Similar gender exposure based on skin tests

Presentation • Asymptomatic renal involvement ○ Renal dysfunction unusual • Flu-like acute respiratory infection • Hemophagocytic syndrome • Chronic form mimics tuberculosis

Laboratory Tests • Radioimmunoassay ○ H. capsulatum polysaccharide antigen • Complement fixation test ○ More sensitive, less specific than immunodiffusion ○ False-positives from cross reactivity with antigens from Blastomyces dermatitidis and Coccidioides immitis • Immunodiffusion • Culture • Biopsy

Treatment • Drugs ○ Amphotericin B ○ Ketoconazole ○ Itraconazole ○ Fluconazole ○ Decreased immunosuppression

Prognosis • Disseminated form fatal without treatment ○ May lead to irreversible renal failure • Localized form in normal individuals (self-limiting)

MACROSCOPIC

Kidney Transplantation

ETIOLOGY/PATHOGENESIS

DIFFERENTIAL DIAGNOSIS Blastomycosis • 8-15 μm in diameter • Thick wall and broad-based budding

Cryptococcosis • 5-10 μm • Capsule-deficient variant causes granulomatous inflammation ○ Fontana-Masson silver stain positive

Candidiasis • Budding yeasts may resemble Histoplasma

Coccidioidomycosis • 30-60 μm in diameter • Endospores (2-5 μm in diameter) resemble Histoplasma

Tuberculosis • Caseating granulomas with acid-fast bacilli

Sarcoidosis • Noncaseating granulomas without microorganisms

Drug-Induced Acute Interstitial Nephritis • No microorganisms

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Necrotizing granulomas • Yeast forms smaller than erythrocytes

SELECTED REFERENCES 1.

General Features

2.

• Discrete nodular masses • Papillary necrosis • Diffuse inflammation and necrosis

3. 4.

Guimarães LF et al: Invasive fungal disease in renal transplant recipients at a Brazilian center: local epidemiology matters. Transplant Proc. 48(7):23062309, 2016 Nieto-Ríos JF et al: Histoplasmosis in renal transplant patients in an endemic area at a reference hospital in Medellin, Colombia. Transplant Proc. 46(9):3004-9, 2014 Nieto-Ríos JF et al: Disseminated histoplasmosis and haemophagocytic syndrome in two kidney transplant patients. Nefrologia. 32(5):683-4, 2012 Sethi S: Acute renal failure in a renal allograft: an unusual infectious cause of thrombotic microangiopathy. Am J Kidney Dis. 46(1):159-62, 2005

209

Kidney Transplantation

Candidiasis KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Candida infection of kidney ○ Typically in immunocompromised patients

• Fungal organisms ○ Pseudohyphae ○ Budding yeast forms • Cortical abscesses ○ May be centered around glomeruli ○ Rare cortical infarcts • Granulomatous interstitial inflammation

ETIOLOGY/PATHOGENESIS • Candida albicans, Candida glabrata, Candida parapsilosis, and Candida tropicalis

CLINICAL ISSUES • Therapeutic options ○ Fluconazole, amphotericin B, reduction of immunosuppressive agents, transplant nephrectomy

MACROSCOPIC • • • •

Cortical abscesses Papillary necrosis Pyelitis Mycotic pseudoaneurysm

TOP DIFFERENTIAL DIAGNOSES • • • •

Aspergillosis Mucormycosis Fusariosis Pseudallescheriasis

DIAGNOSTIC CHECKLIST • Budding yeast and pseudohyphae combination characteristic • Fungal cultures needed for definitive identification

Microabscesses

Candida Colonization of Papilla

Granulomatous Inflammation

Candida Yeast and Pseudohyphae

(Left) Gross photograph of the capsular surface of this kidney at autopsy shows many small abscesses ſt due to disseminated candidiasis from Candida tropicalis. (Right) Gross photograph shows prominent small and large green lesions ﬈ on the renal papilla of this autopsy specimen representing colonization by Candida. (Courtesy C. Abrahams, MD.)

(Left) Fungal organisms ſt are intermixed with TammHorsfall protein (uromodulin) and show focal necrosis and rupture of the tubules ﬈ associated with a granulomatous reaction ﬉ and scattered interstitial lymphocytes ﬊. The glomerulus st is normal. (Right) Gomori methenamine silver stain shows yeast forms ﬈ within the renal tubules and a few pseudohyphae ﬇, a helpful finding in the identification of Candida.

210

Candidiasis

MACROSCOPIC

Definitions

General Features

• Candida infection of kidney ○ Typically in immunocompromised patients

• Abscesses ○ Cortical abscesses – Miliary distribution ○ Perinephric abscesses • Pyelitis ○ Fungus balls • Papillary necrosis ○ 20% of patients with disseminated candidiasis at autopsy • Mycotic pseudoaneurysm or occlusion of major renal arteries in allograft

ETIOLOGY/PATHOGENESIS Infectious Agents • Candida albicans ○ Normal flora of skin, gastrointestinal, and genitourinary tracts • Candida glabrata ○ Previously named Torulopsis glabrata (or torulopsosis) • Candida parapsilosis • Candida tropicalis • Candida krusei

CLINICAL ISSUES Epidemiology • Incidence ○ 8 cases per 100,000 persons in United States ○ 4th most common nosocomial bloodstream infection ○ Risk factors include diabetes, chemotherapy, immunosuppression • Age ○ Neonates – Very low birth weight babies susceptible to invasive candidiasis ○ Elderly – > 65 years • Ethnicity ○ Higher incidence among African Americans

Presentation • Renal dysfunction or acute kidney injury ○ May be due to ureteral or bladder obstruction by Candida

Kidney Transplantation

TERMINOLOGY

MICROSCOPIC Histologic Features • Cortical abscesses ○ May be centered around glomeruli ○ Rare cortical infarcts • Fungal organisms ○ Pseudohyphae ○ Budding yeast forms, 4-6 μm ○ May be in glomerular capillaries &/or arterioles • Granulomatous interstitial nephritis ○ Multinucleated giant cells (atypical feature) ○ May contain predominantly budding yeast forms

DIFFERENTIAL DIAGNOSIS Aspergillosis • Septate hyphae with 45° branching

Mucormycosis • Nonseptate hyphae with 90° branching

Cryptococcosis • Capsule-deficient variant mimics Candida yeast forms

Fusariosis

Laboratory Tests

• Septate hyphae

• Fungal culture • Direct microscopy

Pseudallescheriasis • Septate hyphae

Treatment • Drugs ○ Fluconazole ○ Echinocandins ○ Voriconazole ○ Amphotericin B • Reduction of immunosuppression • Renal allograft nephrectomy

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Budding yeast and pseudohyphae combination characteristic • Fungal cultures needed for definitive identification

SELECTED REFERENCES

Prognosis

1.

• Up to 50% mortality in disseminated or invasive candidiasis

2.

IMAGING CT Findings • Hypodense lesions correspond to renal abscesses • Fungus balls • Papillary necrosis

3. 4. 5.

Patel MH et al: Invasive fungal infections in renal transplant patients: a single center study. Ren Fail. 39(1):294-298, 2017 Andes DR et al: The epidemiology and outcomes of invasive Candida infections among organ transplant recipients in the United States: results of the Transplant-Associated Infection Surveillance Network (TRANSNET). Transpl Infect Dis. 18(6):921-931, 2016 Bagnasco SM et al: Fungal infection presenting as giant cell tubulointerstitial nephritis in kidney allograft. Transpl Infect Dis. 14(3):288-91, 2012 Wasi N et al: A rare case of acute renal failure due to massive renal allograft infiltration with Candida glabrata. Nephrol Dial Transplant. 23(1):374-6, 2008 Meehan SM et al: Granulomatous tubulointerstitial nephritis in the renal allograft. Am J Kidney Dis. 36(4):E27, 2000

211

Kidney Transplantation

Cryptococcosis KEY FACTS

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• Cryptococcus neoformans • Cryptococcus gattii ○ Found in Pacific Northwest of USA and Canada • Present in soil • Inhalation of airborne fungal forms

• Granulomatous interstitial inflammation • Fungal organisms ○ Clear halo; mucicarmine, PAS, and silver stain positive – Capsule-deficient variant of Cryptococcus (all silver stains) ○ 5-10 μm in diameter ○ Narrow-based budding • Tubulointerstitial inflammation with tubulitis

CLINICAL ISSUES • 1/100,000 in general population • 2.8-5.0% in solid organ transplant patients ○ 0.8-5.8% in renal transplant patients • Rare in children before puberty • Male predilection • Amphotericin B • Fluconazole

MACROSCOPIC • Papillary necrosis ○ May be present in cryptococcal pyelonephritis

TOP DIFFERENTIAL DIAGNOSES • • • • • • • •

Blastomycosis Candidiasis Histoplasmosis Coccidioidomycosis Paracoccidioidomycosis Sarcoidosis Drug-induced acute interstitial nephritis Tuberculosis

Granulomatous Interstitial Nephritis

Multinucleated Giant Cells

Gomori Methenamine Silver

Mucicarmine

(Left) Hematoxylin and eosin shows prominent granulomatous interstitial inflammation that involves primarily the renal medulla in a kidney transplant patient. (Right) Hematoxylin and eosin shows large, multinucleated giant cells ﬈ and Cryptococci with characteristic clear halos ﬊ that are distinct from the adjacent foamy macrophages in this kidney allograft, which ultimately resulted in a transplant nephrectomy.

(Left) Gomori methenamine silver shows strong staining of numerous round cryptococcal organisms ﬈ within the prominent granulomatous inflammation throughout the renal cortex. (Right) Mucicarmine stain highlights the thick outer capsule ﬈ that characterizes Cryptococcus.

212

Cryptococcosis

MICROSCOPIC

Definitions

Histologic Features

• Cryptococcal infection, typically in immunocompromised patients

• Granulomatous interstitial inflammation ○ Absence of significant granulomatous or inflammatory response in severely immunocompromised patients • Fungal organisms ○ Capsule is mucicarmine, PAS, and silver stain positive ○ Capsule-deficient variant of Cryptococcus (all silver stains) ○ 5-10 μm in diameter ○ Narrow-based budding ○ May be present in glomerular capillaries within macrophages • Prominent tubulointerstitial inflammation • Tubulitis • Necrotizing and crescentic glomerulonephritis ○ Single report in association with pulmonary cryptococcosis with resolution after antifungal therapy

ETIOLOGY/PATHOGENESIS Environmental Exposure • Present in soil • Inhalation of airborne fungal forms

Infectious Agents • Cryptococcus neoformans • Cryptococcus gattii ○ Found in Pacific Northwest of USA and Canada ○ Isolated from eucalyptus trees in subtropical and tropical regions • Rare isolates ○ Cryptococcus laurentii, Cryptococcus albidus

Pathogenetic Factors • Polysaccharide capsule, melanin, urease, laccases, and phospholipase B

CLINICAL ISSUES Epidemiology • Incidence ○ 1/100,000 in general population – 2-7/1,000 in AIDS patients ○ 2.8-5.0% in solid organ transplant patients – 0.8-5.8% in renal transplant patients ○ Common cause of meningoencephalitis in sub-Saharan Africa • Age ○ Rare in children before puberty • Sex ○ Male predilection

Presentation • Acute renal failure • Proteinuria

Laboratory Tests • Cryptococcal antigen test • Fungal culture

Treatment • Drugs ○ Amphotericin B, fluconazole • Reduction of immunosuppressive agents in transplant patients

Prognosis • Graft loss in 9% of renal transplant patients

MACROSCOPIC

DIFFERENTIAL DIAGNOSIS Blastomycosis • 8-15 μm in diameter • Broad-based budding

Candidiasis • Budding yeasts • Pseudohyphae

Histoplasmosis • 2-4 μm in diameter • Endemic to Mississippi and Ohio River Valleys

Coccidioidomycosis • Spherules with characteristic endospores • Endemic to Southwest USA, Mexico, and South America

Paracoccidioidomycosis • Endemic to South America and Brazil • Clear halo, Gomori methenamine silver positive

Tuberculosis • Caseating necrosis • Acid-fast bacilli present

SELECTED REFERENCES 1. 2.

3.

4.

5. 6.

General Features • Papillary necrosis ○ May be present in cryptococcal pyelonephritis

Kidney Transplantation

TERMINOLOGY

7.

Marques S et al: Cryptococcosis in renal transplant recipients: a single-center experience. Transplant Proc. 48(7):2289-2293, 2016 Pongmekin P et al: Clinical characteristics and mortality risk factors of cryptococcal infection among HIV-negative patients. J Med Assoc Thai. 97(1):36-43, 2014 Yang YL et al: Cryptococcosis in kidney transplant recipients in a Chinese university hospital and a review of published cases. Int J Infect Dis. 26:15461, 2014 Silveira FP et al: Cryptococcosis in liver and kidney transplant recipients receiving anti-thymocyte globulin or alemtuzumab. Transpl Infect Dis. 9(1):22-7, 2007 Iglesias JI et al: AIDS, nephrotic-range proteinuria, and renal failure. Kidney Int. 69(11):2107-10, 2006 Nakayama M et al: A case of necrotizing glomerulonephritis presenting with nephrotic syndrome associated with pulmonary cryptococcosis. Clin Exp Nephrol. 9(1):74-8, 2005 Singh N et al: Allograft loss in renal transplant recipients with cryptococcus neoformans associated immune reconstitution syndrome. Transplantation. 80(8):1131-3, 2005

213

Kidney Transplantation

Mucormycosis KEY FACTS

TERMINOLOGY • Fungus infection, typically in immunocompromised patients • Synonym ○ Invasive zygomycosis

ETIOLOGY/PATHOGENESIS • Mucormycetes ○ Order: Mucorales – Most common genera (in descending frequency): Rhizopus, Rhizomucor, Cunninghamella, Apophysomyces, Saksenaea, Absidia, Mucor • Risk factors: Diabetes, immunosuppression, malnutrition, malignancy

CLINICAL ISSUES • • • •

Renal involvement in 20% of disseminated disease patients Fever, flank pain Rapidly progressive Complete debridement of infected tissue

• Amphotericin B • Reduction of immunosuppression

MACROSCOPIC • Large infarcts • Arterial thrombosis ○ Main renal, arcuate, or interlobar artery

MICROSCOPIC • • • • •

Nonseptate fungal hyphae with 90° branching Cortical necrosis Thrombi Microabscesses Granulomatous interstitial nephritis

TOP DIFFERENTIAL DIAGNOSES • Aspergillosis • Candidiasis • Pseudallescheriasis or fusariosis

Disseminated Mucormycosis

Angioinvasive Mucormycosis

Glomerular Fungi

Gomori Methenamine Silver

(Left) Jones methenamine silver stain shows numerous hyphae ﬈, typical of mucormycosis in an area of cortical necrosis. (Courtesy E. Bracamonte, MD.) (Right) Jones methenamine silver stain shows numerous fungal nonseptate branching hyphae ﬈ invading the arterial lumen in an area with extensive cortical necrosis. (Courtesy E. Bracamonte, MD.)

(Left) Fungal organisms that are consistent with Rhizopus have widely disseminated through the kidneys and other organs at autopsy of an immunocompromised patient after hematopoietic stem cell transplantation. A thrombus ſt is distending a glomerular capillary. (Right) Many hyphae, some with right angle branching ſt, are noted at autopsy of this 57-year-old man with disseminated Rhizopus and stem cell transplantation for acute myelogenous leukemia.

214

Mucormycosis

MACROSCOPIC

Synonyms

General Features

• Invasive zygomycosis

• Arterial thrombosis ○ Main renal artery ○ Arcuate &/or interlobar arteries • Large infarcts • Papillary necrosis (rare)

Definitions • Fungal infection typically in immunocompromised patients

ETIOLOGY/PATHOGENESIS Infectious Agents • Mucormycetes ○ Order: Mucorales – Most common genus: Rhizopus – Other genera (in descending frequency): Rhizomucor, Cunninghamella, Apophysomyces, Saksenaea, Absidia, Mucor ○ Ubiquitous organisms found in soil and decaying matter, including moldy bread

CLINICAL ISSUES Epidemiology • Incidence ○ Risk factors – Diabetes mellitus – Leukemia, solid malignancies – Immunosuppression – Malnourishment

Site • Sinus and brain (rhinocerebral): ~ 50% of cases • Lungs, skin • GI, kidney: Less common

MICROSCOPIC Histologic Features • Nonseptate fungal hyphae with 90° branching ○ Found in infarcts, granulomata, thrombi, microabscesses • Cortical necrosis • Thrombi • Arteritis • Microabscesses • Granulomatous interstitial nephritis ○ Frequent multinucleated giant cells

DIFFERENTIAL DIAGNOSIS Aspergillosis • Septate hyphae with 45° branching

Pseudallescheriasis • Septate hyphae

Fusariosis • Septate hyphae

Candidiasis • Pseudohyphae and budding yeast forms

Presentation

Tuberculosis

• • • • •

• Caseating necrosis in granulomas with acid-fast bacilli

Acute kidney injury Fever Flank pain Rapidly progressive course Hematuria

Laboratory Tests • Fungal cultures • Biopsy

Bacterial Pyelonephritis • Prominent neutrophilic infiltrate or abscesses • No fungal organisms

Sarcoidosis • Granulomatous inflammation without microorganisms

DIAGNOSTIC CHECKLIST

Treatment

Pathologic Interpretation Pearls

• Surgical approaches ○ Complete debridement of infected tissue ○ Allograft nephrectomy • Drugs ○ Antifungal therapy – Amphotericin B ○ Reduction of immunosuppressive agents

• Difficult to distinguish fungal species based on morphologic evaluation alone • Fungal cultures establish diagnosis

Prognosis • Poor: > 50% mortality in disseminated disease ○ Mucormycosis isolated to kidney may have better prognosis

Kidney Transplantation

TERMINOLOGY

SELECTED REFERENCES 1. 2. 3. 4. 5. 6.

Song Y et al: Mucormycosis in renal transplant recipients: review of 174 reported cases. BMC Infect Dis. 17(1):283, 2017 Dhaliwal HS et al: Diagnosed only if considered: isolated renal mucormycosis. Lancet. 385(9984):2322, 2015 Park W et al: Allograft mucormycosis due to Rhizopus microsporus in a kidney transplant recipient. Transplant Proc. 46(2):623-5, 2014 Gupta KL et al: Mucormycosis of the transplanted kidney with renal papillary necrosis. Exp Clin Transplant. 11(6):554-7, 2013 Kuy S et al: Renal mucormycosis: a rare and potentially lethal complication of kidney transplantation. Case Rep Transplant. 2013:915423, 2013 Gupta KL et al: Renal zygomycosis: an under-diagnosed cause of acute renal failure. Nephrol Dial Transplant. 14(11):2720-5, 1999

215

Kidney Transplantation

Aspergillosis KEY FACTS ○ Voriconazole

ETIOLOGY/PATHOGENESIS • Aspergillus species, ubiquitous fungi in environment ○ A. fumigatus, A. flavus, A. niger

MACROSCOPIC

CLINICAL ISSUES

MICROSCOPIC

• Incidence ○ 0.1% in kidney transplant patients after 1 year ○ 30-40% renal involvement in disseminated aspergillosis – 50-100% mortality rate for invasive aspergillosis • Presentation ○ Fever ○ Flank pain ○ Hematuria • Laboratory tests ○ Cultures • Treatment ○ Renal allograft nephrectomy ○ Nephrostomy drainage and systemic antifungal therapy

• Microorganisms, fungus ○ Septate hyphae with 45° angle branching ○ Vascular invasion • Tubulointerstitial inflammation, neutrophil rich

• Abscesses, cortical or perinephric

TOP DIFFERENTIAL DIAGNOSES • • • • •

Candidiasis Mucormycosis Pseudallescheriasis Fusariosis Bacterial pyelonephritis

Cortical Necrosis

Glomerular Aspergillosis

Gomori Methenamine Silver

Gomori Methenamine Silver

(Left) Hematoxylin & eosin shows a necrotic area of renal cortex with prominent neutrophilic inflammation and many acute angle branching hyphae ﬈ that are characteristic of Aspergillus. (Right) Hematoxylin & eosin shows septate hyphae ﬈ with 45° angle branching, which has entirely replaced a glomerulus in this autopsied kidney of a patient with disseminated aspergillosis.

(Left) Gomori methenamine silver shows a fungus ball consisting of numerous septate hyphae ﬈ with acute angle branching in the renal medulla of a patient with disseminated aspergillosis. (Right) Gomori methenamine silver reveals hyphal elements characteristic of Aspergillus ﬈ within a glomerulus. The diameter of Aspergillus hyphae is uniform and less broad than those of pseudallescheriasis or mucormycosis. Septate hyphae with acute angle branching help to exclude mucormycosis.

216

Aspergillosis

Definitions • Aspergillus infection of kidney in immunosuppressed or immunocompromised patients

ETIOLOGY/PATHOGENESIS Environmental Exposure

IMAGING CT Findings • Hypodense lesions in kidney

MACROSCOPIC

• Ubiquitous fungus in environment

General Features

Infectious Agents

• Abscesses ○ Cortical ○ Perinephric

• Aspergillus ○ A. fumigatus ○ A. flavus ○ A. niger ○ A. terreus ○ A. nidulans

CLINICAL ISSUES Epidemiology • Incidence ○ 0.1% in kidney transplant patients after 1 year • Sex ○ M:F = 4:1

Site • Kidneys ○ 30-40% involvement in disseminated aspergillosis ○ Isolated involvement in some deceased donor allografts – Probable transmission from deceased donor or during organ procurement – Fungi account for up to 2.5% of isolates cultured from perfusion solutions used for kidney preservation

MICROSCOPIC Histologic Features • Necrosis • Suppurative inflammation • Fungal organisms ○ Septate hyphae with 45° angle branching ○ 3-4 μm uniform diameter of hyphae ○ Vascular invasion • Thrombosis ○ Hemorrhagic infarcts

DIFFERENTIAL DIAGNOSIS Candidiasis • Pseudohyphae and budding yeast forms

Mucormycosis • Nonseptate hyphae with 90° angle branching

Pseudallescheriasis

Presentation

• Septate hyphae

• Fever • Flank pain • Hematuria

Fusariosis

Laboratory Tests

• Caseating granulomas with acid-fast bacilli

• Serologic tests ○ Enzyme-linked immunoassay – Detection of galactomannan antigen of Aspergillus ○ Immunodiffusion ○ Complement fixation • Cultures • Direct microscopy

• Septate hyphae

Tuberculosis

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Fungal cultures useful to confirm diagnosis

SELECTED REFERENCES 1.

Treatment • Surgical approaches ○ Renal allograft nephrectomy ○ Nephrostomy drainage and systemic antifungal therapy • Drugs ○ Voriconazole: 1st-line agent for invasive aspergillosis ○ Itraconazole, amphotericin B • Reduction of immunosuppressive agents in transplant patients

Prognosis • 50-100% mortality rate for invasive aspergillosis • Risk factors for mortality

Kidney Transplantation

○ Disseminated infection, leukopenia, serum galactomannan level

TERMINOLOGY

2. 3.

4. 5. 6.

7.

Sadagah L et al: Renal allograft Aspergillus infection presenting with obstructive uropathy: A case report. Transplant Proc. 49(1):193-197, 2017 Desbois AC et al: Prognosis of invasive aspergillosis in kidney transplant recipients: A case-control study. Transplant Direct. 2(8):e90, 2016 Guimarães LF et al: Invasive fungal disease in renal transplant recipients at a Brazilian center: Local epidemiology matters. Transplant Proc. 48(7):23062309, 2016 Heylen L et al: Invasive aspergillosis after kidney transplantation: CaseControl Study. Clin Infect Dis. ePub, 2015 Hoyo I et al: Epidemiology, clinical characteristics, and outcome of invasive aspergillosis in renal transplant patients. Transpl Infect Dis. 16(6):951-7, 2014 Meng XC et al: Renal aspergillosis after liver transplantation: Clinical and imaging manifestations in two cases. World J Gastroenterol. 20(48):18495502, 2014 Singh N et al: Donor-derived fungal infections in organ transplant recipients: guidelines of the American Society of Transplantation, infectious diseases community of practice. Am J Transplant. 12(9):2414-28, 2012

217

Kidney Transplantation

Coccidioidomycosis KEY FACTS

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• Coccidioides immitis ○ Limited to California San Joaquin Valley, Southwestern United States, and Mexico • Coccidioides posadasii ○ Limited to Southwestern United States, Mexico, and South America • Acquired through inhalation of fungal spores (arthroconidia) in environment • Dimorphic fungi

• Coccidioides spherules ○ Round, thick walled (PAS and methenamine silver positive) ○ 10-80 μm in diameter ○ Contain numerous endospores (2-5 μm in diameter) • Necrotic nodules ○ Septate hyphae seen in transitional form • Granulomatous interstitial inflammation ○ Absent in severely immunocompromised patients

CLINICAL ISSUES

TOP DIFFERENTIAL DIAGNOSES

• Flu-like symptoms • Surgical resection for some with pulmonary, bone, or joint involvement • Fluconazole: 1st-line agent

• Blastomycosis • Cryptococcosis

MACROSCOPIC

DIAGNOSTIC CHECKLIST • Spherules and endospores characteristic for Coccidioides • Rare renal involvement

• Perinephric abscess

Immature Coccidioides Spherules

Mature Spherule With Endospores

Numerous Immature Spherules and Endospores

Immature Spherules and Endospores

(Left) Jones methenamine silver shows numerous immature Coccidioides spherules ﬈ compressing the glomerulus ﬊ within the urinary space in a patient with disseminated disease. (Courtesy E. Bracamonte, MD.) (Right) Periodic acid-Schiff shows a thick-walled mature spherule ﬈ with endospores ﬊ characteristic of Coccidioides among the renal tubules (possibly within a peritubular capillary) of this autopsy kidney. (Courtesy E. Bracamonte, MD.)

(Left) Periodic acid-Schiff shows numerous immature spherules ﬈ and endospores ﬉ that are characteristic of Coccidioides immitis within the Bowman space compressing the adjacent glomerulus ﬊. (Courtesy E. Bracamonte, MD.) (Right) Jones methenamine silver shows immature spherules ﬈ and endospores ﬉ of Coccidioides within the urinary space and compressing the adjacent glomerulus ﬊. (Courtesy E. Bracamonte, MD.)

218

Coccidioidomycosis

Synonyms • Valley fever

ETIOLOGY/PATHOGENESIS Infectious Agents • Coccidioides immitis ○ Geographically limited to California San Joaquin Valley, Southwestern United States, and Mexico • Coccidioides posadasii ○ Geographically limited to Southwestern United States, Mexico, and South America • Acquired through inhalation of fungal spores (arthroconidia) • Dimorphic fungi

CLINICAL ISSUES Epidemiology • Incidence ○ ~ 3% among renal transplant patients in endemic regions ○ Typically occurs within 1st year post transplantation • Sex ○ Pregnancy is risk factor for disseminated disease • Ethnicity ○ African, Asian, and Hispanic descent more likely than Caucasians to develop disseminated disease

Presentation • Flu-like symptoms ○ Fever ○ Cough ○ Myalgia ○ Rash • Eosinophilia • Acute renal failure

Prognosis • Good in limited disease • Poor in disseminated disease ○ Mortality rate > 50% ○ Up to 75% mortality rate in transplant patients

IMAGING Radiographic Findings • Pyelocalyceal alterations similar to tuberculosis in transplant kidneys • Compression of renal transplant artery may be detected by angiogram

MACROSCOPIC General Features • Perinephric abscess

MICROSCOPIC Histologic Features • Coccidioides spherules ○ Round, thick walled (PAS and methenamine silver positive) ○ 10-80 μm in diameter ○ Contain numerous endospores (2-5 μm in diameter) • Necrotic nodules ○ Septate hyphae seen in transitional form • Granulomatous interstitial inflammation ○ Absent in severely immunocompromised patients

DIFFERENTIAL DIAGNOSIS

Laboratory Tests

Blastomycosis

• Skin test ○ 10-50% of those in endemic areas test positive ○ Coccidioidin ○ Spherulin • Enzyme immunoassay ○ IgA ○ IgM – Antibodies difficult to detect in immunosuppressed patients • Immunodiffusion assay • Complement fixation test • Culture ○ Sputum ○ Other body fluids • Direct microscopy

• 8-15 μm in diameter

Treatment • Surgical approaches ○ Surgical resection for some with pulmonary, bone, or joint involvement • Drugs ○ Fluconazole

Kidney Transplantation

○ Amphotericin B – 2nd-line agent – Used for disseminated disease or azole-resistant strains of Coccidioides ○ Reduction of immunosuppressive agents

TERMINOLOGY

Cryptococcosis • Capsule-deficient variant, silver stain positive • No endospores

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Spherules and endospores characteristic for Coccidioides • Rare renal involvement

SELECTED REFERENCES 1.

2.

3.

Singh N et al: Donor-derived fungal infections in organ transplant recipients: guidelines of the American Society of Transplantation, infectious diseases community of practice. Am J Transplant. 12(9):2414-28, 2012 Baden LR et al: Case records of the Massachusetts General Hospital. Case 352009. a 60-year-old male renal-transplant recipient with renal insufficiency, diabetic ketoacidosis, and mental-status changes. N Engl J Med. 361(20):1980-9, 2009 Braddy CM et al: Coccidioidomycosis after renal transplantation in an endemic area. Am J Transplant. 6(2):340-5, 2006

219

Kidney Transplantation

Paracoccidioidomycosis KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Synonyms ○ South American blastomycosis ○ Brazilian blastomycosis

• • • • •

ETIOLOGY/PATHOGENESIS • Paracoccidioides brasiliensis ○ Endemic to South America and Brazil

CLINICAL ISSUES • Male predilection • Laboratory testing ○ Enzyme-linked immunoassay ○ Complement fixation test ○ Culture ○ Biopsy • Drugs ○ Itraconazole: 1st-line agent

Granulomatous interstitial nephritis Glomerular granulomas Glomerular capillary thrombi Glomerular fibrinoid necrosis Yeast forms with clear halos

TOP DIFFERENTIAL DIAGNOSES • • • • • •

Cryptococcosis Lobomycosis Blastomycosis Tuberculosis Sarcoidosis Drug-induced acute interstitial nephritis

Granulomatous Inflammation

Granulomata

Gomori Methenamine Silver

Electron Microscopy

(Left) Light microscopy shows a granuloma in the cortex with prominent Langhans giant cells ﬈ surrounded by mononuclear cells in a Brazilian male with renal failure. (Courtesy A. Billis, MD.) (Right) Periodic acidSchiff shows interstitial and glomerular granulomas ﬈ with glomerular destruction ﬊ in a severe case with a miliary pattern of systemic involvement. Lungs, lymph nodes, and oral mucosa are commonly involved and, less commonly, the kidney, spleen, bones, and meninges. (Courtesy A. Billis, MD.)

(Left) Grocott-Gomori methenamine silver stain of granulomas containing fungal bodies shows strong peripheral silver uptake ﬈. An adjacent glomerulus contains a granuloma ﬊. The mechanism of glomerular involvement is believed to be embolic lodging of fungi in the capillaries with subsequent thrombus and inflammation. (Courtesy A. Billis, MD.) (Right) Electron micrograph of mononuclear giant cells shows 3 Paracoccidioides brasiliensis fungal organisms ſt with a clear halo. (Courtesy A. Billis, MD.)

220

Paracoccidioidomycosis

Abbreviations • Paracoccidioidomycosis (PCM)

Synonyms • South American blastomycosis • Brazilian blastomycosis

ETIOLOGY/PATHOGENESIS Environmental Exposure • Agricultural and construction workers

Infectious Agents • Paracoccidioides brasiliensis ○ Endemic in South America ○ Dimorphic fungus – Septate hyphae at room temperature – Yeast forms at body temperature ○ Acquired through inhalation

CLINICAL ISSUES Epidemiology • Incidence ○ Rare • Age ○ Usually > 30 years ○ Rare in children or teenagers • Sex ○ Strong male predilection – M:F = 15-78:1

Presentation

Treatment • Drugs ○ Itraconazole has low rate of relapse ○ Ketoconazole, sulfonamide, amphotericin B ○ Reduction of immunosuppressive agents in transplant patients

Prognosis • Poor in rare juvenile form of disease • Untreated: Up to 25% mortality rate • Treated: Good prognosis

MICROSCOPIC Histologic Features • Glomeruli ○ Granulomas – May resemble cellular crescents ○ Fibrinoid necrosis ○ Capillary thrombi • Tubules and Interstitium ○ Granulomatous interstitial nephritis – Associated with Paracoccidioides organisms – Caseating necrosis occasionally present – Multinucleated giant cells ○ Prominent acute inflammation – Pyogenic abscesses ○ Fungal organisms – Characteristic clear halos – Budding yeast forms, 12-14 μm in diameter • Vessels: No specific lesions

DIFFERENTIAL DIAGNOSIS

• Asymptomatic • Acute renal failure

Cryptococcosis

Laboratory Tests

Blastomycosis

• Skin test ○ Positive result indicates exposure not active disease • Enzyme-linked immunoassay ○ Detects antibodies to gp43 – High sensitivity and specificity • Complement fixation test ○ May cross react with Histoplasma capsulatum antigen • Immunodiffusion • Western blot ○ High sensitivity and specificity • Fungal culture ○ Sabouraud dextrose agar ○ Up to 30 days of growth – Growth of yeast form at 37°C confirms diagnosis • Direct microscopy ○ Wet mount with potassium hydroxide – 1 large yeast with budding forms resembles "pilot wheel" • Biopsy

• 8-15 μm in diameter • DNA confirmation probe may cross react with Paracoccidioides

Natural History • Generally asymptomatic in immunocompetent hosts

Kidney Transplantation

TERMINOLOGY

• Capsule positive for mucicarmine stain

Tuberculosis • Caseating granulomas • Acid-fast bacilli present • Coinfection with PCM reported

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Fungal culture or serologic tests confirm infection by P. brasiliensis

SELECTED REFERENCES 1. 2. 3. 4.

Góes HF et al: Paracoccididoidomycosis in a renal transplant recipient. Rev Inst Med Trop Sao Paulo. 58:12, 2016 Batista MV et al: Recipient of kidney from donor with asymptomatic infection by Paracoccidioides brasiliensis. Med Mycol. 50(2):187-92, 2012 Zavascki AP et al: Paracoccidioidomycosis in organ transplant recipient: case report. Rev Inst Med Trop Sao Paulo. 46(5):279-81, 2004 Shikanai-Yasuda MA et al: Paracoccidioidomycosis in a renal transplant recipient. J Med Vet Mycol. 33(6):411-4, 1995

221

Kidney Transplantation

Microsporidiosis KEY FACTS

TERMINOLOGY • Infection by 1 of many species of Microsporidia fungi particularly affecting immunocompromised hosts

ETIOLOGY/PATHOGENESIS • Kidney involved in minority of cases, typically as part of systemic infection • 14 species infect humans • Encephalitozoon intestinalis and Encephalitozoon cuniculi most common organisms in disseminated cases

CLINICAL ISSUES • • • • • •

• Outcome good if treated with antibiotics and immunocompetence can be improved

MICROSCOPIC • Acute and chronic interstitial nephritis • Intraluminal and intracellular 1- x 2-μm ovoid spores in aggregates in tubules ○ Purple with Brown-Hopps stain ○ Giemsa-positive central body • EM identification by unique polar tube • Acute and chronic interstitial nephritis

ANCILLARY TESTS

Diarrhea usual presentation Acute or chronic renal failure Renal allograft dysfunction Fever Other sites: Lung, brain, heart, liver, eye Treatment: Fumagillin, albendazole

• EM reveals pathognomonic coiled polar tube in spores

TOP DIFFERENTIAL DIAGNOSES • Toxoplasmosis • Candidiasis and other fungal infections

Tubulointerstitial Nephritis

Chronic Tubulointerstitial Nephritis

Intracellular Spores

Polar Tube Coil

(Left) Acute and chronic interstitial nephritis due to microsporidiosis is shown. Tubules contain inflammatory cells ﬈, debris, and organisms. The tubular epithelium is ballooned and contains aggregates of organisms ſt. (Right) PAS stain of active interstitial nephritis in an HIV-positive patient with microsporidiosis shows that the organisms in tubules ﬈ are not strongly stained, which allows differentiation from usual fungi.

(Left) Electron micrograph shows intracellular Microsporidia spores ﬈ in tubular epithelium. (Right) High-power electron micrograph shows the pathognomonic polar tube coils ﬈, a unique structure of Microsporidia spores. Here, 5 coils are present, typical of Encephalitozoon intestinalis.

222

Microsporidiosis

Definitions • Infection by 1 of many species of Microsporidia fungi particularly affecting immunocompromised hosts

ETIOLOGY/PATHOGENESIS Infectious Agents • Microsporidia ○ Ubiquitous obligate intracellular eukaryotic pathogens – Unique group of fungi with reduced cell organelles (no mitochondria, Golgi) ○ Infects all phyla of organisms – Initially identified as silkworm pathogen in 1857 and as human pathogen 50 years ago ○ 14 species infect humans – Most common: Enterocytozoon bieneusi, Encephalitozoon intestinalis, and Encephalitozoon cuniculi ○ Unique method of inserting spore contents into cell via polar tube that behaves like hypodermic needle • Kidney involved in minority of cases, typically as part of systemic infection ○ E. intestinalis and E. cuniculi most common organisms in disseminated cases ○ E. cuniculi infects dogs and cats • E. bieneusi limited to GI tract and hepatobiliary system • Immunodeficiency predisposes to disease ○ HIV patients (CD4 < 100/mm³), transplant recipients (3 weeks to 7 years post transplant) • Transmission in donor kidneys, liver and heart reported

CLINICAL ISSUES Presentation • Acute renal failure • Chronic renal failure • Renal allograft dysfunction ○ Kidney involved in ~ 40% of recipients with microsporidiosis • Fever • Diarrhea • Weight loss • Other sites: Lung, brain, heart, liver, eye

○ Intraluminal and intracellular 1- x 2-μm ovoid spores in aggregates – No budding or pseudohyphae ○ Purple with Gram stain (Brown-Hopps, Brown-Brenn) ○ Giemsa stains central body (nucleus) ○ PAS weakly stained in contrast to other fungi ○ Silver positivity in punctate pattern (posterior body) ○ Positive acid-fast stain (red with Ziehl-Neelsen) ○ Calcofluor white fluorochrome stains cell wall polysaccharide (fluoresces blue with DAPI filter) • Interstitium ○ Acute and chronic inflammation ○ Neutrophils, eosinophils, monocytes, lymphocytes, plasma cells • Glomeruli ○ No specific feature ○ HIV-associated glomerular disease may be present • Vessels ○ No specific feature

ANCILLARY TESTS Immunohistochemistry • Negative for Toxoplasma gondii antigens

PCR • Identification of species in paraffin-embedded tissues

Electron Microscopy • Intracellular spores ○ Distinctive and pathognomonic coiled polar tube in spore – Allows speciation

DIFFERENTIAL DIAGNOSIS Toxoplasmosis • Similar size • Negative on Brown-Brenn or Brown-Hopps stain • Positive for anti-Toxoplasma antigens by IHC

Candidiasis and Infection by Other Fungi • PAS-positive cell walls • Budding spores, pseudohyphae

SELECTED REFERENCES

Treatment

1.

• Drugs ○ Fumagillin, albendazole ○ Tapering immunosuppressive drugs

2. 3.

Prognosis • Good if treated with antibiotics and immunocompetence can be improved

MICROSCOPIC Histologic Features • Acute and chronic interstitial nephritis • Tubules ○ Severe tubular injury, disruption, and destruction

Kidney Transplantation

TERMINOLOGY

4.

5.

6.

7.

Bukreyeva I et al: Enterocytozoon bieneusi microsporidiosis in stem cell transplant recipients treated with fumagillin1. Emerg Infect Dis. 23(6):10391041, 2017 Smith RM et al: Three cases of neurologic syndrome caused by donorderived microsporidiosis. Emerg Infect Dis. 23(3):387-395, 2017 Kicia M et al: Prevalence and molecular characteristics of urinary and intestinal microsporidia infections in renal transplant recipients. Clin Microbiol Infect. 22(5):462.e5-9, 2016 Hocevar SN et al: Microsporidiosis acquired through solid organ transplantation: a public health investigation. Ann Intern Med. 160(4):21320, 2014 Nagpal A et al: Disseminated microsporidiosis in a renal transplant recipient: case report and review of the literature. Transpl Infect Dis. 15(5):526-32, 2013 Lanternier F et al: Microsporidiosis in solid organ transplant recipients: two Enterocytozoon bieneusi cases and review. Transpl Infect Dis. 11(1):83-8, 2009 Orenstein JM: Diagnostic pathology of microsporidiosis. Ultrastruct Pathol. 27(3):141-9, 2003

223

Kidney Transplantation

Tuberculosis KEY FACTS

TERMINOLOGY

MACROSCOPIC

• Infection by Mycobacterium tuberculosis

• Destruction of pelvic calyces and papillae with caseous cheesy material • Large, tumor-like nodules of chalky material may replace renal parenchyma

ETIOLOGY/PATHOGENESIS • Reactivated latent infection or hematogenous dissemination of active pulmonary infection

CLINICAL ISSUES • Genitourinary TB accounts for 30% of extrapulmonary TB in developed countries • Diagnosis ○ Sterile pyuria ○ Mycobacterial cultures positive in 6-8 weeks ○ Mantoux skin test ○ Polymerase chain reaction detection of mycobacterial nucleic acid ○ Interferon-γ release assay (ELISA blood test) – M. tuberculosis antigens in latent and active infection stimulate production of host interferon-γ

MICROSCOPIC • Caseating granulomatous inflammation ○ Central necrosis rimmed by histiocytes, plasma cells, lymphocytes, and few multinucleated giant cells • AA amyloidosis with longstanding infection • Cases of glomerulonephritis reported in endemic areas

ANCILLARY TESTS • Acid-fast bacteria stain (Ziehl-Neelsen) ○ Rare organisms positive at periphery of necrosis

TOP DIFFERENTIAL DIAGNOSES • M. avium-intracellulare infection, sarcoidosis, BCG therapy interstitial nephritis, drug-induced interstitial nephritis

Caseous Necrosis in Renal Pelvis

Necrotizing Granulomatous Inflammation

Lymphohistiocytic Inflammation and Giant Cells

Acid-Fast Bacilli in Granulomas

(Left) Gross photograph of a nephrectomy specimen with renal tuberculosis shows a dilated pelvicalyceal system with ulcerated papillae and caseous necrotic material ﬊. (Courtesy L. Fajardo, MD.) (Right) Caseating granulomatous inflammation is seen in a kidney resected for renal tuberculosis. Extensive necrosis ﬊ is present in the granulomas with associated lymphohistiocytic infiltrate.

(Left) Necrotizing granulomas are seen in the pelvic wall, extending into the medulla of a kidney with tuberculous infection. In addition to the histiocytes, occasional multinucleated giant cells ﬊ are seen rimming the necrosis. (Right) Acid-fast bacteria stain of the kidney shows scattered acid-fast bacilli ﬈ in the granulomas. (Courtesy G. Berry, MD.)

224

Tuberculosis

Abbreviations

Prognosis • Treatment failure due to nonadherence to regimen or drug-resistant organisms

• Tuberculosis (TB)

Definitions • Infection of kidney by Mycobacterium tuberculosis

ETIOLOGY/PATHOGENESIS Infectious Agents • M. tuberculosis: Most common organism • Mycobacterium bovis: Bovine bacillus rarely causes disease • Mycobacterium avium-intracellulare in immunosuppressed state

Pathogenesis • Acute infection • Hematogenous dissemination of active pulmonary TB • Reactivated latent infection

CLINICAL ISSUES Epidemiology • Incidence ○ Genitourinary TB accounts for 30% of extrapulmonary TB in developed countries – Higher frequency in immunosuppressed individuals (HIV, transplantation, dialysis), endemic areas, and drug-resistant TB infection • Sex ○ More common in men with genital TB

Site • Pelvic calyces and renal medulla

Presentation • Lower urinary tract infection symptoms • Constitutional symptoms of fever, weight loss unusual

Laboratory Tests • Urine ○ White blood cells on microscopy ○ Urine bacterial cultures negative (sterile pyuria) • Mantoux skin test • Polymerase chain reaction detection of mycobacterial nucleic acid • Interferon-γ release assay (ELISA blood test; QuantiFERON) ○ M. tuberculosis antigens in latent and active infection stimulate production of host interferon-γ

Treatment • Options, risks, complications ○ Hypertension, chronic renal failure • Surgical approaches ○ Nephrectomy for cavitary lesions ○ Relief of obstruction for ureteral strictures • Drugs ○ Multidrug regimen with isoniazid, rifampicin, pyrazinamide, and ethambutol (or streptomycin) for months

IMAGING Radiographic Findings • Calyceal distortion, ureteric strictures on intravenous pyelography • Calcification of necrotic papillae

Kidney Transplantation

TERMINOLOGY

MACROSCOPIC General Features • Ulceration/destruction of pelvic calyces and papillae with caseous cheesy material • Large, tumor-like nodules of chalky material may replace renal parenchyma

MICROSCOPIC Histologic Features • Caseating granulomatous inflammation ○ Early infection in medulla but can affect entire kidney ○ Central necrosis rimmed by histiocytes, plasma cells, lymphocytes, and few multinucleated giant cells • Extensive tubular atrophy, interstitial fibrosis, and variable glomerulosclerosis • Severe interstitial inflammation may be present • AA amyloidosis with longstanding infection • Glomerulonephritis reported with systemic TB in endemic areas ○ IgA nephropathy: Most common

ANCILLARY TESTS Histochemistry • Acid-fast bacteria stain (Ziehl-Neelsen) ○ Reactivity: Positive ○ Staining pattern: Rare organisms at periphery of necrosis

DIFFERENTIAL DIAGNOSIS Mycobacterium avium-intracellulare Infection • Sheets of macrophages, lacks caseation

Sarcoidosis • Granulomas lack caseating necrosis

Bacille Calmette-Guérin Granulomatous Interstitial Nephritis • Temporal association to BCG therapy

Drug-Induced Interstitial Nephritis • Granulomas lack caseating necrosis

SELECTED REFERENCES 1. 2. 3. 4.

Verma AK et al: Renal tuberculosis presenting as acute pyelonephritis - a rarity. Indian J Tuberc. 63(3):210-213, 2016 Sourial MW et al: Genitourinary tuberculosis in North America: a rare clinical entity. Can Urol Assoc J. 9(7-8):E484-9, 2015 Sun L et al: Be alert to tuberculosis-mediated glomerulonephritis: a retrospective study. Eur J Clin Microbiol Infect Dis. 31(5):775-9, 2012 Eastwood JB et al: Tuberculosis and the kidney. J Am Soc Nephrol. 12(6):1307-14, 2001

225

Kidney Transplantation

Malakoplakia KEY FACTS

TERMINOLOGY • Chronic bacterial infection with abundant macrophages containing granular eosinophilic cytoplasm and MichaelisGutmann (MG) bodies often forming plaque or mass lesion

ETIOLOGY/PATHOGENESIS • Escherichia coli most common • Defective intracytoplasmic macrophage bactericidal function ○ Partially digested bacterial products form nidus for calcium and iron deposition • Altered immune status is predisposing factor

CLINICAL ISSUES • M:F = 1:4 • Involves renal pelvicalyceal system and parenchyma ○ Bilateral involvement common (30-50% of cases) • Presentation ○ Fever, chills

○ Flank pain, loin tenderness ○ Palpable mass when present; can be mistaken for neoplasm, prompting nephrectomy • Surgical therapy now less common with antibiotics and better imaging techniques

MACROSCOPIC • Yellow-tan nodules in calyces and renal parenchyma

MICROSCOPIC • Sheets of macrophages with foamy eosinophilic cytoplasm in mass lesion • Characteristic cytoplasmic inclusions (MG bodies) ○ Calcium positive (von Kossa stain) ○ Iron positive (Prussian blue stain)

TOP DIFFERENTIAL DIAGNOSES • Renal cell carcinoma • Xanthogranulomatous pyelonephritis • Megalocytic interstitial nephritis

Malakoplakia Mimicking Neoplasm

Sheets of Macrophages

Cytoplasmic Granules on PAS Stain

Michaelis-Gutmann Bodies

(Left) Gross photograph of a kidney with malakoplakia shows multiple tan-yellow masses ﬊ mimicking a neoplasm. (Courtesy R. Rouse, MD.) (Right) Hematoxylin and eosin of renal malakoplakia is shown. The mass identified in the nephrectomy specimen is composed of sheets of macrophages with granular eosinophilic cytoplasm ﬊.

(Left) A renal biopsy shows sheets of interstitial macrophages with PAS(+) cytoplasmic granules ﬇, characteristic of malakoplakia. Patchy areas of interstitial neutrophils and neutrophil casts were seen elsewhere in the biopsy (not shown). (Right) A renal biopsy with features of malakoplakia is shown. In addition to sheets of macrophages, the biopsy showed characteristic cytoplasmic inclusions referred to as Michaelis-Gutmann bodies ﬇.

226

Malakoplakia

MACROSCOPIC

Synonyms

General Features

• Malacoplakia

• • • •

Definitions • Chronic bacterial infection with abundant macrophages containing granular eosinophilic cytoplasm and MichaelisGutmann bodies often forming plaque or mass lesion

ETIOLOGY/PATHOGENESIS Infectious Agents • Escherichia coli most common

Defective Intracytoplasmic Macrophage Bactericidal Function • Decreased lysosomal degradation of bacteria • Inability of cells to release lysosomal enzymes • Partially digested bacterial products form nidus for calcium and iron deposition

Altered Immune Status Is Predisposing Factor • AIDS • Immunosuppressive therapy • Malignancies

CLINICAL ISSUES Epidemiology

Enlarged kidneys Yellow-tan nodules in calyces and parenchyma Dilated pelvicalyceal system if urinary obstruction Renal calculi rare

MICROSCOPIC Histologic Features • Sheets of macrophages with foamy eosinophilic cytoplasm in mass lesion • Characteristic cytoplasmic inclusions, (Michaelis-Gutmann) bodies ○ 4-10 μm in diameter, basophilic ○ Periodic acid-Schiff positive ○ Calcium positive (von Kossa stain) ○ Iron positive (Prussian blue stain) • Admixed lymphocytes and plasma cells

ANCILLARY TESTS Electron Microscopy • Michaelis-Gutmann bodies have central crystalline core, intermediate lucent area, and peripheral lamellar rings of deposited mineral

DIFFERENTIAL DIAGNOSIS

• Age ○ Infancy to 9th decade – 5th decade most common • Sex ○ M:F = 1:4

Renal Cell Carcinoma

Site

• Macrophages foamy and lipid laden • Absence of Michaelis-Gutmann bodies • Usually associated with "staghorn" calculus

• Urinary bladder: Most common • Renal pelvicalyceal system and parenchyma ○ Bilateral involvement common (30-50% of cases)

Presentation • • • •

Fever, chills Flank pain, loin tenderness Palpable mass when present, can be mistaken for neoplasm Acute renal failure in bilateral disease and renal transplants

Laboratory Tests • Urinalysis shows pyuria and proteinuria • Urine cultures may be positive for E. coli

Treatment • Surgical approaches ○ Nephrectomy if suspected malignancy ○ Surgical therapy now less common with antibiotics and better imaging techniques • Drugs ○ Fluoroquinolone

Prognosis • High mortality rate (70%) in early reports • Significant improvement in mortality rate with early diagnosis and antibiotic therapy with fluoroquinolone

Kidney Transplantation

TERMINOLOGY

• Immunohistochemistry with CKAE1/CAM5.2 (epithelial cells), CD163, and CD68 (macrophages) is helpful

Xanthogranulomatous Pyelonephritis

Megalocytic Interstitial Nephritis • Macrophages have strongly PAS-positive granular eosinophilic cytoplasm • Absence of Michaelis-Gutmann bodies

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • High level of suspicion needed for biopsy diagnosis • von Kossa stains and immunostains are useful

SELECTED REFERENCES 1. 2.

3. 4. 5. 6.

Nieto-Ríos JF et al: Malakoplakia after kidney transplantation: case report and literature review. Transpl Infect Dis. ePub, 2017 Das DP et al: Co-existing malakoplakia and xanthogranulomatous pyelonephritis of kidney: two different spectrum of same disease process. Urol Ann. 8(2):252-4, 2016 Lusco MA et al: AJKD atlas of renal pathology: malakoplakia. Am J Kidney Dis. 68(4):e27-8, 2016 Kobayashi A et al: Malakoplakia of the kidney. Am J Kidney Dis. 51(2):326-30, 2008 August C et al: Renal parenchymal malakoplakia: ultrastructural findings in different stages of morphogenesis. Ultrastruct Pathol. 18(5):483-91, 1994 al-Sulaiman MH et al: Renal parenchymal malacoplakia and megalocytic interstitial nephritis: clinical and histological features. Report of two cases and review of the literature. Am J Nephrol. 13(6):483-8, 1993

227

Kidney Transplantation

Malakoplakia

Macrophage Infiltrate Mimics Neoplasm

Basophilic Michaelis-Gutmann Bodies

Necrosis and Acute Inflammation in Malakoplakia

Acute Inflammation in Malakoplakia

Perinephric Tissue Involvement

Nephrectomy for Suspected Neoplasm

(Left) Renal malakoplakia, as shown here, is composed of sheets of plump macrophages ﬇ with granular cytoplasm mimicking a neoplasm. A few residual preserved renal tubules ﬉ are also observed. (Right) Hematoxylin and eosin of renal malakoplakia at high magnification shows the typical basophilic MichaelisGutmann bodies ﬉ within the cytoplasm of the macrophages.

(Left) Renal malakoplakia can show areas of necrosis and acute inflammation ﬊ along with the characteristic macrophages with eosinophilic cytoplasm ﬉. (Right) Scattered foci of necrosis and acute inflammation ﬊ can be seen in renal malakoplakia along with macrophages with granular cytoplasm. On the other hand, renal tuberculosis has caseating granulomas with central acellular necrosis surrounded by epithelioid histiocytes.

(Left) Sheets of histiocytes in malakoplakia extending into the pelvic adipose tissue ﬊ are shown. Similar changes in the capsular surface cause perinephric adhesions. The macroscopic findings of a nephrectomy closely resemble renal cell carcinoma. (Right) End-stage native kidney with multiple cysts ﬊ was removed for a suspected renal neoplasm. The microscopic examination of the mass was compatible with renal malakoplakia. The patient had undergone renal transplantation 2 years earlier.

228

Malakoplakia

Lack of Epithelial Cell Component (Left) CD163 (macrophage marker) of the renal mass lesion diagnosed as malakoplakia is shown. The macrophages are positive for CD163. The cytokeratin stain is negative, ruling out the possibility of renal cell carcinoma. (Right) AE1/AE3 immunohistochemistry of the native kidney mass diagnosed as renal malakoplakia in a renal transplant recipient is negative. The lesional cells have granular cytoplasm and lack staining for cytokeratin ﬊ but are positive for CD163 (macrophage marker, not shown).

Macrophages in Renal Cell Carcinoma

Kidney Transplantation

Immunohistochemical Confirmation of Macrophages

Granular Cytoplasm of Malakoplakia Cells (Left) CD163 stain of a renal cell carcinoma demonstrates the lack of staining in the lesional neoplastic cells ﬊, while the admixed histiocytes are positive. Renal malakoplakia is composed of sheets of macrophages, and the histological findings can be mistaken for renal cell carcinoma. (Right) Periodic acid-Schiff of renal malakoplakia highlights the granular cytoplasm of the macrophages ﬊ and also stains the characteristic Michaelis-Gutmann bodies ﬈.

Refractile Michaelis-Gutmann Bodies

Calcium Deposits in Michaelis-Gutmann Bodies (Left) Giemsa stain of renal malakoplakia fails to reveal organisms, but the MichaelisGutmann bodies ﬇ are highlighted by their refractile appearance. Other stains for microorganisms, such as acidfast bacilli, Gram, and Gomori methenamine-silver, were also negative. (Right) von Kossa stain highlights the MichaelisGutmann bodies ﬈ in malakoplakia. Partially digested bacterial products form the nidus for the calcium [von Kossa (+)] and iron deposition in these bodies.

229

Kidney Transplantation

Nocardiosis KEY FACTS

ETIOLOGY/PATHOGENESIS • Nocardia asteroides most common agent in temperate climates • Infection due to inhalation of organisms or direct inoculation into skin via trauma • Opportunistic infection in immunocompromised host • Can occur in even healthy individuals

CLINICAL ISSUES • Nocardia infection rare ○ Occurs in ≤ 2% of renal transplant recipients • Kidney involvement rare ○ Secondary to hematogenous spread • Presentation ○ Fever, chills ○ Pain and tenderness in loin or around renal allograft ○ Skin involvement with draining sinuses may be present • Laboratory tests ○ Aerobic bacterial cultures of blood, body fluids, or tissue

○ More sensitive molecular detection methods increasingly used • Sulfonamides are 1st-line treatment ○ If resistant, alternative agents, such as amikacin, imipenem, and 3rd generation cephalosporins may be used ○ Treatment for 6-12 months required, especially in immunocompromised patients • Central nervous system involvement ○ Poor prognostic sign

MICROSCOPIC • Multiple necrotizing microabscesses with neutrophils • Rare case of mesangiocapillary glomerulonephritis reported in patient with Nocardia pneumonia

ANCILLARY TESTS • Branched, thin filamentous bacteria seen on GrocottGomori methenamine silver stain and Gram stain

Necrotizing Microabscesses in Nocardia Infection

Nocardial Microabscesses With Multinucleated Giant Cells

Filamentous Nocardial Organisms on GMS Stain

Filamentous Nocardial Organisms on Gram Stain

(Left) Nocardia infection in the kidney is characterized by multiple necrotizing microabscesses ﬊ with associated renal parenchymal damage. (Right) Hematoxylin and eosin characteristically shows a mixed inflammatory infiltrate (neutrophils, lymphocytes, and plasma cells) with multinucleated giant cells ﬈, which should raise the consideration of nocardiosis. Well-formed granulomas (not shown) are not present. (Courtesy A. Husain, MD.)

(Left) GMS stain shows slender, branching filamentous organisms ﬊ characteristic of Nocardia infection within the microabscesses. (Courtesy A. Husain, MD.) (Right) Gram stain highlights a small collection of tangled filamentous Nocardia organisms ﬊ within the microabscesses.

230

Nocardiosis

IMAGING

Definitions

General Features

• Acute inflammation of renal parenchyma by Nocardia

• Renal parenchymal or perinephric abscesses

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

Infectious Agents

Histologic Features

• Nocardia asteroides most common agent in temperate climates • Nippostrongylus brasiliensis more common in tropical climates ○ Less common species include Nocardia caviae, Nocardia nova, and Nocardia farcinica • Ubiquitous organisms in soil ○ Infection route by inhalation or direct inoculation into skin via trauma or animal bite

• Multiple necrotizing microabscesses with neutrophils ○ Multinucleated giant cells may be seen • Acute inflammation may affect glomeruli along with adjacent tubulointerstitium • Blood vessels usually spared • Rare case of mesangiocapillary glomerulonephritis reported in patient with Nocardia pneumonia

Predisposing Factors

Histochemistry

• Opportunistic infection in immunocompromised host ○ Solid organ transplantation ○ Corticosteroid therapy, HIV infection • Can occur with chronic lung disease, diabetes, carcinoma, or even healthy individuals

• Grocott-Gomori methenamine silver ○ Reactivity: Positive ○ Staining pattern: Branched, thin filamentous bacteria • Gram ○ Reactivity: Positive ○ Staining pattern: Branched, thin filamentous bacteria • Periodic acid-Schiff and acid-fast stains ○ Filamentous bacteria less well seen

CLINICAL ISSUES Epidemiology • Incidence ○ Nocardia infection rare – Occurs in ≤ 2% of renal transplant recipients – Lungs most commonly affected organs, followed by skin and disseminated infection – Kidney involvement rare □ Secondary to hematogenous spread

Site • Kidney parenchyma and perinephric tissues

Presentation • • • •

Fever, chills Pulmonary symptoms with septicemia Pain and tenderness in loin or around renal allograft Skin involvement with draining sinuses may be present

Laboratory Tests • Aerobic bacterial cultures of blood, body fluids, or tissue • Urine cultures positive for Nocardia if pyelitis present • More sensitive molecular detection methods increasingly used

Treatment • Drugs ○ Sulfonamides are 1st-line treatment – If resistant, alternative agents, such as amikacin, imipenem, and 3rd generation cephalosporins used ○ Prolonged treatment for 6-12 months required, especially in immunocompromised patients

Prognosis • Central nervous system involvement ○ Poor prognostic sign

Kidney Transplantation

TERMINOLOGY

ANCILLARY TESTS

DIFFERENTIAL DIAGNOSIS Nonnocardial Bacterial Pyelonephritis • Histochemical stains and cultures useful

Pauci-Immune Glomerulonephritis and Vasculitis • Glomerular crescents identified and vasculocentric inflammation may be seen • Neutrophils seen but well-defined abscesses absent

Acute Tubulointerstitial Nephritis • Predominantly lymphocytic infiltrate • Neutrophils seen but well-defined abscesses absent

Infectious Granulomatous Pyelonephritis • Epithelioid histiocytes and multinucleated giant cells present at periphery of necrosis • Mycobacterial and fungal infections should be excluded

SELECTED REFERENCES 1. 2. 3. 4. 5. 6. 7. 8.

Coussement J et al: Nocardia infection in solid organ transplant recipients: a multicenter European case-control study. Clin Infect Dis. 63(3):338-45, 2016 Shrestha S et al: Different faces of Nocardia infection in renal transplant recipients. Nephrology (Carlton). 21(3):254-60, 2016 Santos M et al: Infection by Nocardia in solid organ transplantation: thirty years of experience. Transplant Proc. 43(6):2141-4, 2011 Belhocine W et al: Nocardia carnea infection in a kidney transplant recipient. Transplant Proc. 42(10):4359-60, 2010 Einollahi B et al: Invasive fungal infections following renal transplantation: a review of 2410 recipients. Ann Transplant. 13(4):55-8, 2008 D'Cruz S et al: Isolated nocardial subcapsular and perinephric abscess. Indian J Pathol Microbiol. 47(1):24-6, 2004 Jose MD et al: Mesangiocapillary glomerulonephritis in a patient with Nocardia pneumonia. Nephrol Dial Transplant. 13(10):2628-9, 1998 Raghavan R et al: Fungal and nocardial infections of the kidney. Histopathology. 11(1):9-20, 1987

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SECTION 5

Liver Transplantation

Pathologic Classification of Liver Allograft Diseases Gross Evaluation of Failed Native Liver Evaluation of Failed Liver Allograft Evaluation of the Donor Liver History of Liver Transplantation

234 236 240 242 246

Posttransplant Surgical Complications Preservation Injury Hepatic Artery Thrombosis Portal Vein Thrombosis Bile Duct Stricture, Leak, Sludge, Biloma Hepatic Venous Outflow Obstruction Hyperperfusion Syndrome

248 250 252 254 256 260

Allograft Rejection T-Cell-Mediated Rejection, Liver Antibody-Mediated Rejection, Liver Chronic (Ductopenic) Rejection

262 266 270

Recurrent Diseases in Liver Allograft Recurrent Hepatitis B Virus Recurrent Hepatitis C Virus Fibrosing Cholestatic HBV or HCV Hepatitis Recurrent Autoimmune Hepatitis Recurrent Primary Biliary Cholangitis Recurrent Primary Sclerosing Cholangitis Recurrent Fatty Liver Disease

272 274 278 280 282 284 286

Infections 288 290 292 294 296 300

Cytomegalovirus Herpes Simplex Virus Adenovirus, Liver Hepatitis E Virus Epstein-Barr Virus, Liver Fungal Infections, Liver

Late-Graft Dysfunction Plasma Cell-Rich Rejection Graft-vs.-Host Disease, Liver

302 304

Liver Transplantation

Pathologic Classification of Liver Allograft Diseases

Pathologic Classification • Based on pathogenesis, divided into broad categories of alloimmune, nonalloimmune, recurrent diseases, and infections

Abbreviations • • • • • • • • •

Hepatitis B virus (HBV) Hepatitis C virus (HCV) Cytomegalovirus (CMV) Epstein-Barr virus (EBV) Herpes simplex virus (HSV) Antibody-mediated rejection (AMR) Graft-vs.-host disease (GVHD) Primary biliary cirrhosis (PBC) Primary sclerosing cholangitis (PSC)

Definitions • Endotheliitis ○ Lymphocytes lifting or undermining endothelial cells

ALLOIMMUNE RESPONSE Cellular/T-Cell-Mediated Rejection • Acute rejection ○ Mixed portal inflammatory cell infiltrates ○ Bile duct damage/inflammation ○ Subendothelial venous inflammation (endotheliitis) – Portal vein or central vein

Chronic (Ductopenic) Rejection • Early ○ Lymphocytic cholangitis – Atypical duct epithelium (may mimic dysplasia) ○ Perivenular hepatocyte dropout • Late ○ Loss of interlobular bile ducts ○ Loss of hepatic arteries ○ Foam cell arteriopathy ○ Perivenular necrosis or dropout ○ Decreased portal inflammation • Ductopenic rejection ○ > 50% of portal tracts without bile ducts – > 20 portal tracts must be evaluated (may include multiple biopsies) ○ Cholestasis ○ Ductular proliferation ○ Perivenular fibrosis

Antibody-Mediated Rejection • Hyperacute rejection ○ Fibrin thrombi in portal and central veins ○ Neutrophilic &/or fibrinoid arteritis ○ Widespread or massive hemorrhagic necrosis ○ Sinusoidal congestion and fibrin deposition ○ Hemorrhage into portal tracts • Acute AMR ○ Portal and sinusoidal infiltration by neutrophils ○ Portal and periportal edema and ductular reaction ○ Portal and periportal hemorrhage 234

○ Periportal coagulative necrosis ○ Patchy hepatocyte ballooning and necrosis ○ Cholestasis

TERMINOLOGY

Graft-vs.-Host Disease • Bile duct epithelial damage ○ Damage to majority of ducts with minimal duct loss ○ Lymphocytic infiltration of bile ducts; minimal inflammation otherwise ○ Sloughing or necrosis of bile duct epithelium • Endotheliitis • No definitive features distinguish acute from chronic GVHD

Drug-Induced Vanishing Bile Duct Syndrome • History of medication use associated with ductopenia ○ Amoxicillin/clavulanate ○ Chlorpromazine ○ Phenytoin

NONALLOIMMUNE DISEASES Preservation Injury • • • •

Zone 3 injury Neutrophilic lobular infiltration Steatosis Biliary/cholestatic changes

Hepatic Venous Outflow Obstruction • Zone 3 changes ○ Sinusoidal dilatation/congestion ○ Perivenular and sinusoidal fibrosis ○ Central vein narrowing or obliteration

Hyperperfusion Syndrome • Early ○ Endothelial denudation of portal veins and periportal sinusoids ○ Sinusoidal dilatation and congestion ○ Cholestasis ○ Ischemic cholangitis • Late ○ Portal vein branch thrombosis ○ Biliary stricture ○ Nodular regenerative hyperplasia

Bile Duct Stricture, Leak, Sludge, Biloma • Features of large bile duct obstruction • Bile lakes and infarcts • Features of ascending cholangitis

Hepatic Artery Thrombosis • Bile duct necrosis • Necrosis of hepatocytes and portal connective tissue • Ongoing ischemia causes biliary strictures, fibrosis, and duct loss

Portal Vein Thrombosis • Hepatocyte swelling and necrosis • Hemorrhage • Thrombosis in branches of portal vein

Posttransplant Lymphoproliferative Disorder • EBV positive in majority

Pathologic Classification of Liver Allograft Diseases

RECURRENT DISEASES Autoimmune Hepatitis • • • •

Portal tract inflammation with numerous plasma cells Central vein inflammation with numerous plasma cells Acidophil bodies Parenchymal collapse/necrosis

Fatty Liver Disease • • • •

Steatosis Lobular inflammation Hepatocyte injury Perivenular sinusoidal fibrosis

Hepatitis B Virus • Early ○ Mild lobular hepatitis ○ Scattered acidophil bodies and Kupffer cells ○ Minimal portal inflammation • Late ○ Chronic hepatitis pattern ○ Portal inflammation ○ Interface/lobular activity ○ Progressive fibrosis ○ Occasional ground-glass hepatocytes • Fibrosing cholestatic hepatitis ○ Rapid progression to severe fibrosis and allograft failure ○ High viral load

Hepatitis C Virus • Early ○ Mild lobular disarray/inflammation ○ Kupffer cell aggregates ○ Scattered acidophil bodies • Late ○ Portal inflammation (± lymphoid aggregates) with interface and lobular activity ○ Focal bile duct lymphocytic infiltration • Fibrosing cholestatic hepatitis ○ Rapid progression to severe fibrosis and allograft failure ○ High viral load • Following viral eradication ○ Persistent necroinflammatory activity common ○ Progression of fibrosis seen in some cases

Primary Biliary Cirrhosis • • • •

Positive antimitochondrial antibodies Florid duct lesions Portal inflammation Bile ductular reaction

• Duct sclerosis • Periductal fibrosis • Portal inflammation with bile ductular reaction

INFECTIONS Adenovirus • Intranuclear viral inclusions with smudgy appearance • Necrotic areas without zonal distribution

Liver Transplantation

• Types ○ Early lesions – Preserved hepatic architecture ○ Polymorphic – Efface portal architecture ○ Monomorphic ○ Classic Hodgkin type

Cytomegalovirus • • • •

"Owl-eye" intranuclear inclusions Intracytoplasmic basophilic granules Lobular inflammation and disarray Scattered microabscesses

Herpes Simplex Virus • Cowdry types A and B intranuclear inclusions • Foci of hepatocyte necrosis • Formation of syncytia

Hepatitis E Virus • Acute ○ Lobular inflammation with acidophil bodies and lymphocytic infiltrate • Chronic ○ Portal lymphocyte-predominant inflammation and mild lobular inflammation ○ Minority develop cirrhosis

Epstein-Barr Virus • Portal mononuclear cell infiltrates • Sinusoidal infiltration with lymphocytes in linear, bead-like pattern • Mild lobular disarray with focal hepatocyte necrosis

Fungal Infections • Candidiasis ○ Most common fungal infection ○ Budding yeasts and pseudohyphae • Aspergillosis ○ 2nd most common fungal infection ○ Branching septate hyphae • Cryptococcosis ○ 3rd most common fungal infection ○ Mucicarmine-positive and PAS-positive capsule

SELECTED REFERENCES 1.

2. 3. 4.

Whitcomb E et al: Biopsy specimens from allograft liver contain histologic features of hepatitis C virus infection after virus eradication. Clin Gastroenterol Hepatol. 15(8):1279-1285, 2017 Naini B et al: Liver transplant pathology: review of challenging diagnostic situations. Surg Pathol Clin. 6(2):277-93, 2013 Te HS et al: Hepatitis E virus infection in a liver transplant recipient in the United States: a case report. Transplant Proc. 45(2):810-3, 2013 Banff Working Group et al: Liver biopsy interpretation for causes of late liver allograft dysfunction. Hepatology. 44(2):489-501, 2006

Primary Sclerosing Cholangitis • Characteristic endoscopic retrograde cholangiopancreatography findings 235

Liver Transplantation

Gross Evaluation of Failed Native Liver

TERMINOLOGY Definitions • Native liver: Transplant patient's original liver

CLINICAL IMPLICATIONS Common Indications for Transplantation • Acute liver failure ○ Fulminant viral etiology – Hepatitis A virus – Hepatitis B virus (HBV) – Hepatitis E virus – Autoimmune hepatitis ○ Drug toxicity (e.g., acetaminophen) ○ Toxin exposure (e.g., Amanita phalloides mushroom ingestion) ○ Acute fatty liver of pregnancy and HELLP syndrome ○ Wilson disease ○ Neonatal iron storage disease ○ Fatty acid oxidation defects ○ Unknown etiology (50% of cases) • Decompensated cirrhosis ○ Chronic viral hepatitis – HBV – Hepatitis D virus – Hepatitis C virus (HCV) ○ Autoimmune hepatitis ○ Chronic cholestatic disorders – Primary biliary cirrhosis – Primary sclerosing cholangitis – Biliary atresia – Alagille syndrome – Progressive familial intrahepatic cholestasis ○ Steatohepatitis – Alcoholic liver disease – Nonalcoholic steatohepatitis ○ Wilson disease ○ Genetic hemochromatosis ○ α-1-antitrypsin deficiency

○ Sarcoidosis • Hepatic tumor ○ Hepatocellular carcinoma ○ Hepatoblastoma ○ Metastatic neuroendocrine tumor ○ Epithelioid hemangioendothelioma ○ Biliary papillomatosis ○ Selected (rare) cases of cholangiocarcinoma • Metabolic disorder ○ Hereditary amyloidosis ○ Glycogen storage diseases ○ Niemann-Pick disease ○ Oxaluria ○ Galactosemia ○ Erythropoietic protoporphyria ○ Urea cycle defects • Vascular disorder ○ Budd-Chiari syndrome ○ Venoocclusive disease ○ Hepatoportal sclerosis ○ Nodular regenerative hyperplasia ○ Ischemic cholangiopathy • Hepatic involvement by fibropolycystic disease

MACROSCOPIC General Approach to Native Liver • Observe safety precautions ○ Treat every liver as if infected by HCV • Photograph anterior and posterior aspects of intact organ • Record organ weight and overall dimensions • Describe outer appearance ○ Presence of cirrhosis ○ Wrinkled capsule ○ Visible lesions ○ Patency of hepatic veins • Dissect hilar structures ○ Dissect gallbladder free or remove in continuity with hilar soft tissue

Normal-Appearing Liver With Glycogen Storage Disease (Left) This native liver of a patient with glycogen storage disease type III is grossly normal appearing, which is typical of many metabolic diseases. (Right) Although this section of the native liver appears grossly normal, the organ weighed only 900 g, and sections revealed diffuse recent centrilobular hepatocyte necrosis of unknown etiology.

236

Centrilobular Necrosis

Gross Evaluation of Failed Native Liver

Native Liver Removed for Decompensated Cirrhosis • Remove TIPS stent carefully, if any • Identify any small, incidental hepatocellular carcinomas ○ Cut entire liver into thin (0.5 cm) sections – Separate completely from each other • Identify by location, measure, and sample any lesion that differs from surrounding cirrhotic regenerative nodules (by size, color, texture, etc.) ○ Dysplastic nodules often bulge from cut surface of liver

Native Liver Removed for Acute Hepatic Failure • If no established etiology ○ Snap freeze liver for genetic and enzymatic studies ○ Place in glutaraldehyde for electron microscopic examination • Gross dictation should include ○ Description of degree and distribution of parenchyma loss ○ Degree and extent of nodular regeneration (if any) • Obtain routine sections from both lobes

Native Liver Removed for Tumor • Review medical record to identify previously ablated hepatic lesions ○ Estimate degree of gross tumor necrosis for all hepatocellular carcinomas ablated prior to transplantation ○ Sample ablated lesions generously to document any residual viable tumor • Proper TNM staging requires that each hepatocellular carcinoma be accurately measured and generously sampled to identify vascular invasion ○ Record size and location of each lesion ○ Estimate gross percentage of tumor necrosis for each lesion ○ Each lesion should be entirely submitted or sampled generously • Sample hilar structures, which represent margins

• Submit any hilar lymph nodes • Inspect hepatic veins and sample if grossly involved by tumor • Capsular margins not usually clinically relevant

Native Liver Removed for Metabolic Disease • If no established etiology ○ Snap freeze liver for genetic and enzymatic studies ○ Place in glutaraldehyde for electron microscopic examination • Obtain routine sections from both lobes

Liver Transplantation

○ Hilar structures (extrahepatic bile ducts, portal vein, hepatic artery) removed by dissecting hilar soft tissues en bloc in most cases – In selected cases, individually dissect hepatic artery, extrahepatic bile ducts, &/or portal vein branches ○ Submit sections that include portal vein, hepatic artery, and extrahepatic bile ducts ○ Submit representative section of gallbladder, noting presence of gallstones • Thin (0.5 cm) sections of entire liver (coronal or sagittal) ○ Note location and size of focal lesions and sample ○ Bile duct adenomas – Generally whitish tan, well circumscribed, and firm ○ Cavernous hemangiomas – Reddish and spongy – Often contain whitish tan, firm areas of sclerosis ○ Small, calcified nodules usually represent burned-out granulomas ○ Obtain tissue for electron microscopy in selected cases ○ Freeze tissue for diagnostic studies in selected cases ○ Freeze tissue for research per institutional protocol ○ Submit several sections from both liver lobes, generally avoiding hepatic capsule

Native Liver Removed for Vascular Disorder • Describe degree of gross parenchyma necrosis &/or dropout • Carefully dissect and sample ○ Hepatic artery ○ Portal vein ○ Hepatic veins

Native Liver Removed for Fibropolycystic Disease • Describe gross appearance of included portion of extrahepatic biliary tree • Assess appearance of parenchyma ○ Submit representative samples from various parts of liver • Describe and sample any cystic lesions within liver • Correlate with renal pathology ○ Helpful to establish precise diagnosis

Liver Removed for Specific Disease States • Primary sclerosing cholangitis ○ Critical to identify any occult hilar cholangiocarcinoma – Submit all hilar soft tissue for histologic examination ○ In rare cases of known hilar cholangiocarcinoma – Submit all hilar soft tissue for evaluation of response to neoadjuvant therapy • Acute fatty liver of pregnancy and HELLP syndrome ○ Prepare frozen sections for oil red O or Sudan black stains

SELECTED REFERENCES 1. 2. 3.

4.

Kohl CA et al: A new liver explant fixation technique. Arch Pathol Lab Med. 132(12):1859-60, 2008 Ghali P et al: Liver transplantation for incidental cholangiocarcinoma: analysis of the Canadian experience. Liver Transpl. 11(11):1412-6, 2005 Kirimlioglu H et al: Hepatocellular carcinomas in native livers from patients treated with orthotopic liver transplantation: biologic and therapeutic implications. Hepatology. 34(3):502-10, 2001 Ludwig J et al: The preparation of native livers for morphological studies. Mod Pathol. 7(7):790-3, 1994

237

Liver Transplantation

Gross Evaluation of Failed Native Liver Massive Necrosis Due to Herbal Supplement

Budd-Chiari Syndrome

Cirrhosis Secondary to Steatohepatitis

Primary Sclerosing Cholangitis

Occult Cholangiocarcinoma in Primary Sclerosing Cholangitis

Incidental Hepatocellular Carcinoma in Chronic HCV Hepatitis

(Left) Fulminant massive hepatic necrosis caused by a herbal supplement is shown. In the right half of this liver section, there is complete panlobular parenchymal extinction with lobular collapse. Some viable hepatic parenchyma remains on the left. (Right) Note the extensive centrilobular necrosis and large thrombi ſt in this liver from a patient with BuddChiari syndrome.

(Left) Gross specimen demonstrates micronodular cirrhosis due to alcoholic steatohepatitis. Sections revealed no steatosis since the patient had been abstinent for 6 months prior to transplantation. (Right) These liver sections are from a patient with biliary cirrhosis due to primary sclerosing cholangitis and are characterized by green discoloration and a fine pattern of cirrhosis.

(Left) Biliary cirrhosis due to primary sclerosing cholangitis is shown. Sections from the thickened wall of a cholangiectatic intrahepatic bile duct ſt revealed a 0.5-cm occult cholangiocarcinoma. (Right) Macronodular cirrhosis due to chronic hepatitis C virus (HCV) with an incidental 1.9cm hepatocellular carcinoma ſt is shown. Thin sectioning of cirrhotic native livers can reveal occult hepatocellular carcinomas, dysplastic nodules, and macroregenerative nodules.

238

Gross Evaluation of Failed Native Liver

Siderotic Dysplastic Nodule (Left) Macronodular cirrhosis due to chronic HCV hepatitis with an occult 1-cm dysplastic nodule ſt is shown. An ablated 4-cm hepatocellular carcinoma was present in another part of the liver (not shown). (Right) Occult siderotic dysplastic nodule (0.8 cm) ſt is shown in a background of macronodular cirrhosis due to chronic HCV hepatitis. This lesion is not clinically significant but needs to be submitted in its entirety to exclude hepatocellular carcinoma.

Ablated Hepatocellular Carcinoma

Liver Transplantation

Dysplastic Nodule

Partially Ablated Hepatocellular Carcinoma (Left) Completely ablated hepatocellular carcinoma ſt is shown in a background of macronodular cirrhosis due to chronic HCV. The entire lesion should be submitted for histologic evaluation to determine whether any viable tumor remains. (Right) Partially ablated hepatocellular carcinoma ſt is shown in a background of micronodular cirrhosis due to nonalcoholic steatohepatitis. In this case, the main lesion is entirely necrotic, but there are multiple satellite nodules ﬇ of viable tumor.

Dysplastic Nodule

Partially Ablated Hepatocellular Carcinoma (Left) Gross photograph shows a dysplastic nodule ſt within a liver that contained numerous dysplastic nodules. (Right) Partially ablated hepatocellular carcinoma is shown arising in a background of macronodular cirrhosis due to chronic HCV hepatitis. The lesion measures 3.5 cm in the greatest dimension, but most of the "lesional" tissue represents adjacent cirrhotic parenchyma that is necrotic. The actual residual tumor ſt is only 1.4 cm in the greatest dimension.

239

Liver Transplantation

Evaluation of Failed Liver Allograft

CLINICAL IMPLICATIONS Common Causes of Allograft Failure Leading to Retransplantation • Primary nonfunction • Technical surgical complications ○ Bile duct injury – Necrosis – Dehiscence – Stricture ○ Hepatic artery thrombosis ○ Portal vein thrombosis ○ Hepatic venous outflow obstruction • Antibody-mediated (humoral) rejection • Chronic (ductopenic) allograft rejection • Recurrence of primary liver disease ○ Chronic hepatitis C virus ○ Chronic hepatitis B virus ○ Autoimmune hepatitis ○ Primary biliary cirrhosis ○ Primary sclerosing cholangitis ○ Steatohepatitis – Alcoholic – Nonalcoholic

MACROSCOPIC Specimen Handling

Clinical Presentation • Early graft failure ○ Allograft failure in immediate posttransplant period suggests following diagnoses – Primary nonfunction – Hepatic artery thrombosis – Portal vein thrombosis – Antibody-mediated rejection ○ Primary nonfunction associated with – Older donor age – Presence of severe donor graft macrovesicular steatosis – Prolonged cold or warm ischemia time ○ Hepatic artery thrombosis

Failed Liver Allograft (Left) This liver allograft was removed 4 days after transplantation. The time-zero donor biopsy revealed 60% macrovesicular steatosis. At the time of retransplantation, all vascular anastomoses were intact. (Right) This section of a failed allograft with the diagnosis of primary nonfunction reveals extensive geographic areas of recent necrosis, involving ~ 75% of the parenchyma.

240

– Most common in pediatric transplant and cases with aberrant donor arterial anatomy ○ Portal vein thrombosis – Rare complication often in pediatric patients receiving split donor allografts or requiring portal vein reconstruction ○ Antibody-mediated rejection – Rare complication often in small percentage of ABOincompatible allografts • Late graft failure ○ Failure months to years after transplantation – Most often result of ischemic biliary complications or disease recurrence leading to cirrhosis ○ Allograft failure due to chronic (ductopenic) rejection was important issue in cyclosporine era – Incidence steadily decreased and now rare – Occurs primarily with insufficient immunosuppression ○ Disease recurrence is leading cause of allograft failure ○ Ischemic cholangiopathy remains important cause of late allograft dysfunction and loss – Due primarily to variety of vascular complications – Also due to use of ABO-incompatible donor organs

• Early allograft failure ○ Extent and distribution of parenchymal necrosis should be described in detail ○ Careful dissection of hilar structures to identify and sample hepatic artery and portal vein – Bile duct is of key importance to diagnose vascular obstruction • Late allograft failure ○ Careful dissection of hilar structures to identify and sample – Hepatic artery to diagnose foam cell arteriopathy or thrombosis – Portal vein to diagnose portal vein thrombosis

Primary Nonfunction

Evaluation of Failed Liver Allograft Late Graft Failure • Chronic (ductopenic) rejection ○ Extensively sample extrahepatic hepatic artery and large intrahepatic branches to search for foam cell arteriopathy ○ Describe degree of intrahepatic duct loss and fibrosis • Ischemic cholangiopathy ○ May have necrosis or strictures involving extrahepatic biliary tree or biloma due to anastomotic dehiscence ○ In some cases, examination of portal vein and hepatic artery may reveal organizing thrombus • Cirrhosis due to disease recurrence ○ Histologic features generally mirror those evident in de novo native liver setting

General Features • Sections to be submitted ○ Hepatic artery ○ Portal vein ○ Bile duct ○ Necrotic and nonnecrotic parenchyma from both lobes to identify early allograft failure ○ Routine samples from right and left lobes of cirrhotic liver in cases of – Disease recurrence – Late allograft failure

MICROSCOPIC

SELECTED REFERENCES

Early Graft Failure

1.

• In primary nonfunction, geographic areas of parenchymal necrosis vary in extent ○ Intrahepatic vascular thrombi may be present • Hepatic artery thrombosis ○ Difficult to identify in some cases ○ Usually extensive geographic parenchymal necrosis ○ Correlate with imaging studies – Particularly after thrombolytic therapy ○ Careful microscopic examination of hepatic artery may reveal dissection or intimal flap – Responsible for development of thrombosis in some cases • Portal vein thrombosis usually easily apparent ○ May be associated with intimal proliferation of donor portion of vein • Antibody-mediated rejection may exhibit only geographic areas of parenchymal necrosis ○ Immunofluorescent or immunohistologic demonstration of C4d deposition in vascular structures may aid in diagnosis – C4d may be absent with intensive plasmapheresis therapy ○ Rare reports of arterial vasculitis

Portal Vein Thrombosis

Liver Transplantation

– Extrahepatic bile ducts to diagnose ischemic cholangiopathy

2. 3.

4. 5. 6.

7. 8.

9.

Ackermann O et al: The long-term outcome of hepatic artery thrombosis after liver transplantation in children: role of urgent revascularization. Am J Transplant. 12(6):1496-503, 2012 Zoepf T et al: Optimized endoscopic treatment of ischemic-type biliary lesions after liver transplantation. Gastrointest Endosc. 76(3):556-63, 2012 Demetris A et al: Update of the International Banff Schema for Liver Allograft Rejection: working recommendations for the histopathologic staging and reporting of chronic rejection. An International Panel. Hepatology. 31(3):792-9, 2000 Bäckman L et al: Causes of late graft loss after liver transplantation. Transplantation. 55(5):1078-82, 1993 Hubscher SG et al: Primary biliary cirrhosis. histological evidence of disease recurrence after liver transplantation. J Hepatol. 18(2):173-84, 1993 Colonna JO 2nd et al: Biliary strictures complicating liver transplantation. Incidence, pathogenesis, management, and outcome. Ann Surg. 216(3):34450; discussion 350-2, 1992 Ludwig J et al: Ischemic cholangitis in hepatic allografts. Mayo Clin Proc. 67(6):519-26, 1992 Quiroga J et al: Cause and timing of first allograft failure in orthotopic liver transplantation: a study of 177 consecutive patients. Hepatology. 14(6):1054-62, 1991 Ludwig J et al: The acute vanishing bile duct syndrome (acute irreversible rejection) after orthotopic liver transplantation. Hepatology. 7(3):476-83, 1987

Ischemic Cholangiopathy (Left) CT scan performed 4 days post transplant demonstrates regions of parenchymal necrosis within the allograft. A thrombus is also present in the portal vein ﬈. (Right) Section of an allograft that was removed 6 weeks after transplantation due to hepatic artery thrombosis demonstrates extensive bile duct necrosis (ischemic cholangiopathy).

241

Liver Transplantation

Evaluation of the Donor Liver Performance of Donor Biopsy

TERMINOLOGY Definitions • Donor biopsy ○ Biopsy evaluated by frozen section prior to implantation of donor organ ○ Utilized primarily on deceased donor organs – Occasionally performed for living-donor allografts • "Time-zero" donor biopsy ○ Obtained at time of vascular anastomoses between donor organ and recipient (when blood flow to allograft is started) ○ Evaluate by permanent section after transplant operation completed • Extended donor criteria ○ Donor organs from patients with following characteristics – Age > 65 years – After cardiac death – Serum HCV RNA(+) – Serum HBV core antibody (+) – Human T-cell lymphotrophic virus (+) – Hypernatremia – Prior history of malignancy – Extended intensive care unit stay – Extended cold ischemia time ○ Utilized for patients unlikely to qualify for usual donor organs or in emergent need of transplant (i.e., fulminant hepatic failure)

CLINICAL IMPLICATIONS Utility of Donor Biopsy • Assess likelihood of adequate allograft function in immediate posttransplant period • Document baseline allograft disease (e.g., degree of fibrosis) • Evaluate degree of preservation injury • Diagnose mass lesions

• Not uniformly performed at most transplant centers • Donor biopsies often omitted when no donor condition exists and when surgeon believes organ is in good condition by visual inspection

MICROSCOPIC Evaluation of Donor Organ on Frozen Section • Sample adequacy ○ Submit biopsies in RPMI or on Telfa pad moistened with preservation fluid – Submerging specimen in saline causes artifactual hepatocyte swelling and damage that hinders histologic evaluation ○ Either needle or wedge biopsies can be used – Easier to obtain high-quality section from wedge biopsy, but shallow wedge biopsies do not always reflect true organ donor condition • Macrovesicular steatosis ○ Express as overall percentage of hepatocytes that contain large fat droplets ○ Evaluation performed as gestalt overall assessment, not on "worst" portion of biopsy ○ While percentage of macrovesicular steatosis may impact decision to utilize donor organ, there is no uniformly accepted cutoff for declining organ for transplantation – Severely steatotic livers (> 40-50%) □ May function more poorly in immediate posttransplant period □ May have slight increase in rate of primary nonfunction □ Have been utilized successfully in many patients who require emergent transplantation ○ Ballooned &/or swollen hepatocytes are not part of assessment of steatosis ○ Appearance of uniform, medium-sized clear cytoplasmic vacuoles in hepatocytes represents ice crystal artifact – Can be mistaken for steatosis

Frozen Section Artifact (Left) This frozen section exhibits significant sectioning artifact, which should not be confused with hepatocyte necrosis. (Right) This permanent section reveals normal liver, corresponding with the frozen section that had areas mimicking hepatocyte necrosis.

242

Corresponding Permanent Section

Evaluation of the Donor Liver • Isolated noncaseating granuloma • Hyalinized nodules/granulomas • Minimal spotty neutrophilic inflammation (i.e., surgical hepatitis) • Kupffer cell iron deposition • Genetic hemochromatosis without fibrosis • α-1-antitrypsin deficiency without fibrosis • Mild centrilobular sinusoidal fibrosis without active steatohepatitis • Any degree of lipofuscin pigment deposition

Donor Frozen Section Findings That Exclude Organ Use

8.

• • • •

Malignancy Cirrhosis Hepatic abscess Hepatocellular adenoma ○ No actual data regarding use of such organs • Fibropolycystic disease involving liver • Active parasitic or fungal infection • Significant steatohepatitis

Donor Frozen Section Findings Typically Without Clinical Significance • • • • • •

Minimal unexplained portal &/or lobular inflammation Rare scattered acidophil bodies Mild macrovesicular steatosis Bile duct adenoma and bile duct hamartoma Focal nodular hyperplasia Cavernous hemangioma

Liver Transplantation

– Artifact can be avoided by carefully blotting biopsy dry before freezing ○ Biopsies submerged in saline will exhibit artifactual diffuse hepatocyte swelling that can make it hard to evaluate for steatosis ○ Microvesicular steatosis is almost impossible to recognize reproducibly in H&E frozen sections – Presence has no bearing on posttransplant allograft function – Should not be routinely assessed or reported • Hepatocyte necrosis ○ Very recent centrilobular necrosis due to premortem or preharvesting donor hypoperfusion can be difficult to assess in frozen sections ○ Affected hepatocytes appear slightly shrunken with slightly condensed-appearing cytoplasm and should be present in distinct centrilobular distribution ○ Livers from donors who received maximal pressor support &/or were hypernatremic prior to death may exhibit scattered random foci of individual hepatocyte necrosis ○ No uniformly accepted cutoff for degree of hepatocyte necrosis that will lead to organ being declined for transplantation – Organs with > 10% hepatocyte necrosis not usually utilized • Fibrosis ○ Degree of portal fibrosis may be overestimated in shallow wedge biopsies ○ Unexplained mild portal fibrosis or even periportal fibrosis generally does not exclude organ from being utilized, but organs with bridging fibrosis are unlikely to be used • Inflammation ○ Minimal or mild unexplained portal and lobular inflammation can be ignored ○ Florid sinusoidal neutrophil infiltration may signify septicemia and should be communicated to donor team ○ Presence of abscess is contraindication for use of organ ○ Scattered clusters of lobular neutrophils may represent so-called "surgical hepatitis" and can be ignored

Evaluation of Serum HCV RNA or HBcAg(+) Organ Donors • Grading and staging per usual criteria • Evaluation of degree of steatosis • Organs with stage 3 or 4 fibrosis generally not utilized

Evaluation of "Time-Zero" Donor Biopsies • Degree of preservation injury ○ Related to length of cold and warm ischemia time ○ Can be seen even in living related-donor allografts ○ Centrilobular hepatocyte ballooning and cholestasis usually only features present at this point • Presence of significant hepatocyte necrosis is generally indicative of premortem or procurement liver injury • Evaluation of preexisting chronic donor liver disease

SELECTED REFERENCES 1.

2.

3. 4.

5. 6. 7.

9.

10.

11. 12.

13. 14. 15.

16. 17.

Fiorentino M et al: Predictive value of frozen-section analysis in the histological assessment of steatosis before liver transplantation. Liver Transpl. 15(12):1821-5, 2009 Koçbiyik A et al: Role of postreperfusion subcapsular wedge biopsies in predicting initially poor graft function after liver transplantation. Transplant Proc. 41(7):2747-8, 2009 Lo IJ et al: Utility of liver allograft biopsy obtained at procurement. Liver Transpl. 14(5):639-46, 2008 Nikeghbalian S et al: Does donor's fatty liver change impact on early mortality and outcome of liver transplantation. Transplant Proc. 39(4):11813, 2007 Alkofer B et al: Extended-donor criteria liver allografts. Semin Liver Dis. 26(3):221-33, 2006 Deshpande R et al: Can non-heart-beating donors replace cadaveric heartbeating liver donors? J Hepatol. 45(4):499-503, 2006 Feng S et al: Characteristics associated with liver graft failure: the concept of a donor risk index. Am J Transplant. 6(4):783-90, 2006 Nocito A et al: When is steatosis too much for transplantation? J Hepatol. 45(4):494-9, 2006 Perez-Daga JA et al: Influence of degree of hepatic steatosis on graft function and postoperative complications of liver transplantation. Transplant Proc. 38(8):2468-70, 2006 Renz JF et al: Utilization of extended donor criteria liver allografts maximizes donor use and patient access to liver transplantation. Ann Surg. 242(4):55663; discussion 563-5, 2005 Busuttil RW et al: The utility of marginal donors in liver transplantation. Liver Transpl. 9(7):651-63, 2003 Zamboni F et al: Effect of macrovescicular steatosis and other donor and recipient characteristics on the outcome of liver transplantation. Clin Transplant. 15(1):53-7, 2001 Crowley H et al: Steatosis in donor and transplant liver biopsies. Hum Pathol. 31(10):1209-13, 2000 Markin RS et al: Frozen section evaluation of donor livers before transplantation. Transplantation. 56(6):1403-9, 1993 D'Alessandro AM et al: The predictive value of donor liver biopsies for the development of primary nonfunction after orthotopic liver transplantation. Transplantation. 51(1):157-63, 1991 Kakizoe S et al: Frozen section of liver biopsy for the evaluation of liver allografts. Transplant Proc. 22(2):416-7, 1990 Todo S et al: Primary nonfunction of hepatic allografts with preexisting fatty infiltration. Transplantation. 47(5):903-5, 1989

243

Liver Transplantation

Evaluation of the Donor Liver

Ice Crystal Artifact

Frozen Section and Ice Crystal Artifact

Macrovesicular Steatosis

Corresponding Permanent Section

Steatosis and Ice Crystal Artifact

Corresponding Permanent Section

(Left) Ice crystal artifact produces uniform, small bubbles within each hepatocyte. This can be minimized by blotting the tissue dry before freezing. (Right) A combination of sectioning and ice crystal artifact makes this donor frozen section difficult to evaluate. The permanent section revealed 5% macrovesicular steatosis.

(Left) This frozen section was read as 20% macrovesicular steatosis. Microvesicular steatosis is also present but can be ignored. (Right) The corresponding permanent section of the biopsy revealed 10% macrovesicular steatosis. Posttransplant allograft function was excellent.

(Left) This donor wedge biopsy frozen section exhibits a combination of macrovesicular steatosis, microvesicular steatosis, and ice crystal artifact, which makes evaluation difficult. It was diagnosed as 30% macrovesicular steatosis. (Right) The corresponding permanent section of the biopsy reveals 20% macrovesicular steatosis. Posttransplant allograft function was excellent.

244

Evaluation of the Donor Liver

Osmotic Damage (Left) This donor biopsy frozen section was read as 40-50% macrovesicular steatosis. The organ was utilized for transplantation, and posttransplant allograft function was excellent. (Right) This donor wedge biopsy was received partially submerged in saline. The frozen section reveals osmotic damage to the hepatocytes in the left 1/2 of the field ﬈, which precludes proper histologic evaluation. The portion of the biopsy ﬊ that was not in contact with saline is normal.

Recent Hepatocyte Necrosis

Liver Transplantation

Macrovesicular Steatosis

Mild Mononuclear Portal Infiltrate (Left) This donor biopsy frozen section revealed scattered foci of recent hepatocyte necrosis ﬈. The donor had received maximal doses of pressor agents prior to brain death. This organ was utilized for transplantation. Serum aminotransferase levels were high in the immediate posttransplant period, but ultimately, the allograft functioned well. (Right) Mild mononuclear cell infiltrates involving a few portal tracts are a nonspecific finding that do not preclude use of the organ.

Fibrinoid Necrosis

Sinusoidal Neutrophils (Left) Apparent fibrinoid necrosis of a portal hepatic arteriole ſt is evident in this donor biopsy. The donor had experienced uncontrolled severe hypertension prior to brain death. This organ was utilized for transplantation, and postoperative allograft function was good. (Right) A few neutrophils ﬈ scattered within the sinusoids is not a contraindication using the donor organ for transplantation.

245

Liver Transplantation

History of Liver Transplantation

TERMINOLOGY Definitions • Auxiliary liver transplantation ○ Supplemental transplantation of allograft without removal of native liver ○ Historical importance • Orthotopic liver transplantation ○ Removal of native liver and replacement with allograft in original location of liver

CHRONOLOGY AND EVOLUTION Timeline

Modern Era

• Spans nearly 60 years ○ Advances in kidney transplantation instrumental to advancing liver transplantation

• 1989 ○ Dr. Russell Strong – 1st successful living-donor liver transplantation □ Utilized left liver lobe – 17-month-old boy received liver from 29-year-old mother – Performed in Brisbane, Australia • 1989 ○ Dr. Christoph Broelsch – 1st successful living-donor liver transplantation in United States □ Utilized left liver lobe – 2-year-old girl received liver from mother – Performed at University of Chicago Medical Center • 1994-1999 ○ Living-donor liver transplantation – Utilizing right liver lobe • 2010 ○ 6,291 liver transplantation procedures in United States • 2015 ○ Nonalcoholic steatohepatitis surpasses hepatitis C as leading indication for liver transplantation

Research Era • 1955 ○ Dr. C. Stuart Welch – 1st canine auxiliary liver transplantation □ Performed in Albany, NY • 1958-1959 ○ Dr. Thomas E. Starzl – 1st canine orthotopic liver transplantation □ Performed as multivisceral transplantation (liver, intestines) – Rejection ensued, resulting in death 5-10 days after surgery □ Radiation therapy did not improve outcomes

Early Clinical Era • 1963-1964 ○ 1st attempts with human liver transplantation – Dr. Thomas E. Starzl □ 5 attempts at University of Colorado □ Maximum survival of 23 days – Dr. Francis D. Moore □ 1 attempt (Peter Bent Brigham Hospital, Boston, MA) – Dr. J. Demirleau □ 1 attempt in Paris, France • 1964-1967 ○ Moratorium on human liver transplantation – Self-imposed by transplant community due to poor clinical outcomes and challenges in optimizing surgical techniques • 1967 ○ Dr. Carl Groth – 1st human liver transplants with 1-year survival – Aided by use of triple immunosuppressive drug regimen □ Azathioprine □ Prednisone □ Antilymphocyte globulin • 1969 ○ Dr. Thomas Starzl – 1st inborn error of metabolism (Wilson disease) cured by liver transplantation • 1969-1973 246

○ Dr. Thomas Starzl – Unsuccessful xenotransplantation of chimpanzee liver to human recipients • 1979 ○ Dr. Roy Calne – Use of cyclosporine-based immunosuppressive regimen – Dramatic improvement in patient survival • 1983 ○ Liver transplantation considered viable therapeutic option, no longer an experimental procedure ○ Transplant hepatology emerging as subspecialty

ORGAN ALLOCATION POLICY Definition of Death • Prior to 1968 ○ Cessation of heartbeat • 1968 to present ○ Brain death

Child-Pugh Score • Used to assess prognosis of chronic liver disease, particularly cirrhosis • Created by Drs. C. G. Child and J. G. Turcotte in 1964 (University of Michigan) ○ Modified by Dr. R. Pugh in 1972 • Sum of 5 criteria (each criterion scored from 1-3 for 15 total points) ○ Serum bilirubin ○ Serum albumin ○ International normalized ratio (INR) for prothrombin time ○ Ascites ○ Hepatic encephalopathy • Categorized into classes A, B, and C ○ Class A = 5-6 points

History of Liver Transplantation

Year

Landmark Event

Pioneers

1955

1st canine auxiliary liver transplantation

C. Stuart Welch

1958-1959

1st canine liver transplantation, part of multivisceral transplantation

Thomas Starzl

1963-1964

1st human liver transplantations

Thomas Starzl, Francis Moore, J. Demirleau

1967

Azathioprine-based immunosuppression regimen

Carl Groth

1969

1st inborn error of metabolism cured by liver transplantation

Thomas Starzl

1979

Cyclosporine-based immunosuppression regimen

Roy Calne

1989

1st living-donor liver transplantation

Russell Strong

1995

Banff classification for liver allograft pathology created

2010

Over 6,000 liver transplant procedures performed in United States

2015

Nonalcoholic steatohepatitis surpasses hepatitis C as leading indication for liver transplantation

– 2-year survival: 85% ○ Class B = 7-9 points – 2-year survival: 57% ○ Class C = 10-15 points – 2-year survival: 35%

Model for End-Stage Liver Disease Score • Introduced in 2002 ○ Used by United Network for Organ Sharing for prioritization of organ allocation • Utilizes following laboratory data ○ Serum bilirubin ○ Serum creatinine ○ INR (based on prothrombin time) • Model for end-stage liver disease (MELD) score = (9.57 x ln[serum creatinine (mg/dL)] + 3.78 x ln[serum bilirubin (mg/dL)] + 11.2 x ln(INR) + 6.43) ○ If patient requires hemodialysis, value for creatinine is automatically set to 4.0 • 3-month mortality rate for MELD scores ○ > 40: ~ 70% ○ 30-39: 53% ○ 20-29: 20% ○ 10-19: 6% ○ < 10: 2%

Pediatric End-Stage Liver Disease Score • Scoring system for children < 12 years old • Uses following laboratory data ○ Serum bilirubin ○ Serum albumin ○ INR • Pediatric end-stage liver disease score = 4.8 x ln[serum bilirubin (mg/dL)] + 18.57 x ln[INR] - 6.87 x ln[albumin (g/dL)] + 4.36 (< 1 year old) + 6.67 (if patient has growth failure, i.e., > 2 standard deviations below mean for gender and age)

LIVER ALLOGRAFT PATHOLOGY Timeline • 1968 ○ Dr. K. A. Porter

Liver Transplantation

Major Milestones in Liver Transplantation

– Early description of human liver allograft dysfunction, including allograft rejection • 1984 ○ Dr. D. Snover et al – Systematic analysis of pathologic features of T-cellmediated rejection • 1985 ○ Dr. A. J. Demetris et al – Comprehensive description of histologic features of liver allograft dysfunction • 1995 ○ Banff classification of liver allograft pathology – Update on chronic rejection in 1999

SELECTED REFERENCES 1. 2. 3. 4. 5. 6.

7.

8. 9.

10.

11. 12. 13.

14.

Busuttil RW et al: The first report of orthotopic liver transplantation in the Western world. Am J Transplant. 12(6):1385-7, 2012 Vilarinho S et al: Liver transplantation: from inception to clinical practice. Cell. 150(6):1096-9, 2012 Starzl TE et al: Themes of liver transplantation. Hepatology. 51(6):1869-84, 2010 Groth CG: Forty years of liver transplantation: personal recollections. Transplant Proc. 40(4):1127-9, 2008 Otte JB: History of pediatric liver transplantation. where are we coming from? where do we stand? Pediatr Transplant. 6(5):378-87, 2002 Starzl TE: The saga of liver replacement, with particular reference to the reciprocal influence of liver and kidney transplantation (1955-1967). J Am Coll Surg. 195(5):587-610, 2002 Demetris A et al: Update of the International Banff Schema for Liver Allograft Rejection: working recommendations for the histopathologic staging and reporting of chronic rejection. An International Panel. Hepatology. 31(3):792-9, 2000 Banff schema for grading liver allograft rejection: an international consensus document. Hepatology. 25(3):658-63, 1997 Demetris AJ et al: Pathology of hepatic transplantation: a review of 62 adult allograft recipients immunosuppressed with a cyclosporine/steroid regimen. Am J Pathol. 118(1):151-61, 1985 Snover DC et al: Orthotopic liver transplantation: a pathological study of 63 serial liver biopsies from 17 patients with special reference to the diagnostic features and natural history of rejection. Hepatology. 4(6):1212-22, 1984 Penn I et al: Liver transplantation in man. Ann N Y Acad Sci. 170:251-258, 1970 Porter KA: Pathology of liver transplantation. Transplant Rev. 2:129-70, 1969 Starzl TE et al: Clinical and pathologic observations after orthotopic transplantation of the human liver. Surg Gynecol Obstet. 128(2):327-39, 1969 Starzl TE et al: Orthotopic homotransplantation of the human liver. Ann Surg. 168(3):392-415, 1968

247

Liver Transplantation

Preservation Injury KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Tissue damage sustained during harvesting, preservation, transportation, and reperfusion

• Hepatocyte ballooning and microvesicular/small droplet steatosis at zone 3, imparting distinct pale appearance on low-power view • Hepatocyte detachment from each other, scattered acidophil bodies, and spotty necrosis • Confluent zone 3 necrosis in severe cases, can also involve periportal areas • Cytoplasmic and canalicular cholestasis, more pronounced at zone 3 • Varying degree of neutrophilic infiltrates in lobules • No significant portal inflammation in general

ETIOLOGY/PATHOGENESIS • Cold ischemia ○ Prolonged storage in preservation solutions • Warm ischemia ○ Compromised blood flow to liver at body temperature before and during harvesting ○ Resumption of blood flow after implantation

CLINICAL ISSUES • Elevation of serum transaminases and poor bile production within 24-48 hours after revascularization • Enzyme levels typically decrease progressively within several days if graft survives injury • Complete resolution in most cases • Graft failure in rare occasions (primary nonfunction)

TOP DIFFERENTIAL DIAGNOSES • • • • •

Antibody-mediated rejection Hepatic artery thrombosis Hepatic vein stenosis and thrombosis Biliary obstruction "Surgical hepatitis"

Zone 3 Hepatocyte Ballooning

Zone 3 Necrosis

Coagulative Necrosis

Lipopeliosis

(Left) Mild preservation injury features hepatocyte ballooning around the terminal hepatic venule ﬊, imparting a distinctive pale appearance at zone 3 on lowpower view. Note the presence of a small unremarkable portal tract ﬈. (Right) This case of preservation injury shows zone 3 necrosis ﬊. Note the presence of microvesicular/small droplet steatosis and occasional neutrophils in the lobules.

(Left) This posttransplant day 1 biopsy shows severe preservation injury with extensive coagulative necrosis. Clinically, the allograft showed primary nonfunction that required retransplantation. (Right) This posttransplant day 6 biopsy shows large fat droplets in extracellular spaces ﬈, which are released from damaged steatotic hepatocytes secondary to preservation injury. Note the presence of inflammatory cells around fat droplets.

248

Preservation Injury

MICROSCOPIC

Synonyms

Histologic Features

• Preservation/reperfusion injury • Ischemia and reperfusion injury • Harvesting injury

• Hepatocyte injury, primarily at zone 3 ○ Hepatocyte ballooning and microvesicular/small droplet steatosis – Imparts distinct pale appearance on low-power view ○ Hepatocyte detachment from each other, scattered acidophil bodies, and spotty necrosis ○ Confluent necrosis in severe cases – Can also involve periportal areas ○ Damaged hepatocytes may release existing fat into extracellular spaces (lipopeliosis) • Varying degree of neutrophilic infiltrates in lobules ○ No significant portal inflammation, in general • Biliary/cholestatic changes ○ Cytoplasmic and canalicular cholestasis – More pronounced at zone 3 ○ Bile duct degeneration and detachment of duct epithelium from basement membrane ○ Ductular reaction, sometimes with ductular cholestasis and bile plugs ○ No significant bile duct injury in most cases • Resolving preservation injury with regenerative changes of hepatocytes ○ Increased mitotic activity, nuclear enlargement, frequent binucleation, and thickened cell plates ○ Mild ballooning and cytoplasmic cholestasis may persist for several weeks ○ Zone 3 histiocytes and other inflammatory cells

Definitions • Tissue damage sustained during harvesting, preservation, transportation, and reperfusion • One of major causes of initial graft dysfunction

ETIOLOGY/PATHOGENESIS Ischemic Injury • 4 stages ○ Prepreservation injury ○ Cold preservation ○ Rewarming ○ Reperfusion injury • 2 types of ischemic injury ○ Cold ischemia – Prolonged storage in preservation solutions (< 12 hours) – Preferentially targets sinusoidal endothelial cells ○ Warm ischemia – Compromised blood flow to liver at body temperature before and during harvesting – Resumption of blood flow after implantation – Primarily damages hepatocytes • Bile duct epithelium, Kupffer cells, and Ito cells sensitive to both cold and warm ischemia • Severity of injury depends on type and duration of ischemic stress • Preexisting donor risk factors ○ Severe large droplet macrovesicular steatosis (> 30%) ○ Donation after cardiac death ○ Prolonged stay in intensive care unit ○ Increasing donor age

CLINICAL ISSUES Presentation • Elevation of serum transaminases and poor bile production within first 24-48 hours after revascularization ○ Enzyme levels typically decrease progressively within several days if graft survives injury ○ Abnormal liver tests may persist for several months if injury is severe • Clinical resolution usually observed within 1-4 weeks

Treatment • No specific therapy

Prognosis • Complete resolution in most cases ○ Graft failure in rare occasions (primary nonfunction) • Higher incidence of subsequent acute and chronic rejection • Higher incidence of biliary complications, such as ischemic cholangiopathy

Liver Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Antibody-Mediated Rejection • Preformed donor-reactive antibodies in recipient • Hemorrhagic necrosis of graft with fibrin thrombi • Fibrinoid necrosis of hepatic artery branches

Hepatic Artery Thrombosis • Doppler ultrasound and angiography are diagnostic • Massive or zone 3 coagulative necrosis • Bile duct necrosis

Hepatic Vein Stenosis and Thrombosis • Doppler ultrasound and venography are diagnostic • Zone 3 congestion, hemorrhage, and necrosis

Biliary Obstruction • Cholangiography is diagnostic • Portal edema, prominent ductular reaction, and neutrophilic portal infiltrates • Portal and periportal fibrosis may develop rapidly

"Surgical Hepatitis" • Clusters of neutrophils in sinusoids, without necrosis

SELECTED REFERENCES 1.

2.

Cannistrà M et al: Hepatic ischemia reperfusion injury: a systematic review of literature and the role of current drugs and biomarkers. Int J Surg. 33 Suppl 1:S57-70, 2016 Ali JM et al: Analysis of ischemia/reperfusion injury in time-zero biopsies predicts liver allograft outcomes. Liver Transpl. 21(4):487-99, 2015

249

Liver Transplantation

Hepatic Artery Thrombosis KEY FACTS

TERMINOLOGY • Thrombotic occlusion of hepatic artery &/or its branches • Ischemic cholangiopathy ○ Bile duct ischemia ○ Complications resulting from hepatic artery thrombosis or other causes

• Can lead to bile leak or fulminant hepatic failure • Long-term complications include bile duct stricture and duct loss

MICROSCOPIC

• Anastomotic complication after liver transplantation ○ Rare in native livers • Bile ducts solely dependent on arterial flow ○ Subject to ischemic injury • May also cause ischemia of hepatocytes and portal connective tissue

• Bile duct epithelial cell necrosis with formation of eosinophilic casts of sloughed epithelial cells ○ Bile leakage into portal connective tissue • Necrosis of hepatocytes and portal connective tissue • In time, chronic ischemia can lead to ○ Biliary strictures ○ Fibrosis ○ Duct loss • May develop secondary infection and abscesses

CLINICAL ISSUES

DIAGNOSTIC CHECKLIST

• Most frequent vascular complication of liver transplantation • Presentation and prognosis depend on severity and timing

• Biopsy may not be representative ○ Findings may be patchy ○ Large ducts not sampled

ETIOLOGY/PATHOGENESIS

Border of Hepatic Infarct Due to Hepatic Artery Thrombosis

Bile Leak in Patient With Hepatic Artery Thrombosis

Eosinophilic Bile Cast

Ischemic Bile Duct Injury

(Left) This section demonstrates a welldelineated area of hepatic parenchymal infarction ſt following hepatic artery thrombosis. (Right) An explanted allograft liver demonstrates an area of bile duct necrosis and a bile leak ſt in a patient who developed hepatic artery thrombosis after transplantation.

(Left) This example of hepatic artery thrombosis shows ischemic bile duct injury with an eosinophilic bile cast ﬈. The cast is the remnants of sloughed, necrotic biliary epithelial cells. (Right) This bile duct in a patient with hepatic artery thrombosis shows evidence of biliary epithelial cell injury st. The biliary epithelial cells are flattened, unevenly spaced, and show focal vacuolization. Unlike acute rejection, however, the duct is not inflamed.

250

Hepatic Artery Thrombosis • Computed tomography angiography ○ Used as confirmatory test

Abbreviations • Hepatic artery thrombosis (HAT)

MACROSCOPIC

Definitions

General Features

• Thrombotic occlusion of hepatic artery &/or its branches • Ischemic cholangiopathy: Bile duct ischemia and complications resulting from HAT or other causes

• May appear grossly normal • Mottled liver parenchyma • Foci of parenchymal necrosis and bile leak

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

Causes of HAT

Histologic Features

• Anastomotic complication after liver transplantation • Rare in native livers

• Bile duct necrosis ○ Denuded, necrotic bile duct epithelium ○ Eosinophilic bile casts of sloughed, necrotic biliary epithelial cells ○ Bile leakage into periductal connective tissue • Necrosis of hepatocytes and portal connective tissue seen in hepatic infarcts • May develop secondary infection and abscesses • Ongoing ischemic injury leads to biliary strictures, fibrosis, and duct loss

Ischemic Injury • Bile ducts solely dependent on arterial flow ○ Acute ischemia leads to biliary ulcers, necrosis, and bile leaks ○ Chronic ischemia leads to scarring, strictures, and duct loss • Localized ischemia of hepatocytes, bile ducts, and portal connective tissue

CLINICAL ISSUES Epidemiology • Incidence ○ 2nd most common cause of liver graft failure ○ Most frequent vascular complication of liver transplantation ○ Transplanted livers most susceptible early after transplant – Allografts lack anastomosing collateral blood supply □ More dependent on arterial inflow – Pediatric and split-liver grafts at greatest risk due to smaller vessels and greater technical difficulty

DIFFERENTIAL DIAGNOSIS Recurrent Primary Sclerosing Cholangitis • May be indistinguishable from chronic ischemia based on histology alone • History of primary sclerosing cholangitis in native liver

Acute or Chronic Allograft Rejection • Acute rejection shows typical inflammatory infiltrates and endotheliitis • Chronic rejection shows senescent duct changes and duct loss ○ Associated with inadequate immunosuppressive medications or multiple episodes of acute rejection

Presentation

Other Biliary Complications

• Varies with acuity and ensuing complication ○ Fever, abdominal pain, and jaundice ○ Bile peritonitis ○ Fulminant hepatic failure • Cholestatic liver function abnormalities ○ Elevated bilirubin, alkaline phosphatase, γ-glutamyl transferase

• Biliary obstruction may present with similar clinical picture • Imaging studies used to exclude obstruction

Treatment

• Shows C4d staining and donor-specific antibodies

• Medical drugs for thrombolysis • Surgical thrombectomy or endovascular revascularization • Liver transplantation/retransplantation

Prognosis • Depends on severity and timing • Can lead to bile leak or fulminant hepatic failure • Long-term complications include bile duct stricture and duct loss

IMAGING

Ischemic Hepatitis • Zone 3 hepatocyte injury due to ischemia of any cause • Only see bile duct ischemia if loss of arterial flow

Antibody-Mediated Rejection

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Biopsy may not be representative ○ Findings may be patchy ○ Large ducts often not sampled

SELECTED REFERENCES 1. 2.

Radiographic Findings • Doppler ultrasound

Liver Transplantation

TERMINOLOGY

3.

Mourad MM et al: Aetiology and risk factors of ischaemic cholangiopathy after liver transplantation. World J Gastroenterol. 20(20):6159-69, 2014 Bekker J et al: Early hepatic artery thrombosis after liver transplantation: a systematic review of the incidence, outcome and risk factors. Am J Transplant. 9(4):746-57, 2009 Deltenre P et al: Ischemic cholangiopathy. Semin Liver Dis. 28(3):235-46, 2008

251

Liver Transplantation

Portal Vein Thrombosis KEY FACTS

TERMINOLOGY • Thrombotic occlusion of portal vein &/or its branches • Portal vein stenosis or stricture can also lead to ischemic injury

ETIOLOGY/PATHOGENESIS • Vascular complication during early or late posttransplant period

CLINICAL ISSUES • Presentation varies with clinical setting ○ May be clinically silent or present with ascites and allograft dysfunction • Can lead to severe hepatic dysfunction and fulminant hepatic failure • Treatment includes restoring portal flow ○ Thrombolysis or thrombectomy with recanalization ○ May require retransplantation

• Transjugular intrahepatic portosystemic shunt alleviates portal hypertension • Liver transplantation or retransplantation may be indicated in some cases • Can be clinically silent or present with allograft dysfunction

MICROSCOPIC • • • • • •

Findings may be patchy and not sampled by needle biopsy Depends on timing, location, and extent of thrombosis Parenchymal ischemic changes and hemorrhage Hepatocyte swelling and necrosis May see thrombus in portal vein branches Late changes include nodular regenerative hyperplasia

TOP DIFFERENTIAL DIAGNOSES • Infection causes nonzonal ("geographic") necrosis • Hepatic artery thrombosis distinguished with imaging studies

Organizing Thrombus in Portal Vein Branch

Periportal Hemorrhage and Ischemic Necrosis

Portal and Periportal Hemorrhage

Large Areas of Hemorrhage in Failed Allograft

(Left) H&E-stained slide from an explanted, failed allograft shows extension of a portal vein thrombus into smaller portal vein branches ﬊. There is also periportal ductal cholestasis ﬉. (Right) H&Estained slide shows marked portal and periportal hemorrhage st in a patient with posttransplant portal vein thrombosis. Also noted is ischemic necrosis of periportal hepatocytes ﬈.

(Left) H&E-stained section shows hemorrhage in the portal tract ﬈ and adjacent lobule ﬇ in a patient with portal vein thrombosis after transplant. (Right) H&Estained section from an explanted failed allograft shows a large area of hemorrhage ﬊.

252

Portal Vein Thrombosis

IMAGING

Abbreviations

Radiographic Findings

• Portal vein thrombosis (PVT)

• Doppler ultrasound • Angiography usually diagnostic

Definitions • Thrombotic occlusion of portal vein &/or its branches • Portal vein stenosis or stricture may also lead to ischemic graft injury

ETIOLOGY/PATHOGENESIS Vascular Complication After Liver Transplantation • Can occur in early or late posttransplant period

Native Liver Involvement • Frequent complication of cirrhosis or hepatocellular carcinoma • Other risk factors include ○ Obstruction ○ Inflammation ○ Hypercoagulable state

CLINICAL ISSUES

MACROSCOPIC General Features

Liver Transplantation

TERMINOLOGY

• May appear grossly normal, especially on surface • Mottled liver parenchyma with foci of parenchymal necrosis

MICROSCOPIC Histologic Features • Depend on timing, location, and extent of thrombosis ○ Findings may be patchy and not sampled by needle biopsy • Parenchymal ischemic changes ○ Hepatocyte swelling and necrosis • Hemorrhage • May see thrombus in portal vein branches • Late changes include nodular regenerative hyperplasia • Ductal cholestasis may be seen

Epidemiology • Incidence ○ Portal vein complications occur in < 2% of liver transplant recipients – Occurs in up to 12% of pediatric liver transplant recipients – Associated with technical complications, small portal vein diameter, pediatric recipient, and pretransplant PVT ○ Transplanted livers most susceptible early after transplant

Presentation • Varies with clinical setting ○ Symptoms related to acuity and ensuing complications • Can be clinically silent or present with allograft dysfunction ○ May present as acute liver failure ○ Cholestatic liver enzyme abnormalities • Ascites ○ Can be massive and lead to hemodynamic instability • Portal hypertension ○ May develop complications of portal hypertension such as variceal bleeding

Treatment • Restore portal flow with thrombolysis or thrombectomy and recanalization ○ ± stent placement • Transjugular intrahepatic portosystemic shunt ○ Alleviates portal hypertension • Liver transplantation/retransplantation

Prognosis • Good allograft and patient survival with recognition and treatment • Can lead to massive hepatic necrosis, allograft loss, or death

DIFFERENTIAL DIAGNOSIS Acute or Chronic Allograft Rejection • Acute rejection shows typical rejection-type infiltrates and endotheliitis • Chronic rejection shows senescent duct changes; associated with refractory or untreated acute rejection

Biliary Complications • Biliary obstruction or biliary ischemia

Ischemic Hepatitis • Often more diffuse process with zone 3 hepatocyte injury • Occurs with parenchymal ischemia of any cause

Hepatic Artery Thrombosis • Frequently results in bile duct ischemia and necrosis • May also cause parenchymal necrosis • Imaging studies aid distinction from PVT

Infection • Causes nonzonal ("geographic") necrosis • Herpes hepatitis and adenovirus infection exhibit viral inclusions in hepatocytes • Immunohistochemistry aids in identification of viral inclusions

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Needle biopsy may not be representative

SELECTED REFERENCES 1.

2. 3.

Jensen MK et al: Management and long-term consequences of portal vein thrombosis after liver transplantation in children. Liver Transpl. 19(3):315-21, 2013 Adeyi O et al: Liver allograft pathology: approach to interpretation of needle biopsies with clinicopathological correlation. J Clin Pathol. 63(1):47-74, 2010 Ponziani FR et al: Portal vein thrombosis: insight into physiopathology, diagnosis, and treatment. World J Gastroenterol. 16(2):143-55, 2010

253

Liver Transplantation

Bile Duct Stricture, Leak, Sludge, Biloma KEY FACTS

TERMINOLOGY • Spectrum of posttransplant biliary complications leading to graft dysfunction

ETIOLOGY/PATHOGENESIS • Bile duct stricture ○ Anastomotic causes (technical complications) ○ Nonanastomotic causes (ischemic &/or immunological mechanisms) • Bile leak: Occurs at anastomotic or T-tube insertion site • Bile sludge: Thick collection of mucus, calcium bicarbonate, and cholesterol crystals ○ Leads to cast and stone formation if left untreated • Biloma: Localized bile collection outside biliary system

CLINICAL ISSUES • Incidence of overall biliary complications: ~ 23% ○ Biliary stricture: ~ 15% (higher incidence for living-donor transplantation); bile leaks: ~ 8.5%

• 2/3 of biliary complications occur in first 3 months ○ Technical causes: Usually occur soon after transplantation ○ Nontechnical causes: Usually occur later

MICROSCOPIC • Features of large bile duct obstruction ○ Portal edema, ductular reaction, and mixed inflammatory cell infiltrates, usually with prominent neutrophils ○ Canalicular cholestasis ○ Portal and periportal fibrosis and eventual progression to biliary cirrhosis in chronic cases • Bile lakes and bile infarcts • Features of ascending cholangitis

TOP DIFFERENTIAL DIAGNOSES • Recurrent primary sclerosing cholangitis • Acute antibody-mediated rejection

Large Bile Duct Obstruction

Portal and Periportal Fibrosis

Ascending Cholangitis

Bile Duct Necrosis and Ulceration

(Left) This case of posttransplant bile duct stricture shows features of large bile duct obstruction characterized by marked portal edema, prominent ductular reaction ﬈, and mild mixed inflammatory cell infiltrates rich in neutrophils. Mild bile duct injury is noted ﬊. (Right) Trichrome stain shows portal and periportal fibrosis in a liver biopsy from a patient who developed posttransplant bile duct stricture due to ischemia. Note the presence of proliferating ductules st.

(Left) This case shows features of ascending cholangitis due to bacterial infection as a consequence of posttransplant biliary complications. Note the presence of neutrophils in the duct lumen ﬊. Bile duct damage and portal edema are evident. (Right) This case shows ulceration of the extrahepatic large bile duct at the anastomotic site. There are granulation tissue formation, fibrosis, and inflammatory cell infiltrates, leading to stricture formation. The lumen is only partially lined by biliary epithelium ﬊.

254

Bile Duct Stricture, Leak, Sludge, Biloma

Synonyms • Large bile duct obstruction

Definitions • Spectrum of posttransplant biliary complications leading to graft dysfunction

ETIOLOGY/PATHOGENESIS Bile Duct Stricture • Anastomotic causes (technical complications) ○ Fibrotic nature of healing process ○ Use of split, living-related, or small-for-size livers ○ Improper surgical techniques ○ Choledochojejunostomy-type biliary reconstruction • Nonanastomotic causes (ischemic &/or immunological mechanisms) ○ Hepatic artery thrombosis or stenosis ○ Obliterative arteriopathy due to chronic rejection ○ Arterial steal syndrome ○ Use of livers from extended criteria donors ○ Use of ABO incompatible livers ○ Prolonged cold ischemia time ○ Severe preservation injury

Bile Leak • Usually occurs at anastomotic or T-tube insertion site • May result from ○ Poor suturing or excessive tension on biliary anastomosis ○ Excessive dissection of periductal tissue or excessive use of electrocautery during organ procurement ○ Local tissue ischemia ○ T-tube dislodgement or removal

Biliary Sludge • Thick collection of mucus, calcium bicarbonate, and cholesterol crystals • Results from reduced bile flow or increased viscosity due to obstruction, ischemia, or infection • Leads to cast and stone formation if left untreated

Biloma • Localized bile collection outside biliary system

CLINICAL ISSUES Epidemiology • Incidence of overall biliary complications: ~ 23% ○ Biliary stricture: ~ 15% (higher incidence for living-donor transplantation); bile leaks: ~ 8.5%

Laboratory Tests • Elevation of bilirubin, γ-glutamyl transpeptidase, and alkaline phosphatase levels • Mild elevation of serum aminotransferase levels • Elevation of urine bilirubin levels • Leukocytosis in patients with cholangitis and abscess

Treatment

Liver Transplantation

TERMINOLOGY

• Medical treatment with ursodeoxycholic acid • Repeated balloon dilatation of stricture with stenting • Diversion of biliary flow for leaks ○ Unclamping of T-tubes, endoscopic sphincterotomy ± stenting, nasobiliary drainage • Extraction of sludge, casts, and stones • Management of hepatic artery thrombosis • Surgical intervention • Retransplantation ○ Indications: Failed medical and surgical treatment, recurrent cholangitis, biliary cirrhosis

IMAGING General Features • Ultrasonography, cholangiography, abdominal CT ○ Locate strictured or dilated bile ducts, leaks, bilomas, sludge/casts/stones, abscesses, or T-tube dislodgement • Doppler ultrasound, angiography ○ Demonstrate hepatic artery thrombosis or stenosis

MICROSCOPIC Histologic Features • Features of large bile duct obstruction ○ Portal edema, ductular reaction, and mixed inflammatory cell infiltrates, usually with prominent neutrophils ○ Canalicular cholestasis ○ Portal and periportal fibrosis and eventual progression to biliary cirrhosis in chronic cases • Bile lakes and bile infarcts • Features of ascending cholangitis ○ Neutrophils infiltrating duct epithelium and within bile duct lumina ○ Bile duct necrosis and ulceration ○ Abscess formation

DIFFERENTIAL DIAGNOSIS Recurrent Primary Sclerosing Cholangitis • Confirmed pretransplant history • Usually occurs > 1 year post transplantation

Presentation

Acute Antibody-Mediated Rejection

• 2/3 of biliary complications occur in first 3 months ○ Technical causes: Usually occur soon after transplantation ○ Nontechnical causes: Usually occur later • Jaundice, pruritus, acholic stools, and dark urine • Fever in patients with bile leaks, cholangitis, or abscess • Signs of peritonitis in patients with bile leaks ○ May be asymptomatic if patient is on corticosteroids

• May show portal changes similar to biliary obstruction • Presence of antidonor antibodies • C4d immunostain may be helpful

SELECTED REFERENCES 1. 2.

Memeo R et al: Management of biliary complications after liver transplantation. World J Hepatol. 7(29):2890-5, 2015 Nemes B et al: Biliary complications after liver transplantation. Expert Rev Gastroenterol Hepatol. 9(4):447-66, 2015

255

Liver Transplantation

Hepatic Venous Outflow Obstruction KEY FACTS

TERMINOLOGY

CLINICAL ISSUES

• Hepatic injury due to impedance of blood flow out of liver • 3 categories according to level of obstruction ○ Heart: Congestive heart failure ○ Hepatic veins &/or inferior vena cava: Budd-Chiari syndrome ○ Terminal hepatic venules and sinusoids: Sinusoidal obstruction syndrome, formerly venoocclusive disease

• Painful hepatomegaly, ascites, jaundice, and liver failure

ETIOLOGY/PATHOGENESIS • Anastomotic stenosis &/or thrombosis of hepatic veins and inferior vena cava in transplant setting ○ Seen in 0.8-9.5% of liver transplants, but higher incidence in living-related or split transplants • Recurrent Budd-Chiari syndrome post transplantation ○ Seen in up to 10% of patients • Sinusoidal obstruction syndrome in liver transplants ○ Rare occurrence

MICROSCOPIC • Zone 3 (centrilobular) changes ○ Sinusoidal dilatation and congestion with ↑ pressure that leads to hepatocyte atrophy and dropout ○ Hemorrhage and hepatocyte necrosis ○ Perivenular and perisinusoidal fibrosis that may lead to fibrous narrowing or complete obliteration of terminal hepatic venules, bridging fibrosis, and cirrhosis • Nodular regenerative hyperplasia may occur

TOP DIFFERENTIAL DIAGNOSES • Other causes of sinusoidal dilatation ○ Nonspecific finding in liver transplants ○ Posttransplant portal vein thrombosis • Ischemic injury due to hepatic artery thrombosis • Isolated central perivenulitis

Zone 3 Sinusoidal Dilatation

Sinusoidal Dilatation With Atrophy

Red Cell Extravasation

Perisinusoidal Fibrosis

(Left) Liver biopsy from a patient with congestive heart failure shows centrilobular (zone 3) sinusoidal dilatation. Only mild hepatocyte atrophy is noted in this case. The terminal hepatic venule (central vein) ﬊ appears unremarkable. (Right) This liver biopsy from a patient with congestive heart failure shows marked centrilobular sinusoidal dilatation and congestion. Hepatocyte atrophy ﬉ due to increased sinusoidal pressure is evident.

(Left) Red blood cell st extravasation into the space of Disse, a perisinusoidal space between hepatocytes and sinusoidal endothelial cells ﬉, occurs due to increased sinusoidal pressure. (Right) Liver biopsy from a patient with congestive heart failure shows perisinusoidal collagen deposition st highlighted by trichrome stain. Note the presence of sinusoidal dilatation with red blood cells.

256

Hepatic Venous Outflow Obstruction ○ Depletion of glutathione, which plays role in hepatocyte necrosis

Abbreviations • Hepatic venous outflow obstruction (HVOO)

CLINICAL ISSUES

Synonyms

Presentation

• Congestive hepatopathy • Congestive hepatic venopathy

• Painful hepatomegaly, ascites, jaundice, and liver failure • Congestive heart failure: Usually progressive process • Budd-Chiari syndrome: Subacute (most common), fulminant, or chronic presentation • Sinusoidal obstruction syndrome: Acute or chronic (insidious) onset

Definitions • Hepatic injury due to impedance of blood flow out of liver • 3 categories according to level of obstruction ○ Heart: Congestive heart failure ○ Hepatic veins &/or inferior vena cava: Budd-Chiari syndrome ○ Terminal hepatic venules and sinusoids: Sinusoidal obstruction syndrome, formerly venoocclusive disease

Laboratory Tests

• Right heart failure, constrictive pericarditis, cardiac amyloidosis, etc.

• Mild elevation of serum aminotransferase levels ○ Marked elevation with severe heart failure or fulminant Budd-Chiari syndrome • Mild elevation of serum alkaline phosphatase and γglutamyl transpeptidase levels • Hyperbilirubinemia ○ > 2 mg/dL for sinusoidal obstruction syndrome • Decreased clearance of immunosuppressive medications such as tacrolimus (FK506, Prograf)

Budd-Chiari Syndrome

Treatment

• Thrombotic causes ○ Hypercoagulable states – Myeloproliferative disorders, antiphospholipid syndrome, pregnancy/postpartum, oral contraceptive use, malignancy, etc. ○ Coagulation factor or inhibitor deficiencies • Nonthrombotic causes ○ Membranous obstruction (fibrous web) ○ Compression or invasion by tumor or other lesions ○ Idiopathic: ~ 10% • Anastomotic stenosis &/or thrombosis in transplant setting ○ Seen in 0.8-9.5% of liver transplants but higher incidence in living-related or split transplants – Surgical anastomotic narrowing – Anastomotic kinking or twisting – Size mismatch between donor and recipient veins – Development of intimal hyperplasia and fibrosis or abnormal intimal flap – Piggyback technique for anastomosis • Recurrent Budd-Chiari syndrome post transplantation ○ Seen in up to 10% of patients

• Congestive heart failure ○ Treatment of underlying causes ○ Lifestyle modifications, including control of sodium and fluid intake ○ Medications to improve cardiac functions ○ Heart transplantation • Budd-Chiari syndrome: Decompression procedures ○ Transjugular intrahepatic portosystemic shunt – Considered for patients with acute presentation ○ Surgical decompression – Considered for nonfulminant or chronic presentation ○ Angioplasty with stent for membranous webs or short segment stenosis • Budd-Chiari syndrome: Medical therapies ○ Thrombolytic and anticoagulation therapies ○ Management of ascites • Sinusoidal obstruction syndrome ○ Fibrinolytic agents, such as tissue plasminogen activator ○ Antithrombotic agents, such as antithrombin III, heparin, and defibrotide ○ Transjugular intrahepatic portosystemic shunt ○ Supportive care • Liver transplantation for advanced liver injury ○ Cirrhosis ○ Fulminant liver failure ○ Biochemical evidence of advanced liver dysfunction

ETIOLOGY/PATHOGENESIS Congestive Heart Failure

Sinusoidal Obstruction Syndrome • Bone marrow or hematopoietic stem cell transplantation • Chemotherapy, particularly regimens with high doses of cyclophosphamide and busulfan • Hepatic radiation • Ingestion of pyrrolizidine alkaloids in herbal medicines and bush tea • Rare occurrence in liver transplants ○ Prevalence of 1.9% in early literature ○ Association with azathioprine (Imuran) use and episodes of T-cell-mediated rejection • Direct injury to sinusoidal endothelial cells and surrounding hepatocytes, mainly at zone 3 ○ High concentration of cytochrome P450 enzymes that metabolize many chemotherapeutic agents

Liver Transplantation

TERMINOLOGY

Prognosis • Congestive heart failure ○ Depends on nature and severity of underlying illness as well as patient's response to medications ○ Liver disease rarely contributes to mortality • Budd-Chiari syndrome ○ Poor for untreated patients – High mortality due to progressive liver failure ○ 38-90% 5-year survival rate after portosystemic shunt 257

Liver Transplantation

Hepatic Venous Outflow Obstruction ○ 65-90% 5-year survival rate after liver transplantation • Sinusoidal obstruction syndrome ○ Severe disease: High mortality (> 90%) ○ Mild or moderate disease: Complete resolution with no significant adverse effect

IMAGING Ultrasonographic Findings • Echocardiogram is useful diagnostic tool for congestive heart failure • Doppler US, hepatic venography, and MR are useful diagnostic tools for Budd-Chiari syndrome ○ Abnormal flow in hepatic veins ○ Large intrahepatic collateral vessels – May show "spider web" venous network • Doppler US is useful diagnostic tool for sinusoidal obstruction syndrome ○ Reversal of flow in portal vein

MACROSCOPIC General Features • "Nutmeg liver" with alternating dark and pale areas due to zone 3 congestion/hemorrhage surrounded by relatively normal or fatty liver parenchyma • Hepatomegaly

MICROSCOPIC Histologic Features • Zone 3 (centrilobular) changes ○ Sinusoidal dilatation and congestion with ↑ pressure – Hepatocyte atrophy and dropout – Red cell extravasation into space of Disse – May extend to zone 2 (mid zone) in severe cases ○ Overt hemorrhage and hepatocyte necrosis in acute and subacute cases ○ Perivenular and perisinusoidal fibrosis – Progression to central-central bridging fibrosis in chronic cases – Nodule formation with centrally located portal tracts ("reversed lobulation" or "cardiac cirrhosis") – Central-portal bridging fibrosis may occur as longterm effect, leading to cirrhosis ○ Fibrous narrowing or complete obliteration of terminal hepatic venules (central veins) – Characteristic lesion for sinusoidal obstruction syndrome – Early lesions: Subendothelial edema, red cell extravasation, fibrin, loose collagen deposition – Collagen becomes more dense with time – Recanalization may develop • Mild steatosis may be seen in viable hepatocytes • Lack of significant lobular inflammation • Cholestasis may be seen in severe cases but usually absent • Portal tracts usually lack significant inflammation, bile duct damage, or ductular reaction • Nodular regenerative hyperplasia may occur ○ Large regenerative nodules may develop in setting of Budd-Chiari syndrome, mimicking focal nodular hyperplasia or hepatocellular adenoma 258

DIFFERENTIAL DIAGNOSIS Other Causes of Sinusoidal Dilatation • Nonspecific finding in liver transplants ○ Altered blood supply ○ Medication effect • Posttransplant portal vein thrombosis ○ Doppler US is useful diagnostic tool ○ May show dilated &/or herniated portal venules • Liver parenchyma adjacent to mass lesion ○ May show inflammatory cell infiltrates and ductular reaction • Systemic inflammatory conditions ○ Castleman disease, Crohn disease, granulomatous disorders, rheumatoid arthritis, etc. • Extrahepatic neoplasms ○ Renal cell carcinoma, Hodgkin lymphoma, etc.

Zone 3 Ischemic Injury • Posttransplant hepatic artery thrombosis ○ Zone 3 hepatocyte swelling, necrosis, and cholestasis ○ No significant sinusoidal dilatation ○ Doppler US is useful diagnostic tool • Preservation injury ○ Typically resolves 2-4 weeks post transplantation ○ Rarely persists as long as 3 months

Isolated Central Perivenulitis • Endotheliitis of central veins with subendothelial mononuclear cell infiltration • Perivenular hepatocyte necrosis and dropout • No significant sinusoidal dilatation

Alcoholic Hepatitis • May show sclerosing hyaline necrosis leading to fibrous obliteration of central veins and noncirrhotic portal hypertension, mimicking sinusoidal obstruction syndrome • History of alcohol use • Steatosis and Mallory-Denk bodies • No significant sinusoidal dilatation

Drug-Induced Acute Hepatitis With Zone 3 Necrosis • • • •

Acetaminophen, niacin, cocaine, etc. History of drug intake Testing drug levels in blood or urine No significant sinusoidal dilatation

SELECTED REFERENCES 1. 2.

3. 4.

Valla DC: Budd-Chiari syndrome/hepatic venous outflow tract obstruction. Hepatol Int. ePub, 2017 Chu HH et al: Longterm outcomes of stent placement for hepatic venous outflow obstruction in adult liver transplantation recipients. Liver Transpl. 22(11):1554-1561, 2016 Fulgenzi A et al: Defibrotide in the treatment of hepatic veno-occlusive disease. Hepat Med. 8:105-113, 2016 Writing committee members et al: 2013 ACCF/AHA guideline for the management of heart failure: a report of the American College of Cardiology Foundation/American Heart Association Task Force on practice guidelines. Circulation. 128(16):e240-327, 2013

Hepatic Venous Outflow Obstruction

Zone 3 Hemorrhagic Necrosis (Left) Liver biopsy from a patient with Budd-Chiari syndrome shows zone 3 sinusoidal dilatation and hepatocyte atrophy ﬉ and dropout. The native central vein is difficult to recognize in this area. (Right) Zone 3 hemorrhage and necrosis ﬊ is seen in a case of acute BuddChiari syndrome. The central vein is obscured. Note the presence of an unremarkable portal tract ﬉.

Bridging Fibrosis

Liver Transplantation

Zone 3 Hepatocyte Dropout

Sinusoidal Obstruction Syndrome (Left) Liver biopsy from a patient with long-term hepatic venous outflow obstruction reveals central ﬊ to portal ﬈ bridging fibrosis (trichrome stain). Note the presence of sinusoidal dilatation and ductular reaction st in the fibrous septa. (Right) Trichrome stain of sinusoidal obstruction syndrome shows subepithelial edema ﬊ and wall fibrosis ﬈ of the central vein, leading to luminal narrowing. Note the presence of sinusoidal congestion and perisinusoidal collagen deposition st around the vein.

Central Vein Obliteration

Nodular Regenerative Hyperplasia (Left) This central vein is completely obliterated in the setting of sinusoidal obstruction syndrome. Perisinusoidal fibrosis is also evident. (Right) Reticulin stain highlights nodular regenerative hyperplasia that can be seen in the setting of venous outflow obstruction. It is characterized by small vague parenchymal nodules ﬈ separated by atrophic cell plates ﬈. Large regenerative nodules can also be seen in the setting of Budd-Chiari syndrome.

259

Liver Transplantation

Hyperperfusion Syndrome KEY FACTS

ETIOLOGY/PATHOGENESIS

• Portal hypertension with intractable ascites

• Excessive portal venous inflow that leads to graft dysfunction due to insufficient graft volume (small-for-size syndrome) ○ Split or living-related liver transplantation ○ Persistent hyperdynamic portal circulation ○ Compensatory decrease in hepatic artery flow (buffer response), leading to ischemic injury ○ Inadequate venous drainage • Predisposing factors ○ Prolonged warm and cold ischemia time ○ Graft steatosis (> 30% large droplet steatosis) ○ Advanced donor age (> 50 years) ○ High model for end-stage liver disease score (> 20) or Child-Pugh grade C recipient

MICROSCOPIC

CLINICAL ISSUES • Usually occurs within first 2 weeks post transplantation • Persistent hyperbilirubinemia

• Early changes ○ Endothelial denudation of portal veins and periportal sinusoids, leading to rupture and hemorrhage ○ Enhanced preservation injury with hepatocyte ballooning, necrosis, and even infarction ○ Sinusoidal dilatation and congestion ○ Hepatocyte regeneration with frequent binucleation and mitotic figures ○ Centrilobular canalicular cholestasis ○ Ischemic cholangitis ○ Ductular reaction and ductular cholestasis • Late changes ○ Small portal vein branch thrombosis with obliterative venopathy ○ Nodular regenerative hyperplasia ○ Biliary stricture

Sinusoidal Dilatation

Cholestasis

Regenerative Changes

Ischemic Cholangitis

(Left) Hyperperfusion syndrome (small-for-size syndrome) shows sinusoidal dilatation and congestion ﬉. Note the presence of focal hepatocyte necrosis ſt in this case. (Right) Marked canalicular cholestasis ﬉ is seen in a case of hyperperfusion syndrome. Also note the presence of frequent binucleated hepatocytes ſt, consistent with regenerative changes.

(Left) This case of hyperperfusion syndrome shows prominent features of hepatocyte regeneration with frequent binucleation ﬉ and mitosis ſt. Canalicular cholestasis is evident. (Right) This case of hyperperfusion syndrome exhibits features of ischemic cholangitis. The injured bile duct shows epithelial detachment from the basement membrane ﬊ and sloughing into duct lumen ﬉. Note the presence of portal veins with endothelial detachment ſt.

260

Hyperperfusion Syndrome

IMAGING

Synonyms

General Features

• Small-for-size syndrome (SFSS)

• Cholangiography to rule out biliary obstruction • Doppler ultrasonography and angiography may show arterial narrowing, thrombosis, and poor liver filling

Definitions • Excessive portal venous inflow that leads to graft dysfunction due to insufficient graft volume

ETIOLOGY/PATHOGENESIS Excessive Portal Venous Flow Into Graft • Split or living-related liver transplantation ○ Small-for-size graft: Graft:recipient body weight ratio < 0.8% or graft volume-to-standard liver volume of recipient < 30% or < 35% ○ Incidence of SFSS: ~ 5-20% – Left lobe grafts > right lobe grafts • Persistent hyperdynamic portal circulation and high portal blood inflow in recipients ○ Shear stress leading to endothelial cell injury and microcirculatory disturbance • Compensatory decrease in hepatic artery flow (buffer response), leading to ischemic injury

Inadequate Venous Drainage • More common in right lobe grafts: Middle hepatic vein or segment 5 and 8 veins are not reconstructed

Predisposing Factors • • • •

Prolonged warm and cold ischemia time Graft steatosis (> 30% large droplet steatosis) Advanced donor age (> 50 years) High model for end-stage liver disease score (> 20) or ChildPugh grade C recipient ○ Preexisting high portal pressure (≥ 15 mm Hg)

CLINICAL ISSUES Presentation • Usually occurs within first 2 weeks post transplantation ○ May occur beyond 1st month • Persistent hyperbilirubinemia • Delayed synthetic function with coagulopathy • Portal hypertension with intractable ascites • Gastrointestinal variceal bleeding, acidosis, renal failure, encephalopathy, and septic complications in severe cases

Laboratory Tests • Markedly elevated serum bilirubin levels (> 10 mg/dL)

Treatment • Reduction of portal blood inflow and portal pressure ○ Portosystemic shunt ○ Splenectomy or splenic artery ligation/embolization ○ Portal vein banding • Pharmacological treatment: Experimental ○ Attenuation of portal shear stress ○ Enhancement of liver regeneration • Retransplantation

Prognosis

MICROSCOPIC

Liver Transplantation

TERMINOLOGY

Histologic Features • Early changes ○ Endothelial denudation of portal veins and periportal sinusoids, leading to rupture and hemorrhage ○ Enhanced preservation injury with hepatocyte ballooning, necrosis, and even infarction ○ Sinusoidal dilatation and congestion ○ Hepatocyte regeneration with frequent binucleation and mitotic figures ○ Centrilobular canalicular cholestasis ○ Ischemic cholangitis ○ Ductular reaction and ductular cholestasis • Late changes ○ Small portal vein branch thrombosis with obliterative venopathy ○ Nodular regenerative hyperplasia ○ Biliary stricture

DIFFERENTIAL DIAGNOSIS Primary Nonfunction • Failure to produce bile and severe coagulopathy immediately following revascularization

Hyperacute Antibody-Mediated Rejection • Presence of preformed donor-reactive antibodies ○ Major ABO blood group isoagglutinins (ABOincompatible transplants) • Fibrin thrombi in portal and central veins

Vascular Thrombosis or Anastomotic Stricture • May occur in hepatic artery, portal vein, or hepatic vein • Doppler ultrasonography and angiography are diagnostic

Preservation Injury • Clinical resolution usually observed within 1-4 weeks

Posttransplant Biliary Complications • Portal edema, ductular reaction, and inflammatory cell infiltrates rich in neutrophils • Cholangiography is diagnostic

SELECTED REFERENCES 1.

2.

3.

4.

Goldaracena N et al: Small-for-size syndrome in live donor liver transplantation-pathways of injury and therapeutic strategies. Clin Transplant. 31(2), 2017 Rajakumar A et al: Small-for-size syndrome: bridging the gap between liver transplantation and graft recovery. Semin Cardiothorac Vasc Anesth. 21(3):252-261, 2017 Shoreem H et al: Small for size syndrome difficult dilemma: lessons from 10 years single centre experience in living donor liver transplantation. World J Hepatol. 9(21):930-944, 2017 Troisi RI et al: Graft inflow modulation in adult-to-adult living donor liver transplantation: a systematic review. Transplant Rev (Orlando). 31(2):127135, 2017

• High mortality rate without retransplantation 261

Liver Transplantation

T-Cell-Mediated Rejection, Liver KEY FACTS

TERMINOLOGY • Immune-mediated inflammation and injury due to genetic mismatch of allograft and recipient, characterized by Tlymphocyte infiltrates • Distinct patterns of early and late T-cell-mediated rejection (TCMR) now recognized

CLINICAL ISSUES • Often asymptomatic but may present with fever, abdominal pain, or nonspecific elevation of liver chemistries • Most cases of TCMR respond to high-dose corticosteroids

MICROSCOPIC • Mixed portal inflammatory cell infiltrates ○ Enlarged, activated, or blastic lymphocytes, eosinophils, neutrophils, macrophages • Bile duct damage ○ Lymphocytic infiltrates and biliary epithelial injury • Subendothelial venous inflammation (endotheliitis)

○ Venous inflammation with lifting and denudation of endothelial cells • May be graded as mild, moderate, or severe or with rejection activity index • Biopsy remains gold standard for diagnosis of TCMR • Classic triad of TCMR features are most characteristic of early TCMR ○ Mixed portal inflammatory cell infiltrates ○ Bile duct damage ○ Endotheliitis ○ 2 of 3 features generally required for diagnosis of early TCMR

TOP DIFFERENTIAL DIAGNOSES • • • • •

Antibody-mediated rejection Chronic viral hepatitis (hepatitis B or C) Biliary complications Recurrent autoimmune hepatitis Posttransplant lymphoproliferative disorder

Mixed Portal Inflammatory Cell Infiltrates in T-Cell-Mediated Rejection

Bile Duct Damage in T-Cell-Mediated Rejection

Portal Vein Endotheliitis in T-CellMediated Rejection

Bile Duct Damage in T-Cell-Mediated Rejection

(Left) H&E stain demonstrates mixed portal inflammatory cell infiltrate in early T-cellmediated rejection (TCMR). The infiltrate includes enlarged, activated lymphocytes, eosinophils, plasma cells, and histiocytes. (Right) H&E stain shows a markedly injured bile duct ﬈ in an inflamed portal tract. The bile duct epithelium is flattened and infiltrated by inflammatory cells.

(Left) H&E stain demonstrates endotheliitis. There is inflammation of the vein wall and associated endothelial cell lifting ſt. (Right) H&E stain demonstrates evidence of bile duct damage ſt. The injured duct shows nuclear hyperchromasia, pleomorphism, uneven spacing, and loss of polarity.

262

T-Cell-Mediated Rejection, Liver

Abbreviations • T-cell-mediated rejection (TCMR)

Synonyms • Acute rejection • Acute cell-mediated rejection • Acute cellular rejection

Definitions • Immune-mediated inflammation and injury of liver allograft ○ Due to genetic mismatch ○ Characterized by T-lymphocyte infiltrates • Distinct patterns of early and late TCMR now recognized ○ Classic histologic picture of TCMR now recognized as characteristic of early TCMR ○ Morphologically distinct patterns of late TCMR also include – Plasma cell-rich rejection – Central perivenulitis – Idiopathic posttransplant chronic hepatitis ○ Early and late patterns are not strictly time delineated and do exhibit overlap – Distinction between early and late variably defined as 3, 6, or 12 months after transplant

ETIOLOGY/PATHOGENESIS Immune-Mediated Inflammatory Process • Recipient immune system recognizes donor antigens in liver allograft as foreign • T-lymphocyte-mediated bile duct, endothelium, and, in some cases, hepatocyte injury

CLINICAL ISSUES Epidemiology • Incidence ○ Affects 12-15% of liver allograft recipients within first 2 years after transplantation – Vast majority occur within 1st year ○ All liver transplant recipients susceptible ○ Risk factors for TCMR – Younger, healthier patients – Older donor age (> 30 years) – Longer cold ischemia time

Presentation • • • • •

Fever Abdominal pain Hepatomegaly Ascites Often asymptomatic

Laboratory Tests • Nonspecific • May see elevations of ○ Transaminases ○ Bilirubin ○ Alkaline phosphatase &/or gamma-glutamyl transferase (GGT)

Natural History • Variable course • Untreated or refractory TCMR can lead to ○ Rapid allograft failure ○ Chronic rejection

Treatment • Drugs ○ Increase immunosuppression – Corticosteroids are standard therapy – May also be managed by adjusting baseline immunosuppression – Other immunosuppressants may be added for steroid-resistant rejection

Liver Transplantation

TERMINOLOGY

Prognosis • Prognosis very good for treated TCMR ○ Most respond to high-dose corticosteroids • Associated with increased risk of chronic rejection ○ Risk of chronic rejection increases with refractory, untreated, or recurrent TCMR

Timing • Can occur any time after transplantation ○ Late TCMR associated with medication changes or noncompliance • Most often presents within first 6 weeks after transplant

MICROSCOPIC Predominant Pattern/Injury Type • Inflammatory

Histologic Features • Biopsy remains gold standard for diagnosis of TCMR • Classic triad of TCMR features are most characteristic of early TCMR ○ Mixed portal inflammatory cell infiltrates ○ Bile duct damage ○ Endotheliitis ○ 2 of 3 features generally required for diagnosis of early TCMR – Features are not specific for TCMR • Mixed portal inflammatory cell infiltrates ○ Lymphocytes are enlarged, blastic, or activatedappearing – Predominantly CD4- and CD8-positive T lymphocytes ○ Lymphocytes admixed with – Eosinophils – Neutrophils – Macrophages – Plasma cells ○ Portal infiltrates may be more lymphocyte-predominant and less activated-appearing in late TCMR • Bile duct damage ○ Lymphocytic infiltration of interlobular bile ducts ○ Biliary epithelial cell injury – Eosinophilia – Vacuolization – Irregular cell shape and uneven lumen – Pleomorphism and uneven spacing of nuclei 263

Liver Transplantation

T-Cell-Mediated Rejection, Liver • Subendothelial venous inflammation (endotheliitis) ○ Venous inflammation with lifting and denudation of endothelial cells ○ Can affect portal veins &/or terminal hepatic venules ○ In severe TCMR, associated with perivenular parenchymal necrosis ○ Endotheliitis rarely affects hepatic arterioles • May see centrilobular perivenular parenchymal necrosis in severe cases of classic early TCMR • Central perivenulitis (perivenular inflammation and hepatocyte dropout) ○ May occur in conjunction with typical portal findings of TCMR or as isolated finding ○ More commonly seen in isolation in late TCMR ○ Venous endotheliitis may be present – But not required for diagnosis • Grading may be performed ○ Banff global assessment based on overall severity – Mild □ Rejection-type infiltrates are generally mild □ Seen in minority of portal triads or perivenular areas □ In isolated central perivenulitis, no confluent necrosis or hepatocyte dropout – Moderate □ Rejection-type infiltrate expands most or all portal triads &/or perivenular areas □ With confluent necrosis/hepatocyte dropout limited to minority of perivenular areas – Severe □ Same as for moderate, with spillover into periportal areas and moderate to severe perivenular inflammation that extends into parenchyma □ Associated with perivenular hepatocyte necrosis ○ Rejection activity index – Semiquantitative scoring of □ Portal inflammation □ Bile duct inflammation and injury □ Endotheliitis – 0-3 points for each category with possible total score of 9

DIFFERENTIAL DIAGNOSIS

– Associated with hepatocyte necrosis &/or dropout • Scattered foci of lobular inflammation or acidophil bodies favor viral hepatitis ○ Beware of confluent necrosis of hepatocytes in severe TCMR • Chronic viral hepatitis more likely to show interface activity and inflammation focused on limiting plate ○ In TCMR, inflammation more centered on portal vein and bile duct • TCMR exhibits more mixed and activated-appearing portal infiltrates and acidophil bodies ○ Late TCMR may mimic chronic hepatitis in containing fewer blastic lymphocytes, less endotheliitis, and more lobular inflammation than early TCMR

Biliary Complications • Biliary obstruction or bile leak may mimic TCMR ○ Both can show signs of bile duct injury, portal inflammation • More neutrophilic infiltrates than TCMR • Portal edema not feature of TCMR • Cholestasis may be present ○ Not specific for biliary complication ○ Incomplete biliary obstruction may not show cholestasis • Chronic biliary obstruction associated with periportal cholate stasis • Acute cholangitis shows collections of neutrophils within bile duct lumina

Recurrent Autoimmune Hepatitis • Recurs in 36-68% of patients transplanted for autoimmune hepatitis • Often numerous plasma cells and marked interface activity

Posttransplant Lymphoproliferative Disorder • Majority of posttransplant lymphoproliferative disorders are B-cell-mediated processes ○ In contrast, TCMR exhibits mostly T lymphocytes

SELECTED REFERENCES 1.

2.

Antibody-Mediated Rejection • Diagnosis based on compatible histology, C4d staining, and detection of donor-specific antibodies at time of biopsy • Can coexist with TCMR ○ Diagnoses are not mutually exclusive

Chronic Viral Hepatitis (Hepatitis B or C) • Newly acquired or recurrent chronic viral hepatitis • Serologic or molecular evidence of viral infection • Distinction from TCMR can be very difficult but clinically important • More stringent criteria used to diagnose TCMR in patients with chronic viral hepatitis ○ Portal inflammation with inflammatory bile duct damage involving ≥ 50% of bile ducts, or ○ Mononuclear perivenular inflammation involving ≥ 50% of terminal hepatic venules 264

3. 4. 5.

Demetris AJ et al: 2016 Comprehensive update of the Banff Working Group on liver allograft pathology: introduction of antibody-mediated rejection. Am J Transplant. 16(10):2816-2835, 2016 Banff Working Group on Liver Allograft Pathology: Importance of liver biopsy findings in immunosuppression management: biopsy monitoring and working criteria for patients with operational tolerance. Liver Transpl. 18(10):1154-70, 2012 Hübscher SG: Transplantation pathology. Semin Liver Dis. 29(1):74-90, 2009 Banff Working Group et al: Liver biopsy interpretation for causes of late liver allograft dysfunction. Hepatology. 44(2):489-501, 2006 Demetris AJ et al: Recurrent hepatitis C in liver allografts: prospective assessment of diagnostic accuracy, identification of pitfalls, and observations about pathogenesis. Am J Surg Pathol. 28(5):658-69, 2004

T-Cell-Mediated Rejection, Liver

Endotheliitis and Mixed Portal Infiltrate (Left) H&E stain shows an area of confluent hepatocyte necrosis ﬉ in severe TCMR. (Right) H&E stain shows a portal tract expanded by a mixed inflammatory cell infiltrate including lymphocytes ﬊ and eosinophils ﬇. There is also evidence of venous endotheliitis ﬈.

Mixed Portal Infiltrate and Endotheliitis

Liver Transplantation

Severe T-Cell-Mediated Rejection With Parenchymal Necrosis

Endotheliitis and Bile Duct Damage in TCell-Mediated Rejection (Left) H&E stain of acute rejection shows a mixed portal inflammatory cell infiltrate composed of activatedappearing lymphocytes, histiocytes, eosinophils, and other inflammatory cells. Endotheliitis is seen in the portal vein ﬇. (Right) H&E stain demonstrates endotheliitis ﬈ of a portal vein branch and mild bile duct damage ſt in an inflamed portal tract.

Central Perivenulitis With Endotheliitis

Central Perivenulitis With Endotheliitis (Left) H&E stain shows perivenular hepatocyte necrosis ﬇ and endotheliitis ﬈ affecting a terminal hepatic vein in a case of severe acute rejection. (Right) H&E stain demonstrates centrilobular perivenulitis with hepatocyte dropout ﬇ and venous endotheliitis ﬊. This finding may be seen in isolation or accompanied by portal features of acute rejection.

265

Liver Transplantation

Antibody-Mediated Rejection, Liver KEY FACTS

TERMINOLOGY • Graft dysfunction mediated by donor-specific antibodies (DSA) directed against donor antigens on endothelium

CLINICAL ISSUES • Hyperacute AMR ○ Severe graft dysfunction over hours to days following revascularization • Acute AMR ○ Usually in 1st several weeks posttransplant ○ Varying degrees of allograft dysfunction • Chronic AMR ○ Usually revealed by protocol biopsies

MICROSCOPIC • Hyperacute AMR ○ Sinusoidal congestion and fibrin deposition ○ Fibrin thrombi in portal and central veins ○ Neutrophilic &/or fibrinoid arteritis

○ Patchy or massive hemorrhagic necrosis • Acute AMR ○ Portal microvascular dilation with endothelial hypertrophy ○ Portal microvasculitis ○ Portal edema, ductular reaction • Chronic AMR (proposed features) ○ At least mild portal &/or perivenular mononuclear cell infiltrate ○ At least moderate portal, periportal, sinusoidal, &/or perivenular fibrosis

ANCILLARY TESTS • DSA testing • C4d immunohistochemistry

TOP DIFFERENTIAL DIAGNOSES • Biliary obstruction • T-cell-mediated rejection

Hyperacute Antibody-Mediated Rejection

Sinusoidal C4d Deposition

Acute Antibody-Mediated Rejection

Portal Microvascular C4d

(Left) A case of hyperacute rejection featuring massive, panacinar hemorrhagic necrosis is shown. A residual portal tract is present ﬊. Note the absence of an inflammatory response. (Right) C4d immunostain shows sinusoidal deposition in a case of severe acute rejection with central perivenulitis ﬊, indicating a role for humoral mechanisms. This is one of the C4d patterns seen in antibody-mediated rejection (AMR).

(Left) An allograft biopsy performed 36 days after transplantation shows features suggestive of biliary obstruction with ductular reaction st; bile duct injury is also noted ﬈. No evidence of biliary obstruction is demonstrated by image studies, however. (Right) Continuous linear C4d immunoreactivity is observed in endothelial cells lining the portal veins ﬉ and capillaries ﬈ in nearly every portal tract present in this biopsy. The patient is also DSA-positive, confirming the diagnosis of acute AMR.

266

Antibody-Mediated Rejection, Liver

Synonyms • Humoral rejection

Definitions • Graft dysfunction mediated by donor-specific antibodies (DSA) directed against donor antigens on endothelium • 3 forms ○ Hyperacute antibody-mediated rejection (AMR) ○ Acute AMR ○ Chronic AMR

ETIOLOGY/PATHOGENESIS Presensitized Donor-Specific Antibodies • Hyperacute or acute AMR ○ ABO blood group isoagglutinins ○ MHC class I antigens

De Novo Donor-Specific Antibodies • Acute and chronic AMR • 8-15% of liver allograft recipients develop de novo DSA ○ MHC class I &/or II antibodies

Mechanisms • • • •

DSA bind to endothelial cells ± activation of complement Initiation of clotting and fibrinolytic cascades Impaired blood flow and tissue damage

CLINICAL ISSUES Presentation • Hyperacute AMR ○ Severe graft dysfunction over hours to days following revascularization – Seen in ABO-incompatible transplants – Rapid rise in serum liver enzyme and bilirubin levels – Thrombocytopenia and hypocomplementemia – Other signs of acute liver failure • Acute AMR ○ Usually in 1st several weeks posttransplant ○ Varying degrees of allograft dysfunction – Rapid allograft failure rarely occurs – Less florid dysfunction with hyperbilirubinemia is more common • Chronic AMR ○ Poorly defined – Typically mild or no liver dysfunction – Usually revealed by protocol biopsies

Treatment • Depends on acuity of injury ○ Plasmapheresis, IVIg, anti-CD20, splenectomy ○ Intensive routine immunosuppression • Retransplantation

Prognosis • Graft failure • Increased incidence of graft complications in recipients who survive early insult

MACROSCOPIC General Features • Hyperacute and severe acute AMR ○ Graft rapidly becomes swollen, cyanotic, and mottled following initial short period of normal reperfusion ○ Thrombosis in large vessels may be seen

Liver Transplantation

○ Biliary complications ○ Hepatic artery complications ○ Conventional acute and chronic rejection

TERMINOLOGY

MICROSCOPIC Histologic Features • Hyperacute AMR ○ Sinusoidal congestion and fibrin deposition ○ Fibrin thrombi in portal and central veins ○ Neutrophilic &/or fibrinoid arteritis ○ Patchy or massive hemorrhagic necrosis ○ Portal edema, hemorrhage, ductular reaction, and cholangitis • Acute AMR ○ Portal microvascular dilation with endothelial hypertrophy ○ Portal microvasculitis (monocytes, eosinophils, and neutrophils) ○ Portal edema, ductular reaction • Chronic AMR (proposed features) ○ At least mild portal &/or perivenular mononuclear cell infiltrate ○ Interface &/or perivenular necroinflammatory activity ○ At least moderate portal, periportal, sinusoidal, &/or perivenular fibrosis

ANCILLARY TESTS Immunohistochemistry • C4d variably positive ○ Sinusoids ○ Portal microvasculature ○ Portal stroma (ABO-incompatible transplants)

DIFFERENTIAL DIAGNOSIS Vascular Thrombosis • Typical Doppler ultrasound and angiographic findings • Zone 3 hepatocyte damage

Biliary Obstruction • Typical cholangiographic findings • Lack of microvasculitis and dilation

T-Cell-Mediated Rejection • Characteristic endotheliitis and bile duct damage • Lack of C4d deposition

SELECTED REFERENCES 1.

2.

Demetris AJ et al: 2016 Comprehensive update of the Banff Working Group on Liver Allograft Pathology: introduction of antibody-mediated rejection. Am J Transplant. 16(10):2816-35, 2016 O'Leary JG et al: Proposed diagnostic criteria for chronic antibody-mediated rejection in liver allografts. Am J Transplant. 16(2):603-14, 2016

267

Liver Transplantation

Antibody-Mediated Rejection, Liver Acute AMR Banff Criteria (2016) Definite for Acute AMR (All 4 Criteria Required)

Comments

(1) Histopathological pattern of injury consistent with acute AMR, including portal microvascular endothelial cell hypertrophy, portal capillary and inlet venule dilation, monocytic, eosinophilic, and neutrophilic portal microvasculitis, portal edema, ductular reaction

h (histopathology)-score (1) Portal microvascular endothelial cell enlargement (portal veins, capillaries, and inlet venules) involving majority of portal tracts with sparse microvasculitis defined as 3-4 marginated &/or intraluminal monocytes, neutrophils, or eosinophils in maximally involved capillary with generally mild dilation (2) Monocytic, eosinophilic, or neutrophilic microvasculitis/capillaritis, defined as at least 5-10 leukocytes marginated &/or intraluminal in maximally involved capillary, prominent portal &/or sinusoidal microvascular endothelial cell enlargement involving majority of portal tracts or sinusoids, with variable but noticeable portal capillary and inlet venule dilation and variable portal edema (3) As above, with marked capillary dilation, marked microvascular inflammation (≥10 marginated &/or intraluminal leukocytes in most severely affected vessels), at least focal microvascular disruption with fibrin deposition, and extravasation of red blood cells into portal stroma &/or space of Disse (subsinusoidal space)

Cholestasis usually present, but variable Active lymphocytic &/or necrotizing arteritis variably present Edema and periportal hepatocyte necrosis are more common/prominent in ABO-incompatible allografts

(2) Positive serum DSA

Often ≥ 5,000 MFI but criteria not established

(3) Diffuse (C4d-score = 3) microvascular C4d deposition on frozen or formalin-fixed, paraffin-embedded tissue in ABO-compatible tissues or portal stromal C4d deposition in ABO-incompatible allografts

C4d-score (formalin-fixed, paraffin-embedded) (0) No C4d deposition in portal microvasculature (1) Minimal (< 10% portal tracts): C4d deposition in > 50% of circumference of portal microvascular endothelia (portal veins and capillaries) (2) Focal (10-50% portal tracts): C4d deposition in > 50% of circumference of portal microvascular endothelia (portal veins and capillaries), usually without extension into periportal sinusoids (3) Diffuse (> 50% portal tracts): C4d deposition in > 50% of circumference of portal microvascular endothelia (portal veins and capillaries), often with extension into inlet venules or periportal sinusoids

(4) Reasonable exclusion of other insults that might cause similar pattern of injury

Most cases will have C4d-score + h-score = 5 or 6

Suspicious for AMR (Both Criteria Required) (1) DSA are positive (2) Nonzero h-score with: C4d-score + h-score = 3 or 4 Indeterminate for AMR (Requires 1 + 2 and 3 or 4) (1) C4d-score + h-score ≥ 2 (2) DSA not available, equivocal, or negative (3) C4d staining not available, equivocal, or negative (4) Coexisting insult might be contributing to injury Formalin-fixed, paraffin-embedded tissues show weaker staining than fresh-frozen tissues, but interpretation of frozen tissues can be more difficult because of background/nonspecific staining and poor preservation of morphology. Sinusoidal staining should be localized to sinusoidal endothelial cells; false-positive staining of connective tissue fibers can occur in livers with subsinusoidal fibrosis. Special stains that help identify capillaries, such as CD31, CD34, &/or PAS, may be needed to help identify involved portal-based capillaries. Fibrin deposition and RBC sludging occur earlier and are more common and prominent in ABO-incompatible allografts. Donor-specific antibodies = DSA; antibody-mediated rejection = AMR; mean fluorescence intensity = MFI.

Chronic AMR Banff Criteria (2016) Probable Chronic AMR (All 4 Criteria Required) (1) Histopathological pattern of injury consistent with chronic AMR; both are required (a) Unexplained and at least mild mononuclear portal &/or perivenular inflammation with interface &/or perivenular necroinflammatory activity (b) At least moderate portal/periportal, sinusoidal &/or perivenular fibrosis (2) Recent (e.g., measured within 3 months of biopsy) circulating HLA DSA in serum samples (3) At least focal C4d(+) (> 10% portal tract microvascular endothelia) (4) Reasonable exclusion of other insults that might cause similar pattern of injury Possible Chronic AMR As above, but C4d staining is minimal or absent

268

Antibody-Mediated Rejection, Liver

Endarteritis (Left) Acute AMR in an allograft biopsy 6 months after transplant is shown. There is portal mononuclear cell infiltrate with microvascular dilation, microvasculitis, and periportal hepatocyte necrosis. DSA are positive in this case. The biopsy also shows features of T-cell-mediated rejection. (Right) Arteritis in hepatic artery branch ﬇ in a patient with positive DSA and C4d deposition is shown. Features of T-cell-mediated rejection are also present.

Acute Antibody-Mediated Rejection

Liver Transplantation

Acute Antibody-Mediated Rejection

Acute Antibody-Mediated Rejection (Left) C4d immunoreactivity in portal microvascular endothelium in a patient with positive DSA 6 months post transplant is shown. (Right) Acute AMR in an allograft biopsy 1 year after transplant is shown. There is dense plasma cell-predominant portal inflammation with microvasculitis ﬈ and C4d immunoreactivity in portal microvascular endothelium ﬊. DSA are positive.

Chronic Antibody-Mediated Rejection

C4d Deposition in Chronic AntibodyMediated Rejection (Left) One month after a 1year posttransplant biopsy that showed acute AMR, a repeat allograft biopsy was performed. Portal inflammation is significantly decreased, but there is portal and periportal fibrosis. (Right) C4d immunoreactivity in portal microvascular endothelium is demonstrated in a biopsy that shows prominent fibrosis in portal tracts.

269

Liver Transplantation

Chronic (Ductopenic) Rejection KEY FACTS ○ PAS stain with diastase digestion or CK7 or CK19 immunohistochemistry may identify bile ducts

TERMINOLOGY • Chronic rejection (CR) • Presents in 2 forms ○ Bile duct loss – Most common finding in allograft biopsy ○ Obliterative (foam cell) arteriopathy – Only seen in large- and medium-sized arteries

TOP DIFFERENTIAL DIAGNOSES • • • •

Ischemic cholangiopathy Recurrent primary biliary cholangitis Recurrent primary sclerosing cholangitis Vanishing bile duct syndrome in drug-induced liver disease

MICROSCOPIC

DIAGNOSTIC CHECKLIST

• Early CR ○ Atypical bile duct epithelium resembling dysplasia ○ Centrizonal perivenular hepatocyte dropout • Late CR ○ Foam cell arteriopathy – Luminal narrowing by subintimal foam cells ○ Loss of interlobular bile ducts • Ductopenic rejection ○ > 50% of portal tracts do not have interlobular bile ducts in 20 portal tracts examined

• Previous episodes of severe or persistent T-cell-mediated (acute cellular) rejection

Bile Duct Loss

Foam Cell Arteriopathy

Bile Duct Loss

Cytokeratin 7

(Left) A portal tract in a case of chronic ductopenic rejection contains a hepatic artery ſt and portal vein ﬇ but no interlobular bile duct. (Right) A medium-sized muscular artery shows intimal foam cell arteriopathy ﬈ in chronic rejection.

(Left) PAS stain with diastase digestion in a case of chronic rejection shows the presence of hepatic arterioles ﬈ and portal vein ﬊ but no interlobular bile ducts. (Right) Immunohistochemical stain for CK7 shows cells in the hepatic progenitor cell compartment ﬈ in the periportal region but no interlobular bile ducts.

270

Chronic (Ductopenic) Rejection

Abbreviations • Chronic rejection (CR)

DIFFERENTIAL DIAGNOSIS

Synonyms

Ischemic Cholangiopathy

• Ductopenic rejection

• Imaging studies may help • CR usually lacks secondary biliary changes such as ○ Ductular reaction ○ Copper staining

Definitions • Presents in 2 forms ○ Obliterative (foam cell) arteriopathy – Only seen in large- and medium-sized arteries ○ Bile duct loss – Most common finding in allograft biopsy

ETIOLOGY/PATHOGENESIS Immune-Mediated Damage to Allograft • May evolve from severe or repeated T-cell-mediated (acute cellular) rejection ○ Result in potentially irreversible damage to interlobular bile ducts &/or endothelium of veins and arteries

CLINICAL ISSUES Presentation • Progressive jaundice and elevated cholestatic enzymes • Onset usually during 1st year post transplant • Occurs later than T-cell-mediated rejection

Treatment • Early CR may respond to potent immunosuppressants such as tacrolimus, OKT3, mycophenolate, or rapamycin

Prognosis • Usually unresponsive to immunosuppression • Retransplantation often needed

MICROSCOPIC Histologic Features • Early CR ○ Lymphocytic cholangitis ○ Atypical bile duct epithelium; may resemble dysplasia ○ Perivenular hepatocyte dropout • Late CR ○ Loss of interlobular bile ducts ○ Loss of hepatic arteries ○ Foam cell arteriopathy with luminal narrowing by subintimal foam cells ○ Portal inflammation typically decreases over time ○ Perivenular necrosis or hepatocyte dropout common • Ductopenic rejection ○ > 50% of portal tracts do not have interlobular bile ducts ○ Need > 20 portal tracts in biopsy for assessment – Serial biopsies may be needed if sample contains < 20 portal tracts ○ PAS stain with diastase digestion or CK7 or CK19 immunohistochemical stains may help identify bile ducts when ductopenia suspected ○ Cholestasis often prominent ○ Ductular reaction, periportal fibrous expansion usually absent

Liver Transplantation

○ Marked perivenular fibrosis with bridging may develop in late CR

TERMINOLOGY

Recurrent Primary Biliary Cholangitis • Florid duct lesions, portal inflammation, and bile ductular reaction • Positive antimitochondrial antibody (AMA)

Recurrent Primary Sclerosing Cholangitis • Characteristic ERCP findings • Duct sclerosis, periductal fibrosis • Portal inflammation with bile ductular reaction

Drug-Induced Vanishing Bile Duct Syndrome • History of medications known to cause ductopenia ○ Amoxicillin/clavulanate (Augmentin) ○ Chlorpromazine ○ Phenytoin

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Previous episodes of severe or persistent T-cell-mediated (acute cellular) rejection

Pathologic Interpretation Pearls • Centrilobular necrosis &/or cholestasis in repeated biopsy specimens ○ Considered warning sign of possible CR • Foam cell arteriopathy ○ Typically occurs in medium- or large-sized arteries  ○ Seldom seen in unaffected allograft biopsy specimens

SELECTED REFERENCES 1.

2. 3.

4. 5.

6.

7.

Demetris AJ et al: 2016 comprehensive update of the Banff Working Group on Liver Allograft Pathology: introduction of antibody-mediated rejection. Am J Transplant. ePub, 2016 Lefkowitch JH: Diagnostic issues in liver transplantation pathology. Clin Liver Dis. 6(2):555-70, ix, 2002 Demetris A et al: Update of the International Banff Schema for Liver Allograft Rejection: working recommendations for the histopathologic staging and reporting of chronic rejection. An International Panel. Hepatology. 31(3):792-9, 2000 Jones KD et al: Interpretation of biopsy findings in the transplant liver. Semin Diagn Pathol. 15(4):306-17, 1998 Noack KB et al: Severe ductopenic rejection with features of vanishing bile duct syndrome: clinical, biochemical, and histologic evidence for spontaneous resolution. Transplant Proc. 23(1 Pt 2):1448-51, 1991 van Hoek B et al: Recurrence of ductopenic rejection in liver allografts after retransplantation for vanishing bile duct syndrome. Transplant Proc. 23(1 Pt 2):1442-3, 1991 Ludwig J et al: Persistent centrilobular necroses in hepatic allografts. Hum Pathol. 21(6):656-61, 1990

271

Liver Transplantation

Recurrent Hepatitis B Virus KEY FACTS ○ Chronic hepatitis pattern of injury with portal inflammation ○ Interface activity ○ Ground-glass hepatocytes may be present

CLINICAL ISSUES • Current risk of recurrent infection is < 10% • Rare progression to cirrhosis in donor liver with institution of antiviral therapy • Transplant due to fulminant hepatitis B virus (HBV) has lower recurrence rate compared to cirrhosis due to chronic HBV hepatitis • High viral load at time of transplantation is risk factor for recurrence • Excellent prognosis with current therapy

MICROSCOPIC • 2-6 weeks: Histologically unremarkable • 6 weeks to 6 months ○ Mild acute lobular hepatitis with scattered acidophil bodies ○ Lobular inflammation and ○ Kupffer cell aggregates • > 6 months

ANCILLARY TESTS • Antihepatitis B surface antigen immunohistochemical stain demonstrates cytoplasmic reactivity within virally infected cells

TOP DIFFERENTIAL DIAGNOSES • T-cell-mediated rejection ○ Portal inflammation in T-cell-mediated rejection mixed with – Prominent eosinophils – Bile duct damage – Endotheliitis ○ Usually no significant lobular inflammation or necrosis except around central veins

Early Recurrent HBV Hepatitis

Recurrent Chronic HBV Hepatitis

Ground-Glass Hepatocytes

IHC for Hepatitis B Surface Antigen

(Left) This case of active early recurrent hepatitis B virus (HBV) hepatitis is characterized by prominent lobular inflammation ſt with numerous apoptotic hepatocytes. The portal tracts ﬇ are relatively unremarkable. (Right) In this case, there are well-developed features of recurrent chronic HBV hepatitis, characterized by fairly dense portal inflammation ﬈ and interface activity.

(Left) Finely granular cytoplasm with peripheral clearing is characteristic of ground-glass hepatocytes in hepatitis B infection. (Right) Immunohistochemistry for hepatitis B surface antigen can be used to highlight virally infected cells.

272

Recurrent Hepatitis B Virus

Abbreviations • Hepatitis B virus (HBV) • Hepatitis B surface antigen (HBsAg) • Hepatitis B core antigen (HBcAg)

ETIOLOGY/PATHOGENESIS Infectious Agents • HBV ○ DNA virus with circular genome

CLINICAL ISSUES Epidemiology • Incidence ○ In 1980s and 1990s, 50% risk of recurrent infection ○ < 10% current risk of recurrent infection ○ Transplant due to fulminant HBV has lower recurrence rate compared to cirrhosis due to chronic HBV hepatitis ○ High viral load at time of transplantation is risk factor for recurrence ○ Other risk factors for recurrence include hepatocellular carcinoma, precore and pre-S mutations, HBV genotypes C and D, and high immunosuppression

Presentation • Often asymptomatic with elevations in liver function tests found on routine laboratory evaluation

Treatment • Drugs ○ Pretransplant treatment with antiviral therapy (tenofovir, entecavir, lamivudine, adefovir, etc.) ± antiHBs immunoglobulin ○ Posttransplant prophylaxis with antiviral therapy to prevent recurrence

Prognosis • Excellent prognosis with current therapy

MICROSCOPIC Histologic Features • 2-6 weeks post transplantation ○ Histologically unremarkable ○ Rare hepatocytes with cytoplasmic and nuclear expression of HBcAg • 6 weeks to 6 months post transplantation ○ Mild acute lobular hepatitis with scattered acidophil bodies, lobular inflammation, and Kupffer cell aggregates; minimal portal inflammation ○ Rare ground-glass hepatocytes in acute phase • > 6 months post transplantation ○ Chronic hepatitis pattern of injury with portal inflammation, interface and lobular activity, and progressive fibrosis ○ Ground-glass hepatocytes may be present ○ Grade and stage may be used, but – Current systems not validated in posttransplant setting

• Rare diffuse hepatocyte swelling and steatosis in massive viral replication (steatoviral hepatitis B) • Rare fibrosing cholestatic hepatitis with prominent hepatocyte ballooning, ductular reaction, cholestasis, and rapid fibrosis

ANCILLARY TESTS Immunohistochemistry

Liver Transplantation

TERMINOLOGY

• Anti-HBsAg demonstrates cytoplasmic reactivity within virally infected cells • Anti-HBcAg (nuclear and cytoplasmic stain) can highlight cells with active viral replication

DIFFERENTIAL DIAGNOSIS T-Cell-Mediated Rejection • Portal inflammation in T-cell-mediated rejection mixed with prominent eosinophils, bile duct damage, and endotheliitis • Usually no significant lobular inflammation or necrosis except around central veins

Other Forms of Chronic Hepatitis • Serologic testing necessary to exclude other chronic viral hepatitis or de novo autoimmune hepatitis (plasma cell hepatitis)

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Usually causes mild graft dysfunction; however, fibrosing cholestatic hepatitis form results in rapid fibrosis and graft failure

Pathologic Interpretation Pearls • Time after transplant is critical in allograft biopsy evaluation for recurrent HBV hepatitis • Serologic evidence of active HBV replication also helpful • Ground-glass hepatocytes are characteristic of chronic HBV infection but can rarely be seen with certain medications

SELECTED REFERENCES 1.

Terrault NA et al: AASLD guidelines for treatment of chronic hepatitis B. Hepatology. 63(1):261-83, 2016 2. Yataco M et al: Long term survival and complications after liver transplantation in patients with chronic hepatitis B. Ann Transplant. 15(2):2734, 2010 3. Beckebaum S et al: Predictive factors of outcome in patients transplanted for hepatitis B. Transplantation. 87(6):872-81, 2009 4. Mani H et al: Liver biopsy findings in chronic hepatitis B. Hepatology. 49(5 Suppl):S61-71, 2009 5. Papatheodoridis GV et al: Current management of hepatitis B virus infection before and after liver transplantation. Liver Int. 29(9):1294-305, 2009 6. Faria LC et al: Hepatocellular carcinoma is associated with an increased risk of hepatitis B virus recurrence after liver transplantation. Gastroenterology. 134(7):1890-9; quiz 2155, 2008 7. Gane EJ et al: Lamivudine plus low-dose hepatitis B immunoglobulin to prevent recurrent hepatitis B following liver transplantation. Gastroenterology. 132(3):931-7, 2007 8. Thung SN: Histologic findings in recurrent HBV. Liver Transpl. 12(11 Suppl 2):S50-3, 2006 9. Phillips MJ et al: Post-transplant recurrent hepatitis B viral liver disease. viralburden, steatoviral, and fibroviral hepatitis B. Am J Pathol. 140(6):1295-308, 1992 10. Demetris AJ et al: Recurrent hepatitis B in liver allograft recipients. Differentiation between viral hepatitis B and rejection. Am J Pathol. 125(1):161-72, 1986

273

Liver Transplantation

Recurrent Hepatitis C Virus KEY FACTS

CLINICAL ISSUES • Pretransplant therapy with direct-acting antivirals has significantly decreased incidence of recurrent HCV hepatitis ○ In untreated patients, reinfection of allograft after transplantation nearly universal – Variable prognosis depends on many factors including donor/recipient age, donor steatosis, fibrosis at 12 months, episodes of T-cell-mediated rejection, immunosuppression, and viral genotype

MICROSCOPIC • Typical HCV recurrence ○ 2-6 months: Features of mild acute hepatitis ○ > 6 months: Establishment of chronic hepatitis with portal inflammation and interface and lobular activity • HCV hepatitis with T-cell-mediated rejection ○ Most cases represent recurrent HCV with only mild features of rejection

○ Treatment should be considered in cases with at least moderate rejection • Plasma cell hepatitis in HCV hepatitis ○ Inflammation with > 30% plasma cells ○ Likely represents variant of T-cell-mediated rejection or de-novo autoimmune hepatitis • Treated HCV hepatitis ○ Histologic features of HCV hepatitis (necroinflammatory activity) often persist despite achievement of sustained virologic response ○ Progressive fibrosis occurs in some patients

TOP DIFFERENTIAL DIAGNOSES • T-cell-mediated rejection ○ > 50% of bile ducts &/or central veins should be affected ○ Perivenular inflammation is not feature of recurrent HCV hepatitis

Early Recurrent HCV Hepatitis

Lymphocytic Portal Infiltrate

Steatosis in HCV, Genotype 3

Periportal Fibrosis (Trichrome Stain)

(Left) Early recurrent hepatitis C virus (HCV) hepatitis is characterized by scattered lobular inflammation, Kupffer cell aggregates ﬊, and acidophil bodies ﬈. Mild steatosis may be seen. Portal tracts are not inflamed. (Right) Chronic HCV hepatitis is characterized by a lymphocytic portal inflammatory infiltrate with interface activity ﬊. Bile ducts ﬈ are usually unremarkable.

(Left) Patients transplanted for hepatitis C-induced cirrhosis due to genotype 3 HCV often have prominent steatosis before the development of chronic hepatitis. Note that the portal tract ﬈ is fairly unremarkable. (Right) Protocol liver biopsy taken at 1 year post transplant demonstrates periportal fibrosis. This patient went on to develop cirrhosis at year 3 despite treatment with interferon and ribavirin.

274

Recurrent Hepatitis C Virus

Abbreviations • Hepatitis C virus (HCV) • Direct-acting antivirals (DAAs) • Sustained virologic response (SVR)

Synonyms • T-cell-mediated rejection (TCMR), formerly acute cellular rejection (ACR)

Definitions • SVR ○ No detectable HCV RNA in serum 12-24 weeks after completion of antiviral therapy

– Degree of necroinflammatory activity – Amount of fibrosis at 1 year post transplant – Severity and number of episodes of TCMR – Hepatic iron – Cholestasis/ballooning • Recurrent HCV likely more aggressive in posttransplant setting ○ 20-30% of untreated patients have advanced fibrosis (bridging or cirrhosis) 5 years post transplant (rapid progressors) ○ 50-70% of untreated patients will be cirrhotic at 10 years post transplant

Liver Transplantation

TERMINOLOGY

MICROSCOPIC Histologic Features

ETIOLOGY/PATHOGENESIS Infectious Agents • HCV ○ Single-stranded RNA virus

CLINICAL ISSUES Presentation • Reinfection of allograft after transplantation is nearly universal in untreated patients ○ Within 1st year, most patients are asymptomatic ○ Elevated aminotransferase levels – Usually 1st indication of recurrent infection

Natural History • In most centers, protocol liver biopsies performed to monitor HCV recurrence ○ Standard protocol includes biopsy at 6 months, 1 year, and every year thereafter ○ Often additional biopsies performed for unexpected elevations in liver function tests • Fibrosis quite variable between patients ○ Treatment algorithms depend on many factors

Treatment • Drugs ○ Most patients now treated with DAAs pretransplant – Posttransplant treatment with DAAs also increasingly common ○ Ribavirin and interferon formerly used to treat HCV hepatitis in pre- and posttransplant setting

Prognosis • SVR achieved in > 90% of patients treated with DAAs • If untreated/failed treatment, prognosis varies and depends on many factors ○ Clinical factors – Donor and recipient age – Pretransplant HCV RNA levels – HCV genotype – Type and level of immunosuppression – Preservation injury – Superimposed metabolic syndrome – Cytomegalovirus infection ○ Pathologic factors – Donor steatosis

• Typical HCV recurrence ○ 2-6 months (graft reinfection) – Features of acute hepatitis characterized by □ Sparse or minimal portal inflammation □ Mild lobular disarray □ Scattered acidophil bodies □ Kupffer cell aggregates □ Mild lobular inflammation – Mild steatosis may also be seen – HCV RNA levels can be quite high in this phase – Evidence of hepatitis C recurrence can occur as early as 1 week post transplantation with high pretransplant viral load ○ ~ 6 months (chronic graft injury) – Establishment of chronic hepatitis characterized by portal inflammation (± lymphoid aggregates) with interface and lobular activity – Focal bile duct infiltration by lymphocytes may be seen (Poulsen lesion) □ Not to be confused with TCMR – Mild steatosis may also be seen – In chronic phase of HCV hepatitis, grade and stage may be cautiously given □ These systems not validated in posttransplant setting – HCV RNA levels decrease during progression to chronic HCV hepatitis • HCV genotype 3 recurrence ○ May have prominent steatosis and even features of steatohepatitis before development of chronic hepatitis • Fibrosing cholestatic HCV ○ Rapid progression to severe fibrosis and graft failure ○ High viral HCV RNA ○ Histologically mimics extrahepatic biliary obstruction • HCV with TCMR ○ Very difficult to diagnose HCV with superimposed TCMR ○ Most cases represent recurrent HCV with only mild features of rejection, such as – Mild bile duct injury – Focal endotheliitis – Classic cases of chronic HCV hepatitis can have focal lymphocytic infiltration of bile ducts □ Should not be interpreted as feature of rejection ○ Increased immunosuppression should be considered when features of TCMR are at least moderate 275

Liver Transplantation

Recurrent Hepatitis C Virus Histologic Features of Recurrent Hepatitis C Virus and T-Cell-Mediated Rejection Feature

Hepatitis C Virus

T-Cell-Mediated Rejection

Portal inflammation

Mononuclear: Lymphocytes, lymphoid aggregates

Mixed: Eosinophils, blastic lymphocytes, neutrophils

Bile duct injury

Only rare lymphocytes infiltrating bile ducts

Mixed infiltrate infiltrating bile ducts

Portal vein endotheliitis

Very rare

Common

Interface activity

Common

Rare

Parenchymal acidophil bodies

Common

Rare except in moderate to severe cases

Central vein inflammation

Rare

May be present in combination with portal features but also as centrilobular variant of rejection

Steatosis

Common, especially genotype 3

Rare

Fibrosis

Present/common

Usually no fibrosis

○ Some cases have portal-based features of HCV and central perivenulitis (centrilobular TCMR) – > 50% of central veins must demonstrate endotheliitis in order to consider rejection if both coexist • Plasma cell hepatitis in HCV infection ○ Defined as hepatitis with plasma cells comprising ≥ 30% of inflammatory cell infiltrate ○ Cases show portal-based plasma cell-rich infiltrates with prominent interface activity ○ Often has central perivenulitis with hepatocyte necrosis and prominent plasma cells ○ Some arise in patients receiving therapy for HCV (pegylated interferon and ribavirin) and lacking serum HCV RNA – Suggests hepatitis may be triggered by HCV treatment ○ Other cases lack an association with therapy for HCV ○ Associated with autoantibodies (ANA, ASMA, etc.), elevated immunoglobulins, and suboptimal immunosuppression ○ Treatment with increased immunosuppression (with avoidance of steroids) results in improved outcomes • Treated HCV hepatitis ○ Histologic features of HCV hepatitis often persist despite achievement of SVR with DAAs – Necroinflammatory activity common – Progressive fibrosis occurs in some patients

DIFFERENTIAL DIAGNOSIS T-Cell-Mediated Rejection • Differentiating recurrent hepatitis C from TCMR is major challenge in liver allograft pathology • Features that favor TCMR ○ Portal inflammation mixed with blastic lymphocytes and eosinophils ○ Prominent portal vein endotheliitis and bile duct infiltration by inflammatory cells ○ Perivenular inflammation with hepatocyte injury is not feature of hepatitis C infection • Criteria for rejection should require more extensive tissue injury ○ 50% of bile ducts should be injured, or > 50% of central veins should have inflammation and hepatocyte dropout for mild rejection

276

○ More liver injury also required for moderate and severe rejection • TCMR usually associated with low viral RNA

Biliary Obstruction • Characterized by portal edema and bile ductular reaction with associated neutrophils • Some have more mixed inflammatory cell infiltrate with occasional portal vein endotheliitis and bile duct injury mimicking TCMR

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Degree of necroinflammatory activity and fibrosis can help predict severity of HCV recurrence • Liver biopsy plays critical role in distinguishing HCV recurrence from other causes of graft dysfunction • Pathologists should be cautious when diagnosing TCMR in setting of HCV infection ○ Increased immunosuppression can result in graft failure due to severe HCV recurrence

Pathologic Interpretation Pearls • Early HCV recurrence lacks portal-based inflammation ○ Usually demonstrates only scattered acidophil bodies • In setting of high HCV viral RNA, be particularly cautious in diagnosing TCMR

SELECTED REFERENCES 1.

2. 3.

4. 5. 6. 7.

8.

Whitcomb E et al: Biopsy specimens from allograft liver contain histologic features of hepatitis C virus infection after virus eradication. Clin Gastroenterol Hepatol. ePub, 2017 Rahimi RS et al: Post-liver transplant hepatitis C therapy. Curr Treat Options Gastroenterol. 13(2):249-58, 2015 Pearlman BL et al: Sustained virologic response to antiviral therapy for chronic hepatitis C virus infection: a cure and so much more. Clin Infect Dis. 52(7):889-900, 2011 Demetris AJ: Evolution of hepatitis C virus in liver allografts. Liver Transpl. 15 Suppl 2:S35-41, 2009 Demetris AJ et al: Plasma cell hepatitis in liver allografts: Variant of rejection or autoimmune hepatitis? Liver Transpl. 14(6):750-5, 2008 Banff Working Group et al: Liver biopsy interpretation for causes of late liver allograft dysfunction. Hepatology. 44(2):489-501, 2006 Demetris AJ et al: Recurrent hepatitis C in liver allografts: prospective assessment of diagnostic accuracy, identification of pitfalls, and observations about pathogenesis. Am J Surg Pathol. 28(5):658-69, 2004 Greenson JK et al: Histologic progression of recurrent hepatitis C in liver transplant allografts. Am J Surg Pathol. 20(6):731-8, 1996

Recurrent Hepatitis C Virus

Superimposed T-Cell-Mediated Rejection (Left) Biopsy taken at 15 months post transplant due to elevations in liver function tests demonstrates portal tract with features of mild recurrent HCV hepatitis (lymphocytic portal inflammation with mild interface activity). (Right) However, the same biopsy also had superimposed features of T-cell-mediated rejection, as 100% of central veins had perivenular inflammation with hepatocyte injury. The infiltrate was mixed with lymphocytes, eosinophils, and scattered plasma cells.

Treated HCV Hepatitis

Liver Transplantation

Mild Recurrent HCV Hepatitis

Treated HCV Hepatitis (Left) Liver biopsy from a patient who achieved a sustained viral response after direct-acting antiviral therapy for HCV infection. There is a persistent mild portal lymphoplasmacytic infiltrate. (Right) Another mildly inflamed portal tract from a patient with treated HCV hepatitis is shown. There was no fibrosis in this case.

Plasma Cell Hepatitis

Plasma Cell Hepatitis (Left) This liver biopsy was taken from a patient who was recently treated for hepatitis C after transplantation. Despite a low viral load, the patient's AST and ALT levels started to rise. Almost all of the central veins have an inflammatory cell infiltrate rich in plasma cells (plasma cell hepatitis) with associated hepatocyte injury ſt. (Right) Portal tracts from the same biopsy also contain a plasma cell-rich infiltrate. There is mild interface activity as well ſt.

277

Liver Transplantation

Fibrosing Cholestatic HBV or HCV Hepatitis KEY FACTS

ETIOLOGY/PATHOGENESIS • Thought to be due to direct viral cytopathic effect resulting from impaired immune response

CLINICAL ISSUES • Usually occurs within 1st year post transplantation • Can occur in setting of increased immunosuppression due to prior episodes of T-cell-mediated rejection • Laboratory values ○ Cholestatic chemistry profile, often with bilirubin > 5 mg/dL ○ AST/ALT levels > 3-4x upper limit of normal ○ High viral load • Poor prognosis: > 90% graft failure within 1 year

MICROSCOPIC • Marked canalicular and hepatocellular cholestasis with ductular reaction • Hepatocellular swelling and ballooning, any location

• Portal fibrosis with delicate fibrous septa extending into sinusoidal spaces • Hepatitis aggressiveness scoring system may be useful in predicting aggressiveness of hepatitis recurrence

TOP DIFFERENTIAL DIAGNOSES • Bile duct obstruction ○ More portal edema ○ Less cholestasis and periportal sinusoidal fibrosis ○ Must exclude clinically • Severe recurrent hepatitis C virus (HCV) infection ○ Cholestasis usually not prominent ○ Delicate perisinusoidal periportal fibrosis unusual in classic recurrent HCV ○ Fibrosing cholestatic hepatitis cannot be reliably distinguished from active recurrent hepatitis with superimposed biliary obstruction

Hepatocellular Ballooning and Cholestasis

Ductular Reaction

Portal Inflammation and Ductular Reaction

Periportal Sinusoidal Fibrosis

(Left) This biopsy shows common findings in fibrosing cholestatic hepatitis (FCH), which include hepatocellular ballooning ﬊ and cholestasis ﬈. (Right) This portal tract is not inflamed; however, there is a marked ductular reaction at the periphery of the portal tract. Note the presence of hepatocellular ballooning/swelling ﬈.

(Left) This portal tract from a patient with FCH due to hepatitis C virus demonstrates portal inflammation with interface activity ſt and a ductular reaction ﬇. (Right) Trichrome stain shows that the fibrosis in FCH is quite unique with periportal sinusoidal fibrosis.

278

Fibrosing Cholestatic HBV or HCV Hepatitis

Abbreviations • Fibrosing cholestatic hepatitis (FCH)

Synonyms • Fibrosing cytolytic liver failure • Fibroviral hepatitis

Definitions • Severe viral-associated cholestatic syndrome associated with rapid graft failure

ETIOLOGY/PATHOGENESIS Infectious Agents • Hepatitis B virus ○ Uncommon now due to appropriate prophylaxis • Hepatitis C virus (HCV)

Mechanisms and Associations • Thought to be due to direct viral cytopathic effect resulting from impaired immune response • Associated with older donor age • Associated with enhanced divergence of viral quasispecies

CLINICAL ISSUES Epidemiology • Relatively uncommon: Frequency of < 2% in patients transplanted for hepatitis C • Usually occurs within 1st year post transplantation

Presentation • Can occur in setting of increased immunosuppression due to prior episodes of T-cell-mediated rejection • Jaundice, encephalopathy, coagulopathy • Laboratory findings ○ Cholestatic chemistry profile, often with bilirubin > 5 mg/dL – No difference in alkaline phosphatase compared to other patients with recurrent HCV ○ AST/ALT levels > 3-4x upper limit of normal ○ High viral load

Treatment • Reduced immunosuppression • Antiviral therapy • Retransplantation ○ Usually not option due to poor outcomes

Prognosis • Poor: > 90% graft failure within 1 year

• Hepatitis aggressiveness score proposed to better classify severe HCV recurrence ○ Features – Ductular reaction mimicking biliary obstruction – Prominent hepatocyte ballooning/swelling present in majority of sample with lobular disarray – Cholestasis (including at least focal canalicular cholestasis) of any degree – Periportal sinusoidal fibrosis ○ Scoring – 0 of 4 features: Nonaggressive hepatitis – 1-2 of 4 features: Aggressive HCV hepatitis – 3-4 of 4 features: FCH ○ Hepatitis aggressiveness score correlated with survival and graft loss

DIFFERENTIAL DIAGNOSIS Bile Duct Obstruction • Has more portal edema and less cholestasis and periportal sinusoidal fibrosis • Before diagnosis of FCH is considered, biliary obstruction must be excluded clinically

Severe Recurrent Hepatitis C Virus Infection • May have some degree of ductular reaction and lobular disarray • Cholestasis usually not prominent • Delicate periportal sinusoidal fibrosis is unusual in classic recurrent HCV • FCH cannot be reliably distinguished from active recurrent hepatitis with superimposed biliary obstruction

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Diagnosis of FCH strongly correlated to graft and patient survival • Distinction from bile duct obstruction is critical, but often requires clinical and radiologic correlation

Pathologic Interpretation Pearls • Hepatocellular and canalicular cholestasis may be early feature of FCH and be missed • FCH rarely diagnosed > 1 year post transplantation • Overdiagnosis of T-cell-mediated rejection in setting of recurrent HCV may result in FCH

SELECTED REFERENCES 1.

2.

MICROSCOPIC Histologic Features • • • •

Marked canalicular and hepatocellular cholestasis Hepatocellular swelling and ballooning, any location Ductular reaction Portal fibrosis with delicate fibrous septa extending into sinusoidal spaces • Some cases can have only mild portal inflammation

Liver Transplantation

TERMINOLOGY

3. 4. 5. 6. 7.

Moreira RK et al: The Hepatitis Aggressiveness Score (HAS): a novel classification system for post-liver transplantation recurrent hepatitis C. Am J Surg Pathol. 37(1):104-13, 2013 Verna EC et al: Cholestatic hepatitis C following liver transplantation: an outcome-based histological definition, clinical predictors, and prognosis. Liver Transpl. 19(1):78-88, 2013 Gane EJ: The natural history of recurrent hepatitis C and what influences this. Liver Transpl. 14 Suppl 2:S36-44, 2008 Xiao SY et al: Fibrosing cholestatic hepatitis: clinicopathologic spectrum, diagnosis and pathogenesis. Int J Clin Exp Pathol. 1(5):396-402, 2008 Dixon LR et al: Early histologic changes in fibrosing cholestatic hepatitis C. Liver Transpl. 13(2):219-26, 2007 Taga SA et al: Cholestatic hepatitis C in liver allografts. Liver Transpl Surg. 4(4):304-10, 1998 Davies SE et al: Hepatic histological findings after transplantation for chronic hepatitis B virus infection, including a unique pattern of fibrosing cholestatic hepatitis. Hepatology. 13(1):150-7, 1991

279

Liver Transplantation

Recurrent Autoimmune Hepatitis KEY FACTS

CLINICAL ISSUES

TOP DIFFERENTIAL DIAGNOSES

• 12-46% of patients transplanted for autoimmune hepatitis (AIH) have recurrent disease • Most present within 1-6 years post transplant • Graft failure due to recurrent AIH is infrequent (~ 6%) • Histologic recurrence may precede clinical and biochemical recurrence

• Plasma cell hepatitis (de novo AIH) ○ Histologically similar to AIH ○ Considered form of rejection ○ Occurs in patients transplanted for conditions other than AIH, particularly HCV ○ Recognition critical due to high risk of graft loss &/or death • Chronic viral hepatitis ○ Dense portal inflammation with interface activity ○ Plasma cells less prominent in HBV and HCV ○ Interface activity usually less prominent in HBV and HCV • T-cell-mediated rejection ○ Mixed infiltrate ○ Prominent bile duct damage ○ Prominent endotheliitis

MICROSCOPIC • Portal tracts ○ Dense mononuclear cell infiltrates ○ Abundant plasma cells ○ Prominent interface activity • Central veins ○ Often central vein inflammation with prominent plasma cells • Parenchyma ○ Clusters of inflammatory cells including plasma cells

Interface Activity and Acidophil Bodies

Plasmacytic Infiltrate

Parenchymal Collapse

Parenchymal Collapse (Trichrome Stain)

(Left) This example of recurrent chronic autoimmune hepatitis demonstrates brisk interface activity and numerous acidophil bodies ſt. (Right) The portal tract infiltrate in this case of recurrent autoimmune hepatitis is composed predominantly of plasma cells.

(Left) Severe recurrent autoimmune hepatitis with panlobular necrosis in this case shows a cluster of plasma cells within the area of parenchymal collapse ſt. (Right) A trichrome stain is useful in highlighting collapse rather than fibrosis.

280

Recurrent Autoimmune Hepatitis

Abbreviations • Autoimmune hepatitis (AIH) • T-cell-mediated rejection (TCMR), formerly acute cellular rejection (ACR)

○ Clusters of inflammatory cells, including plasma cells ○ Acidophil bodies ○ Parenchymal collapse/necrosis • Progressive fibrosis if untreated • No grading/staging system has been validated in posttransplant setting ○ Grade and stage may be cautiously given

CLINICAL ISSUES DIFFERENTIAL DIAGNOSIS

Epidemiology • 12-46% of patients transplanted for AIH have recurrent disease ○ Recurrence likely higher in pediatric population • Risk factors for recurrence ○ Suboptimal immunosuppression ○ Type 1 AIH ○ High IgG levels pretransplant ○ AIH in native liver with marked necroinflammatory activity – However, fulminant hepatic failure due to AIH may have lower risk of recurrence

Presentation • Most present within 1-6 years post transplant • Histologic recurrence may precede clinical and biochemical recurrence • Diagnosis requires most of following ○ Liver transplant for AIH ○ Elevated aminotransferases ○ Elevated immunoglobulins ○ Autoantibodies: Antinuclear antibody (ANA), antismooth muscle antibody (ASMA), &/or anti-liver/kidney microsomal (LKM) antibody – Presence of autoantibodies is less important in posttransplant setting ○ Clinical and histologic response to increased immunosuppression ○ Exclusion of other causes of graft dysfunction ○ Interface hepatitis on liver biopsy

Treatment • Drugs ○ Increased immunosuppression, including corticosteroids and azathioprine

Prognosis • Infrequent cause of graft failure (~ 6%) • Recurrent AIH increases risk of acute and chronic allograft rejection

Liver Transplantation

TERMINOLOGY

Plasma Cell Hepatitis (De Novo AIH) • Histologically similar to AIH ○ Interface hepatitis with prominent plasmacytic infiltrate • Considered form of rejection • Occurs in patients transplanted for conditions other than AIH, particularly pediatric patients and adults with HCV treated with interferon-ribavirin • High autoantibody titers common • Recognition critical due to high risk of graft loss &/or death

Chronic Viral Hepatitis • • • •

Dense portal inflammation with interface activity Plasma cells less prominent in HBV and HCV Interface activity usually less prominent in HBV and HCV Clinically, HBV and HCV will be apparent serologically

T-Cell-Mediated Rejection • Mixed infiltrate of blastic lymphocytes, macrophages, eosinophils, neutrophils • Prominent bile duct damage • Prominent endotheliitis • Centrilobular necrosis can occur with mainly lymphocytic infiltrate • Minimal lobular inflammation • No fibrosis

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Liver biopsy is often essential for diagnosis • Plasma cell-rich infiltrate helps distinguish recurrent AIH from other causes of graft dysfunction • Histologic recurrence can occur before graft dysfunction becomes clinically apparent • Recurrence often occurs > 1 year after transplantation

SELECTED REFERENCES 1. 2.

MICROSCOPIC Histologic Features

3.

• Portal tracts ○ Dense mononuclear cell infiltrates with abundant plasma cells ○ Prominent interface activity ○ May have apparent endotheliitis ○ No significant bile duct damage • Central veins often inflamed with prominent plasmacytic infiltrate • Parenchyma

4.

5.

6. 7.

Kerkar N et al: 'De novo' and 'recurrent' autoimmune hepatitis after liver transplantation: a comprehensive review. J Autoimmun. 66:17-24, 2016 Fiel MI et al: Plasma cell hepatitis (de-novo autoimmune hepatitis) developing post liver transplantation. Curr Opin Organ Transplant. 17(3):287-92, 2012 Liberal R et al: Autoimmune hepatitis after liver transplantation. Clin Gastroenterol Hepatol. 10(4):346-53, 2012 Schreuder TC et al: Autoimmune liver diseases and recurrence after orthotopic liver transplantation: what have we learned so far? Transpl Int. 22(2):144-52, 2009 Tripathi D et al: Autoimmune hepatitis and liver transplantation: indications, results, and management of recurrent disease. Semin Liver Dis. 29(3):286-96, 2009 Banff Working Group et al: Liver biopsy interpretation for causes of late liver allograft dysfunction. Hepatology. 44(2):489-501, 2006 Duclos-Vallée JC et al: A 10 year follow up study of patients transplanted for autoimmune hepatitis: histological recurrence precedes clinical and biochemical recurrence. Gut. 52(6):893-7, 2003

281

Liver Transplantation

Recurrent Primary Biliary Cholangitis KEY FACTS

CLINICAL ISSUES • Prevalence increases progressively post transplantation ○ 30-50% recurrence within 10 years • Most found to have recurrent disease on protocol liver biopsies • Positive antimitochondrial antibody less useful for diagnosis of recurrent primary biliary cholangitis (PBC) • Risk factors for recurrence ○ Male donor ○ Reduced immunosuppression ○ Living donor transplantation • Excellent prognosis; rarely progresses to cirrhosis

MICROSCOPIC • Histologic features are similar to native liver ○ Features can be patchy within core biopsy with some completely normal portal tracts • Periductal, epithelioid, nonnecrotizing granulomatous reaction

○ Definitive for recurrent disease if present • Lymphocytic cholangitis • Cholate stasis may be seen as bile ducts are progressively lost

TOP DIFFERENTIAL DIAGNOSES • T-cell-mediated rejection (TCMR) ○ Often affects most portal tracts – Unlike recurrent PBC ○ Infiltrate more mixed in TCMR and includes prominent eosinophils ○ Granulomatous bile duct injury not seen in TCMR • Chronic rejection ○ Often preceded by multiple episodes of TCMR ○ Bile duct senescent changes more prominent in chronic rejection ○ Lymphocyte-mediated bile duct injury less prominent ○ Granulomatous bile duct injury not seen in chronic rejection

Lymphocytic Cholangitis

Lymphocytic Cholangitis and Periductal Granuloma

Mixed Portal Infiltrate

Lobular Granuloma

(Left) The bile duct in this portal tract is infiltrated by lymphocytes (lymphocytic cholangitis ﬇), a finding seen in primary biliary cholangitis. No granuloma is associated with this injured bile duct. (Right) In this example of recurrent primary biliary cholangitis, a well-formed granuloma is seen adjacent to a bile duct infiltrated by lymphocytes ﬇.

(Left) No granuloma is seen in this example of recurrent primary biliary cholangitis, and the infiltrate is mixed with scattered eosinophils. This finding raises the possibility of T-cell-mediated rejection; however, the patient was 8 years from transplantation, and other portal tracts did not show a mixed infiltrate. (Right) In addition to portal granulomas adjacent to injured bile ducts, scattered lobular granulomas ﬈ may also be seen.

282

Recurrent Primary Biliary Cholangitis

Abbreviations • Primary biliary cholangitis (PBC) ○ Formerly known as primary biliary cirrhosis

Definitions • Recurrence of autoimmune disease affecting interlobular and septal bile ducts

CLINICAL ISSUES Epidemiology • Prevalence increases post transplantation ○ Rare within 1 year post transplantation ○ 10-30% within 5 years ○ 30-50% within 10 years

Presentation • Most cases of recurrent disease discovered by protocol liver biopsies • Sequela of chronic cholestasis if progressive ○ Jaundice ○ Pruritus ○ Xanthomas • Risk factors for recurrence ○ Male donor ○ Reduced immunosuppression ○ Living-related donor transplantation – Suggesting genetic factors in donor

○ Copper deposition can be demonstrated with histochemistry – Rhodanine stain – Orcein stain • Ductular reaction can also be seen ○ Highlighted using cytokeratin 7 immunohistochemistry

DIFFERENTIAL DIAGNOSIS T-Cell-Mediated Rejection • Has lymphocyte-mediated bile duct damage • Often affects most portal tracts ○ Unlike recurrent PBC • Infiltrate more mixed in T-cell-mediated rejection (TCMR) and includes prominent eosinophils • Granulomatous bile duct injury not seen in TCMR • Alkaline phosphatase increases more gradually and persistently in recurrent PBC ○ In contrast to relatively rapid rise seen in TCMR

Chronic Rejection • Both recurrent PBC and chronic rejection can result in progressive bile duct loss ○ But they are relatively easy to distinguish • Often preceded by multiple episodes of TCMR • Bile duct senescent changes more prominent • Lymphocyte-mediated bile duct injury less prominent • Granulomatous bile duct injury not seen

DIAGNOSTIC CHECKLIST

Laboratory Tests

Clinically Relevant Pathologic Features

• Mildly cholestatic chemistry profile (e.g., elevated alkaline phosphatase, GGT, bilirubin) • Positive antimitochondrial antibody less useful for diagnosis of recurrent PBC

• Recurrent PBC must be distinguished from other causes of cholestatic chemistry profile, particularly chronic rejection • Recurrent PBC has only limited effect on graft survival ○ Excellent prognosis

Treatment

Pathologic Interpretation Pearls

• Drugs ○ Ursodeoxycholic acid

• Granulomatous bile duct injury virtually diagnostic in proper clinical setting

Prognosis • Excellent; rarely progresses to cirrhosis

SELECTED REFERENCES 1.

MICROSCOPIC Histologic Features • Histologic features similar to those seen in native liver ○ Can be patchy in core biopsy with some completely normal portal tracts • Variable portal inflammation with lymphoplasmacytic infiltrate • Lymphocytic cholangitis • Periductal, epithelioid, nonnecrotizing granulomatous reaction ○ May be focal ○ Definitive for recurrent disease, if present • Spotty lobular inflammation • Rare lobular granulomas • Cholate stasis may be seen as bile ducts are progressively lost ○ Periportal hepatocyte swelling ○ May have Mallory-Denk bodies

Liver Transplantation

TERMINOLOGY

2. 3. 4.

5. 6.

7. 8.

Carbone M et al: Liver transplantation in PBC and PSC: indications and disease recurrence. Clin Res Hepatol Gastroenterol. 35(6-7):446-54, 2011 Adeyi O et al: Liver allograft pathology: approach to interpretation of needle biopsies with clinicopathological correlation. J Clin Pathol. 63(1):47-74, 2010 Silveira MG et al: Recurrent primary biliary cirrhosis after liver transplantation. Am J Transplant. 10(4):720-6, 2010 Hytiroglou P et al: Recurrence of primary biliary cirrhosis and development of autoimmune hepatitis after liver transplant: a blind histologic study. Hepatol Res. 39(6):577-84, 2009 Li KK et al: Recurrent nonviral liver disease following liver transplantation. Expert Rev Gastroenterol Hepatol. 3(3):257-68, 2009 Abraham SC et al: Histologic abnormalities are common in protocol liver allograft biopsies from patients with normal liver function tests. Am J Surg Pathol. 32(7):965-73, 2008 Banff Working Group et al: Liver biopsy interpretation for causes of late liver allograft dysfunction. Hepatology. 44(2):489-501, 2006 Neuberger J: Recurrent primary biliary cirrhosis. Liver Transpl. 9(6):539-46, 2003

283

Liver Transplantation

Recurrent Primary Sclerosing Cholangitis KEY FACTS

CLINICAL ISSUES • Prevalence of recurrent primary sclerosing cholangitis (PSC) ranges from 6-60% • Most cases recur > 1 year post transplantation • Often have recurrent episodes of cholangitis • Risk factors ○ Active inflammatory bowel disease at time of transplantation ○ Presence of colon at time of transplantation • Good prognosis with > 80% 5-year survival

IMAGING • MRCP or ERCP shows multifocal nonanastomotic strictures

MICROSCOPIC • Features of chronic biliary obstruction ○ Bile ductular reaction with neutrophils ○ Cholate stasis

– Predominately periportal hepatocellular swelling/ballooning (pseudoxanthomatous change/feathery degeneration) – May have Mallory-Denk bodies – Copper deposition ○ Bile duct loss ○ Progressive fibrosis • "Onion skin" fibrosis and fibroobliterative lesions rare ○ If present, favor PSC

TOP DIFFERENTIAL DIAGNOSES • Chronic rejection ○ Bile duct senescent changes are characteristic ○ Cholestasis usually present ○ Ductular reaction usually not present • Postsurgical biliary stricture ○ Biliary strictures away from surgical anastomosis and long after transplantation favor recurrent PSC

Ductular Reaction With Neutrophils

Bile Duct Loss and Cholate Stasis

Fibroobliterative Lesion

"Onion Skin" Fibrosis

(Left) In areas affected by primary sclerosing cholangitis (PSC), a prominent ductular reaction with associated neutrophils is often seen. These features indicate biliary obstruction. (Right) Over time, there is bile duct loss with cholate stasis characterized by hepatocyte swelling ± MalloryDenk bodies ﬈.

(Left) A fibroobliterative lesion seen in PSC is characterized by a fibrous knot ﬈, indicative of an obliterated bile duct. (Right) Rarely in PSC, a medium-sized bile duct with associated "onion skin" fibrosis ﬈ will be sampled. This feature is highly suggestive of recurrent PSC.

284

Recurrent Primary Sclerosing Cholangitis

Abbreviations • Primary sclerosing cholangitis (PSC)

Definitions • Idiopathic destruction of biliary tree often associated with inflammatory bowel disease

CLINICAL ISSUES Epidemiology • Prevalence of recurrent PSC ○ ~ 25%; range: 6-60% • Most recur > 1 year post transplantation ○ Median: 4.6 years post transplantation

Presentation • Recurrent episodes of cholangitis • Biliary strictures with resulting cholestatic chemistry profile ○ Must occur > 90 days post transplant • Risk factors for recurrent PSC ○ Active inflammatory bowel disease at time of transplantation ○ Presence of colon at time of transplantation ○ CMV infection ○ Male sex ○ Extended donor criteria ○ High immunosuppression

Treatment • Retransplantation required in 12%, higher than recurrent primary biliary cholangitis

Prognosis • Generally good ○ Recurrent PSC does not impact graft or patient survival compared to non-PSC patients ○ 80% 5-year patient survival rate post transplantation • If present, inflammatory bowel disease may be exacerbated

IMAGING Radiographic Findings • MRCP or ERCP ○ Multifocal nonanastomotic strictures – Both intra- and extrahepatic bile ducts – Intervening segments of normal or dilated ducts

○ Progressive fibrosis – Biliary-type fibrosis with irregular nodules – Can be patchy on biopsy depending on ducts involved • Rare "onion skin" fibrosis and fibroobliterative lesions ○ Favors recurrent PSC if present ○ Rarely seen in other entities, such as hepatic artery thrombosis • Larger bile ducts (seen in explant liver) show ○ Ulceration ○ Bile sludge ○ Lymphoplasmacytic infiltrate • Canalicular cholestasis usually not present until late

DIFFERENTIAL DIAGNOSIS Chronic Rejection • • • • •

Usually within 1 year post transplantation Usually in setting of prior episodes of acute rejection Cholestasis usually present Ductular reaction usually not present Bile duct senescent changes are characteristic ○ Cytoplasmic eosinophilia ○ Nuclear enlargement and multinucleation ○ Uneven nuclear spacing ○ Loss of polarity • Progressive fibrosis unusual in chronic rejection, except around central vein

Postsurgical Biliary Stricture • Biliary strictures away from surgical anastomosis favor recurrent PSC • Strictures long after transplantation are more suggestive of recurrent PSC • Fibroobliterative lesions and "onion-skinning" not seen in postsurgical biliary strictures

Hepatic Artery Thrombosis • Results in biliary necrosis and stricture • Often demonstrates centrilobular hepatocyte ballooning/necrosis

Small for Size Syndrome • Increased portal blood flow decreases arterial flow • Bile ducts can be damaged from decreased arterial blood • Periportal hepatocyte necrosis usually present

SELECTED REFERENCES 1.

MICROSCOPIC

2.

Histologic Features • Features of chronic biliary obstruction ○ Bile ductular reaction with neutrophilic infiltrate – Luminal collections of neutrophils (acute cholangitis) can indicate bacterial infection (ascending cholangitis) ○ Cholate stasis – Predominately periportal hepatocellular swelling/ballooning (pseudoxanthomatous change/feathery degeneration) – May have Mallory-Denk bodies – Copper deposition ○ Bile duct loss

Liver Transplantation

TERMINOLOGY

3.

4. 5. 6.

7. 8.

Fosby B et al: Recurrence and rejection in liver transplantation for primary sclerosing cholangitis. World J Gastroenterol. 18(1):1-15, 2012 Adeyi O et al: Liver allograft pathology: approach to interpretation of needle biopsies with clinicopathological correlation. J Clin Pathol. 63(1):47-74, 2010 Alabraba E et al: A re-evaluation of the risk factors for the recurrence of primary sclerosing cholangitis in liver allografts. Liver Transpl. 15(3):330-40, 2009 Banff Working Group et al: Liver biopsy interpretation for causes of late liver allograft dysfunction. Hepatology. 44(2):489-501, 2006 Demetris AJ: Distinguishing between recurrent primary sclerosing cholangitis and chronic rejection. Liver Transpl. 12(11 Suppl 2):S68-72, 2006 Maheshwari A et al: Long-term outcome of liver transplantation in patients with PSC: a comparative analysis with PBC. Am J Gastroenterol. 99(3):53842, 2004 Vera A et al: Risk factors for recurrence of primary sclerosing cholangitis of liver allograft. Lancet. 360(9349):1943-4, 2002 Graziadei IW et al: Recurrence of primary sclerosing cholangitis following liver transplantation. Hepatology. 29(4):1050-6, 1999

285

Liver Transplantation

Recurrent Fatty Liver Disease KEY FACTS ○ Ballooning degeneration

CLINICAL ISSUES • 10-30% of patients return to drinking following transplantation for alcoholic cirrhosis ○ But few develop severe disease • Recurrence of nonalcoholic fatty liver disease varies widely between studies ○ 15-60% develop recurrent steatosis within 1 year • Excellent prognosis ○ Only 5-10% develop recurrent cirrhosis

MICROSCOPIC

TOP DIFFERENTIAL DIAGNOSES • Recurrent genotype 3 hepatitis C virus (HCV) ○ Can present with marked steatosis ○ Correlation with viral load and knowledge of HCV genotype is essential • De novo steatohepatitis ○ Histologically indistinguishable from recurrent fatty liver disease ○ Knowledge of pretransplant liver disease is essential

• Steatosis > 5% is necessary for diagnosis of fatty liver disease ○ Steatosis can be macrovesicular or small droplets ○ Microvesicular steatosis does not count toward overall percentage of fat • Generally required to diagnose steatohepatitis ○ Steatosis > 5% ○ Lobular inflammation

Macrovesicular Steatosis

Ballooning Degeneration

Mild Portal Inflammation

Sinusoidal Fibrosis (Trichrome Stain)

(Left) In this example of recurrent steatosis, there is both macrovesicular ﬈ and small droplet steatosis. Ballooned hepatocytes are not seen. (Right) In this patient with recurrent steatohepatitis, there is prominent ballooning degeneration ﬈.

(Left) Some cases of recurrent steatohepatitis will have mild portal inflammation. This finding should not be mistaken for chronic hepatitis or acute rejection. (Right) In this example of recurrent fatty liver disease, the fibrosis is quite dense and surrounds ballooned hepatocytes, some of which contain Mallory-Denk bodies ﬈.

286

Recurrent Fatty Liver Disease

Abbreviations • • • • •

Nonalcoholic fatty liver disease (NAFLD) Nonalcoholic steatohepatitis (NASH) Alcoholic liver disease (ALD) Alcoholic steatohepatitis (ASH) Hepatitis C virus (HCV)

Definitions • Chronic liver disease characterized by ○ Steatosis ○ Hepatocyte ballooning degeneration ○ Progressive fibrosis

ETIOLOGY/PATHOGENESIS Risk Factors • • • •

Alcohol Metabolic syndrome Obesity Medications

CLINICAL ISSUES Epidemiology • Incidence ○ ALD – 10-30% of patients return to drinking following transplantation for alcoholic cirrhosis ○ Nonalcoholic liver disease – Recurrent disease varies widely between studies □ 15-60% develop recurrent steatosis within 1 year – Recurrence of NASH also varies widely □ 5-15% at 1 year ○ De novo steatohepatitis may occur due to posttransplant metabolic derangements caused by immunosuppressive medications – Medications can exacerbate recurrent NASH

– Usually in form of ballooned hepatocytes □ Large hepatocytes with wispy, clear cytoplasm and central nuclei □ Although not required, some ballooned hepatocytes may contain Mallory-Denk bodies • Distinguishing between ALD and NAFLD requires clinical correlation ○ For ALD, following are often more prominent – Ballooning degeneration – Mallory-Denk bodies – Neutrophilic inflammation, including satellitosis (neutrophils surrounding ballooned hepatocytes) • Other features seen but not essential include ○ Megamitochondria ○ Portal inflammation ○ Microvesicular steatosis ○ Glycogenosis • Fibrosis begins around central veins in sinusoidal distribution ○ Central-central and portal-central bridging fibrosis develops as disease progresses ○ Portal tracts eventually become expanded and fibrotic ○ Cirrhosis often micronodular ○ In ASH, central veins may become obliterated (central hyaline sclerosis)

DIFFERENTIAL DIAGNOSIS Recurrent Hepatitis C Virus, Genotype 3 • Can present with marked steatosis • Essential to correlate with viral load and knowledge of HCV genotype

De Novo Steatohepatitis • Histologically indistinguishable from recurrent fatty liver disease • Essential to know pretransplant liver disease

SELECTED REFERENCES

Presentation

1.

• Often clinically silent • Elevated liver enzymes (ALT and AST) on routine follow-up in some cases

2.

Treatment

4.

• Abstinence for recurrent ALD • Reducing risk factors for progression of NAFLD ○ Weight loss ○ Diabetes control ○ Lipid control

Prognosis • Excellent ○ Only 5-10% develop recurrent cirrhosis

Liver Transplantation

TERMINOLOGY

3.

5. 6. 7. 8.

Charlton M: Evolving aspects of liver transplantation for nonalcoholic steatohepatitis. Curr Opin Organ Transplant. 18(3):251-8, 2013 Friman S: Recurrence of disease after liver transplantation. Transplant Proc. 45(3):1178-81, 2013 Patil DT et al: Evolution of nonalcoholic fatty liver disease recurrence after liver transplantation. Liver Transpl. 18(10):1147-53, 2012 Yalamanchili K et al: Nonalcoholic fatty liver disease after liver transplantation for cryptogenic cirrhosis or nonalcoholic fatty liver disease. Liver Transpl. 16(4):431-9, 2010 Malik SM et al: Recurrent disease following liver transplantation for nonalcoholic steatohepatitis cirrhosis. Liver Transpl. 15(12):1843-51, 2009 Ayata G et al: Cryptogenic cirrhosis: clinicopathologic findings at and after liver transplantation. Hum Pathol. 33(11):1098-104, 2002 Charlton M et al: Frequency of nonalcoholic steatohepatitis as a cause of advanced liver disease. Liver Transpl. 7(7):608-14, 2001 Contos MJ et al: Development of nonalcoholic fatty liver disease after orthotopic liver transplantation for cryptogenic cirrhosis. Liver Transpl. 7(4):363-73, 2001

MICROSCOPIC Histologic Features • Generally required to diagnosis steatohepatitis ○ Steatosis > 5% ○ Lobular inflammation ○ Hepatocyte injury 287

Liver Transplantation

Cytomegalovirus KEY FACTS

CLINICAL ISSUES • Cytomegalovirus (CMV) effects in liver transplantation ○ Associated with increased risk of death, graft loss, retransplantation, biliary complications, and severe recurrent hepatitis C virus ○ > 50% of CMV disease involves liver • Without prophylaxis, CMV infection in solid organ transplant patients occurs early in posttransplant period, when immunosuppression is most intense • Liver allograft most prone to CMV infection ○ > 50% of CMV disease affects liver • Prophylaxis delays infection ○ Does not completely inhibit/prevent it

MICROSCOPIC • Cytopathic effect seen in ○ Hepatocytes ○ Endothelial cells ○ Biliary epithelium

○ Kupffer cells • Characteristic cytomegaly and intranuclear, intracytoplasmic inclusions ○ Viral cytopathic effect not always seen • Neutrophilic microabscesses highly suggestive of CMV hepatitis ○ Can also occur in conditions unrelated to CMV

ANCILLARY TESTS • Hematoxylin and eosin readily shows viral cytopathic effect ○ Immunohistochemistry helps confirm presence of virus if needed

TOP DIFFERENTIAL DIAGNOSES • • • •

Graft rejection Graft ischemia Biliary obstruction Other infections

Microabscess and CMV-Infected Hepatocyte

CMV Immunohistochemistry

CMV-Infected Hepatocyte

CMV-Infected Hepatocyte

(Left) A hepatocyte displaying nuclear and cytoplasmic viral cytopathic effect is present in the middle of a cluster of neutrophils (microabscess). Although microabscesses are not entirely specific for cytomegalovirus (CMV), this case nicely illustrates the presence of microabscess in relationship to an infected hepatocyte. (Right) Immunohistochemical staining for CMV highlights the viral inclusion (nuclear in this case).

(Left) A hepatocyte ﬊ infected by CMV demonstrates cytomegaly, a glassy nuclear inclusion, and cytoplasmic inclusions. CMV, unlike some other viruses, causes both nuclear and cytoplasmic inclusions. (Right) This liver biopsy shows mild portal inflammation. At the periphery of the portal tract, there is a CMV-infected hepatocyte with prominent cytoplasmic viral inclusions as well as nuclear viral cytopathic effect ﬊.

288

Cytomegalovirus

Abbreviations • Cytomegalovirus (CMV)

Definitions • CMV infection: Isolation of CMV or detection of viral proteins or nucleic acid in any body fluid or tissue specimen • CMV disease: Fever, neutropenia, or thrombocytopenia with detection of CMV in blood or tissue

ETIOLOGY/PATHOGENESIS Infectious Agents • DNA virus, member of Herpesviridae • CMV has direct immunosuppressive effects that increase risk for other opportunistic infections ○ Potentially leads to greater hepatitis C viral load and exacerbates hepatitis C virus (HCV) recurrence in transplant liver

CLINICAL ISSUES Presentation • Without prophylaxis, CMV infection in solid organ transplant patients occurs early in posttransplant period (most often within first 3 months), when immunosuppression is most intense • Can present with signs of hepatitis, fever • Liver allograft most prone to CMV infection; > 50% of CMV disease affects liver • In delayed onset, infection outside liver seen, especially in gastrointestinal tract • Associated with increased risk of death, graft loss, retransplantation, biliary complications, and severe HCV recurrence • Risk factors ○ Increased immunosuppression ○ Antilymphocyte antibody or cytotoxic drugs

Treatment • Prophylaxis: Given for 3-6 months or longer • Ganciclovir or valganciclovir for risk of developing CMV disease ○ ↑ risk of CMV reactivation during periods of intense immunosuppression (e.g., when treating acute rejection) necessitates restarting prophylaxis during this time • Preemptive therapy ○ Based on detection of CMV reactivation before onset of clinical symptoms ○ Ganciclovir or valganciclovir-ganciclovir recommended mainly for moderate- or low-risk patients, such as CMV seropositive recipients • Antiviral treatment of CMV disease: Intravenous ganciclovir therapy

Prognosis • Direct and indirect effects of CMV may be reduced by prophylaxis with antivirals ○ Late primary infections may complicate posttransplant course • Prophylaxis delays infection but does not completely inhibit/prevent it

Diagnostic Tests • Tissue biopsy is gold standard • PCR

MICROSCOPIC

Liver Transplantation

○ Once prophylaxis discontinued, risk of CMV infection returns

TERMINOLOGY

Histologic Features • Liver parenchymal findings ○ Scattered neutrophil aggregates (microabscesses) in liver parenchyma ○ Microgranulomas may also be seen ○ Typical viral hepatitis appearance (lobular inflammation and lobular disarray) can be seen • Cytopathic effect ○ Affects hepatocytes, endothelial cells, biliary epithelium, Kupffer cells ○ Intranuclear and cytoplasmic inclusions – Intranuclear inclusions: Nuclei enlarged and contain round, glassy aggregate surrounded by clear halo and thickened nuclear membrane – Intracytoplasmic inclusions: Basophilic granules in cytoplasm

ANCILLARY TESTS Immunohistochemistry • CMV antigens confirm presence of virus

DIFFERENTIAL DIAGNOSIS Graft Rejection • Like graft rejection, CMV hepatitis can demonstrate mild portal inflammation, including inflammation of bile ducts and venules • Graft rejection will not have viral inclusions or positive CMV immunohistochemistry

Other Infections, Graft Ischemia, Biliary Obstruction • Parenchymal neutrophil microabscesses can also occur in these conditions • Numerous microabscesses (> 9) within biopsy correlated with CMV infection in 1 report; no other histologic features, including size of microabscess, correlated with etiology of microabscess

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Characteristic intranuclear and cytoplasmic viral inclusions as well as cytomegaly

SELECTED REFERENCES 1. 2.

3. 4.

Lautenschlager I: CMV infection, diagnosis and antiviral strategies after liver transplantation. Transpl Int. 22(11):1031-40, 2009 Lautenschlager I et al: Cytomegalovirus infection of the liver transplant: virological, histological, immunological, and clinical observations. Transpl Infect Dis. 8(1):21-30, 2006 Limaye AP et al: Late-onset cytomegalovirus disease in liver transplant recipients despite antiviral prophylaxis. Transplantation. 78(9):1390-6, 2004 Lamps LW et al: The significance of microabscesses in liver transplant biopsies: a clinicopathological study. Hepatology. 28(6):1532-7, 1998

289

Liver Transplantation

Herpes Simplex Virus KEY FACTS

CLINICAL ISSUES • Herpes simplex virus (HSV) hepatitis is uncommon • HSV hepatitis can occur as early as 5 days after transplantation ○ In liver transplant setting appears to occur early in postoperative course • Mucocutaneous lesions may not be evident in patient • Clinical suspicion and early diagnosis is critical • Risk factors include noncompliance or intolerance of prescribed prophylactic antiviral therapy • Empiric antiviral therapy pending diagnostic evaluation may be life saving

MICROSCOPIC • Coagulative necrosis • 3 M's: Molding, multinucleation, margination: Viral inclusions in hepatocyte nuclei at edge of necrosis ○ Cowdry A: Intranuclear inclusion with halo

○ Cowdry B: Ground-glass smudgy nucleus with chromatin margination ○ Only nuclear inclusions • Random foci of hepatocyte coagulative necrosis containing neutrophils and macrophages

TOP DIFFERENTIAL DIAGNOSES • Elevated liver enzymes may be initial presentation ○ T-cell-mediated rejection must be considered in differential • Cytomegalovirus can have both nuclear and cytoplasmic inclusions ○ HSV has only nuclear inclusions • Adenovirus infection can look similar • Immunohistochemistry can distinguish between HSV and adenovirus

Hepatocyte Infected by HSV Demonstrating Multinucleation

Nuclear Inclusions With Chromatin Margination in HSV Hepatitis

Coagulative Necrosis

HSVI/II Immunohistochemical Stain

(Left) Herpes simplex virus (HSV)-infected hepatocyte seen on touch imprint demonstrates multinucleation, nuclear molding, and chromatin margination. (Courtesy C. Fligner, MD and M. Lawless, MD.) (Right) Cowdry type A ﬇ nuclear inclusion is a central eosinophilic inclusion, and type B ﬊ is characterized by a glassy nucleus with peripheralized chromatin. (Courtesy C. Fligner, MD and M. Lawless, MD.)

(Left) This biopsy was taken from a transplant patient with HSV hepatitis. It shows a rounded area of coagulative necrosis ﬊ with an infected hepatocyte ﬉. (Right) HSVI/II immunohistochemical stain highlights a confluent area of HSV-infected hepatocytes.

290

Herpes Simplex Virus

Abbreviations • Herpes simplex virus (HSV)

ETIOLOGY/PATHOGENESIS Infectious Agents • HSV1 and HSV2 can cause hepatitis

Risk Factors • Liver transplant recipients are immunocompromised and susceptible to viral reactivation, reinfection, or primary infection ○ Overall frequency of HSV hepatitis does not differ among type of solid organ transplant ○ Acquisition of virus from donor organs can occur

CLINICAL ISSUES Epidemiology • Incidence ○ HSV hepatitis uncommon – < 150 cases reported currently – 10 of reported patients underwent liver transplantation as result of HSV hepatitis – 30% of published HSV hepatitis cases found in solid organ transplant recipients

Presentation • In liver transplant setting, appears to occur early in postoperative course ○ Can occur as early as 5 days after transplantation • Mucocutaneous lesions may not always be present

Treatment • Drugs ○ Treatment with acyclovir early in disease may be life saving – May benefit from intravenous acyclovir pending results of liver biopsy and other studies

Prognosis • Of reported cases of HSV hepatitis in native livers, 6 of 10 patients who underwent liver transplantation died within 1st year ○ Only 1 reported death considered related to recurrent HSV infection • Recovery in children better than in adults both before and after liver transplantation for HSV hepatitis • Diffuse involvement of liver associated with high mortality • Focal involvement may be treatable with antiviral therapy • Elevated liver enzymes not associated with increased mortality

MICROSCOPIC Histologic Features • Random foci of hepatocyte coagulative necrosis containing neutrophils and macrophages ○ May develop into confluent necrosis • Nuclear inclusions in hepatocytes at edge of necrotic areas

○ Cowdry type A: Eosinophilic-basophilic intranuclear inclusion surrounded by halo ○ Cowdry type B: Smudgy, glassy nuclear inclusion occupying majority of nucleus with margination of chromatin against nuclear membrane • Formation of syncytia

ANCILLARY TESTS

Liver Transplantation

TERMINOLOGY

Immunohistochemistry • Immunohistochemistry directed against HSV1 and HSV2 confirms diagnosis

DIFFERENTIAL DIAGNOSIS Ischemia • Hemorrhagic necrosis in zone 3 (central vein) distribution • No viral inclusions

Adenovirus • Inclusions may be similar • No syncytial forms • Immunohistochemistry can distinguish between HSV and adenovirus • Other diagnostic studies include ○ Adenovirus antigen detection ○ Polymerase chain reaction assay ○ Virus isolation ○ Serology

Cytomegalovirus • Both usually occur early in posttransplant period ○ Cytomegalovirus (CMV) hepatitis occurs toward end of 1st month post transplantation (peaks 30-40 days after transplantation) ○ HSV hepatitis occurs earlier (20 ± 12 days) • Distinct clinical picture ○ CMV hepatitis rarely fulminant • CMV associated with microabscesses ○ HSV associated with areas of necrosis and no inflammation • CMV can have both nuclear and cytoplasmic inclusions; HSV has only nuclear inclusions

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Focal vs. diffuse involvement

Pathologic Interpretation Pearls • Look for random foci of hepatocyte coagulative necrosis and viral inclusions

SELECTED REFERENCES 1. 2.

3. 4. 5.

Côté-Daigneault J et al: Herpes simplex hepatitis after liver transplantation: case report and literature review. Transpl Infect Dis. 16(1):130-4, 2014 Ambrosioni J et al: Herpes simplex virus load to monitor antiviral treatment after liver transplantation for acute herpetic hepatitis. Antivir Ther. 17(2):401-4, 2012 Basse G et al: Disseminated herpes simplex type-2 (HSV-2) infection after solid-organ transplantation. Infection. 36(1):62-4, 2008 Norvell JP et al: Herpes simplex virus hepatitis: an analysis of the published literature and institutional cases. Liver Transpl. 13(10):1428-34, 2007 Kusne S et al: Herpes simplex virus hepatitis after solid organ transplantation in adults. J Infect Dis. 163(5):1001-7, 1991

291

Liver Transplantation

Adenovirus, Liver KEY FACTS

CLINICAL ISSUES

MICROSCOPIC

• Adenovirus (AdV) infection associated with high rates of graft failure, morbidity, and mortality • Most common predisposing condition for AdV hepatitis in pediatric patients is liver transplantation • Differential diagnosis of pediatric liver transplant patients and patients under heavy immunosuppression with marked elevation of aspartate transaminase (AST)/alanine transaminase (ALT) should include AdV • Serum liver transaminases (AST, ALT) may be markedly elevated in the thousands, with AST > > ALT • Reduce immunosuppression ○ Cidofovir may be considered

• Small or large areas of coagulative hepatocyte necrosis in random (nonzonal) distribution • Viral inclusions with smudgy appearance and chromatin margination • Immunohistochemistry confirms tissue diagnosis • Minimal inflammation may be present

IMAGING

DIAGNOSTIC CHECKLIST

• AdV infected areas in liver can be focal and may require targeted biopsy

• Coagulative necrosis with intranuclear smudgy inclusions on biopsy • Positive AdV immunohistochemistry

TOP DIFFERENTIAL DIAGNOSES • • • •

Herpes simplex virus Varicella zoster virus Cytomegalovirus Drug-induced hepatitis

Random Necrosis in Adenovirus Hepatitis

Intranuclear Viral Inclusion

Adenovirus Immunohistochemistry

Adenovirus Immunohistochemistry

(Left) This low-power view of a liver biopsy shows random areas of coagulative necrosis ﬈. Unlike other causes of hepatitis, viral infection often induces a random distribution of necrosis (not pericentral or periportal).(Courtesy R. Wilcox, MD.) (Right) Highpower view demonstrates a hepatocyte with a smudgy intranuclear viral inclusion ﬈.

(Left) Adenovirus immunohistochemistry highlights numerous infected hepatocytes within and around a necrotic area ﬇. Necrosis in adenovirus hepatitis can range from being spotty to showing massive involvement of the liver. There appears to be no survival advantage to having limited necrosis in the liver. (Right) Immunohistochemistry for adenovirus highlights nuclear viral inclusions in infected hepatocytes. (Courtesy R. Wilcox, MD.)

292

Adenovirus, Liver

Abbreviations • Adenovirus (AdV), aspartate transaminase (AST), alanine transaminase (ALT)

ETIOLOGY/PATHOGENESIS Infectious Agents • Nonenveloped, double-stranded DNA virus ○ Divided into 6 major subgroups ○ Serotypes 1, 2, 5 most commonly associated with hepatic disease

Risk Factors • Children ○ More likely from recent primary exposure – Adult cases more likely from reactivation • Residents in close quarters (i.e., dormitories, military barracks) • Immunocompromised patients ○ Liver transplant (LT) recipients susceptible to de novo AdV infection and reactivation of latent infection • Transmission via direct exposure to infected tissue or blood, aerosolized droplets, oral-fecal spread • Incubation time: 2-14 days

CLINICAL ISSUES Epidemiology • Incidence ○ 2-5% infection rate in pediatric series

Presentation • Asymptomatic to nonspecific fever, rash, upper respiratory infection, cystitis, hepatitis • LT patients present with hepatitis as primary manifestation ○ Markedly elevated AST and ALT should raise AdV in differential diagnosis – Especially with muromonab-CD3 or polyclonal antithymocyte globulin use – Also seen with treatment of acute rejection with increased immunosuppression

Treatment • Withdrawing or reducing immunosuppression is 1st option • No FDA-approved drugs available • Reports of successful treatment with cidofovir, especially pediatric LT recipients ○ Cidofovir use limited by nephrotoxicity • Ribavirin and intravenous immunoglobulin with reported success

• Intranuclear viral inclusions with smudgy appearance and chromatin margination in infected hepatocytes • Bile duct epithelial cells may be infected

ANCILLARY TESTS Immunohistochemistry • Anti-adenovirus antibody can confirm diagnosis

Other • Polymerase chain reaction testing of viral DNA, viral antigen detection, culture isolation of AdV, serologic testing

DIFFERENTIAL DIAGNOSIS Cytomegalovirus • Both cytomegalovirus (CMV) and AdV occur in early posttransplant period • CMV associated with microabscesses ○ Whereas AdV is associated with necrosis and no inflammation • Infected cells are cytomegalic • CMV has both nuclear and cytoplasmic viral inclusions ○ Immunohistochemistry confirms CMV infection

Herpes Simplex Virus • Both AdV and herpes simplex virus (HSV) associated with coagulative necrosis and intranuclear viral inclusions • Immunohistochemistry helpful • Multinucleated cells in HSV but not AdV

Varicella Zoster Virus • Similar to HSV: Coagulative necrosis and intranuclear viral inclusions

Drug-Induced Hepatitis • Relevant drug exposure history • Cholestasis &/or steatosis may be greater • No viral inclusions

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Coagulative necrosis with intranuclear smudgy inclusions on biopsy; positive immunohistochemistry for AdV

Clinical Features • Elevated serum AST/ALT and hepatic failure in immunosuppressed LT recipients • PCR(+), culture positive for AdV

SELECTED REFERENCES 1.

Prognosis • AdV hepatitis in LT patients usually fatal ○ > 50% mortality rate

2. 3.

MICROSCOPIC

4.

Histologic Features

5.

• Coagulative necrosis (small or large areas) without specific zonal distribution • Minimal inflammatory cell response

Liver Transplantation

TERMINOLOGY

6.

Schaberg KB et al: Adenovirus hepatitis: clinicopathologic analysis of 12 consecutive cases from a single institution. Am J Surg Pathol. 41(6):810-819, 2017 Ronan BA et al: Fulminant hepatitis due to human adenovirus. Infection. 42(1):105-11, 2014 Cimsit B et al: Treatment of adenovirus hepatitis with cidofovir in a pediatric liver transplant recipient. Pediatr Transplant. 16(3):E90-3, 2012 Perez D et al: Successful outcome of severe adenovirus hepatitis of the allograft following liver transplantation. Transpl Infect Dis. 9(4):318-22, 2007 Ison MG: Adenovirus infections in transplant recipients. Clin Infect Dis. 43(3):331-9, 2006 Saad RS et al: Adenovirus hepatitis in the adult allograft liver. Transplantation. 64(10):1483-5, 1997

293

Liver Transplantation

Hepatitis E Virus KEY FACTS

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• Transmission mainly via oral-fecal route

• HEV infection usually self-limited ○ May persist & cause chronic hepatitis in immunocompromised patients • Manifestations of HEV infection in immunosuppressed patients can be subtle or nonspecific pathological changes initially, making early clinical diagnosis challenging • Acute HEV ○ Lobular inflammation with acidophil bodies & lymphocytic infiltrate ○ Cholestatic changes but no periductal/portal edema ○ More hepatitic & cholestatic changes seen in immunocompetent than immunocompromised • Chronic HEV ○ Portal lymphocyte-predominant inflammation & mild lobular inflammation ○ Minority develop cirrhosis

CLINICAL ISSUES • Unexplained nonspecific hepatitis in immunosuppressed patients should prompt hepatitis E virus (HEV) IgM & IgG antibody & HEV RNA level testing with recognition of diagnostic assay limitations • In LT patients (unlike immunocompetent patients), acute HEV may not be self-limited ○ Can progress to chronic hepatitis • Anti-HEV IgM seen within incubation period of 2-6 weeks, followed by anti-HEV IgG • In LT patients, HEV antibody seroconversion may never occur ○ HEV RNA level should be obtained if clinically suspected • Wide variability in testing accuracy for HEV antibody & RNA levels exist between laboratories • Up to 10% progress to cirrhosis

HEV Infection, Portal Tract

Lobular Inflammation in Acute Hepatitis E

Chronic HEV Hepatitis

Chronic HEV Hepatitis With Fibrosis

(Left) Liver explant from an HEV-infected patient shows portal tracts with bile ductular reaction. There is an increase in small bile ductules st as well as neutrophils and mild stromal edema. (Courtesy J. Hooper, MD.) HEV hepatitis is characterized by both hepatic as well as cholestatic changes. (Right) In this case of acute hepatitis E, there is extensive lobular inflammation, as well as dead or dying hepatocytes ﬈. Lobular disarray is evident as would be seen in any other form of acute hepatitis. (Courtesy H. Appelman, MD.)

(Left) Liver biopsy from a patient with chronic HEV shows portal tract expansion by lymphocytic inflammation with interface activity. (Right) Reticulin stain highlights fibrosis in this explant liver. (Courtesy J. Hooper, MD.)

294

Hepatitis E Virus

Abbreviations • Hepatitis E virus (HEV)

ETIOLOGY/PATHOGENESIS Infectious Agents • Nonenveloped, single-stranded RNA virus ○ All 4 major genotypes infect humans – Genotypes 3 & 4 can infect other animal species (can be zoonosis) – Consumption of HEV-contaminated undercooked meat □ Major source for HEV infections in developed countries □ May account for some cases after organ transplantation

Risk Factors • Immunosuppression: Renders liver transplant (LT) recipients susceptible to primary HEV infection • Transmission mainly via oral-fecal route ○ Transmission via blood transfusions, solid organ transplantation, consumption of undercooked infected meat

CLINICAL ISSUES Epidemiology • ~ 1-1.5% of LT patients in European countries, 0.1% of LT patients in Japan ○ In British study, 1 in 2,848 blood donors viremic with genotype 3 – 42% of recipients had evidence of infection with persistent infection in immunosuppressed patients ○ Plasma products seem to be specifically causative for HEV infections • Incubation time ○ Range: 2-8 weeks (mean: 6 weeks)

Presentation • Nonspecific fatigue, malaise most common • Nonspecific elevations of liver enzymes (not otherwise explained by other causes of hepatitis) often noted • In LT recipients, severe liver injury or fulminant liver failure has not been reported

Laboratory Tests • Anti-HEV IgM seen within incubation period of 2-6 weeks, followed by anti-HEV IgG ○ In LT patients, HEV antibody seroconversion may never occur & thus HEV RNA level should be obtained if clinically suspected • PCR testing of viral RNA ○ HEV RNA becomes undetectable in serum & stool within weeks after onset of symptoms • Diagnosis in transplanted patients based on HEV RNA testing ○ Antibody assays not sensitive enough

• Up to 10% progress to cirrhosis

MICROSCOPIC Histologic Features • Explant livers from HEV patients ○ Lobular hepatitis ○ Bile ductular reaction [↑ bile ductules (+) associated neutrophils (+) stromal edema] & reactive appearing biliary epithelial cells] ○ Canalicular cholestasis common • Acute HEV in organ transplant patients characterized by predominantly lobular inflammation with no hepatocyte swelling & scattered acidophil bodies ○ Changes can be subtle early in disease process, making diagnosis challenging – In one retrospective study, tissue RNA analysis showed HEV to be present in allograft liver tissue months before diagnosis made ○ Portal tract findings include mild to moderate expansion by lymphocytic inflammation ○ Canalicular cholestasis not feature of HEV infection in allograft liver ○ More lobular plasma cells than in nonallograft HEV • Progressors to chronic hepatitis (57% in one LT series of acute HEV) have dense portal lymphocytic infiltrates with interface activity, fibrosis, & mild to moderate lobular hepatitis

DIFFERENTIAL DIAGNOSIS Hepatitis A, B, & C • Difficult to differentiate etiology of acute or chronic viral hepatitis on histology alone ○ Acute HEV diagnosed based on (+) HEV IgM & serum or stool; HEV RNA needed to confirm HEV infection ○ Negative HAV, HBV, HCV testing to exclude hepatitis

CMV, EBV, HSV, VZV • No viral inclusions in HEV cases • Viral cytopathic effect can be seen in CMV & HSV

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Features of acute viral hepatitis such as lobular inflammation & necrosis • In HEV progressors, typical features of chronic viral hepatitis include portal lymphocytic infiltrates with mild to moderate interface hepatitis • HEV RNA displays PCR positivity

SELECTED REFERENCES 1. 2. 3. 4. 5.

Prognosis • ~ 60% fail to clear HEV infection

Liver Transplantation

TERMINOLOGY

6.

Lenggenhager D et al: Visualization of hepatitis E virus RNA and proteins in the human liver. J Hepatol. 67(3):471-479, 2017 Prost S et al: Detection of viral hepatitis E in clinical liver biopsies. Histopathology. 71(4):580-590, 2017 Behrendt P et al: The impact of hepatitis E in the liver transplant setting. J Hepatol. 61(6):1418-29, 2014 Pischke S et al: Hepatitis E virus infection as a cause of graft hepatitis in liver transplant recipients. Liver Transpl. 16(1):74-82, 2010 Kamar N et al: Hepatitis E virus and chronic hepatitis in organ-transplant recipients. N Engl J Med. 358(8):811-7, 2008 Malcolm P et al: The histology of acute autochthonous hepatitis E virus infection. Histopathology. 51(2):190-4, 2007

295

Liver Transplantation

Epstein-Barr Virus, Liver KEY FACTS

CLINICAL ISSUES • Epstein-Barr virus (EBV) associated with ○ Hepatitis ○ Posttransplant lymphoproliferative disorder (PTLD) • EBV-negative, pediatric liver transplant (LT) patients and patients under heavy immunosuppression with elevated LFTs should have EBV in differential diagnosis • Most PTLD present within ~ 2 years after LT • Immunosuppression reduction is 1st line of management • PTLD masses restricted to liver allograft can present with features of biliary obstruction

MICROSCOPIC • EBV hepatitis characterized by ○ Portal mononuclear infiltrates ○ Sinusoidal infiltration with lymphocytes in linear, beadlike pattern ○ Mild lobular disarray ○ Focal hepatocyte necrosis

○ Scattered atypical lymphocytes • 4 major PTLD categories ○ Early lesions – Preserve hepatic architecture ○ Polymorphic – Can efface portal architecture ○ Monomorphic – Destruction of hepatic architecture with lymphocytic infiltration meeting criteria for lymphomas in immunocompetent patients ○ Classic Hodgkin type (least common) – Must fulfill criteria for classic Hodgkin lymphoma • Not all PTLD is EBV-positive

ANCILLARY TESTS • EBER in situ hybridization on tissue • PCR for immunoglobulin heavy chain or T-cell receptor to confirm clonality of lymphoid proliferation in PTLD cases

EBV Hepatitis

EBV Hepatitis

Sinusoidal Lymphocytic Inflammatory Infiltrates in EBV Hepatitis

EBER In Situ Hybridization

(Left) This low-power view highlights portal inflammatory infiltrates ﬊. (Right) EpsteinBarr virus (EBV) hepatitis is characterized by lymphocytic inflammatory infiltrates within the sinusoids and portal tracts. This micrograph mainly highlights the presence of lymphocytes in the sinusoids.

(Left) High-power view of EBV hepatitis depicts sinusoidal lymphocytic inflammatory infiltrates in a bead-like pattern. (Right) EBER in situ hybridization demonstrates positivity for EBV early RNA st in a portal tract lymphocytic infiltrate.

296

Epstein-Barr Virus, Liver

Abbreviations • Epstein-Barr virus (EBV)

• PTLD masses restricted to liver allograft can present with features of biliary obstruction

Laboratory Tests

Synonyms

• Rising EBV viral load raises suspicion for EBV-related PTLD ○ Confirmatory biopsy necessary for diagnosis

• Human herpesvirus 4

Treatment

• Enveloped, double-stranded DNA virus ○ Types 1 and 2 • In posttransplant lymphoproliferative disorder (PTLD) ○ EBV forms episome in latently infected B cells ○ Immunosuppression results in ↓ T-cell function with lack of T-cell control of B cells → uncontrolled EBVtransformed B-cell proliferation → development of PTLD

• No specific treatment available ○ Symptomatic treatment with rest and adequate hydration • Reduction of immunosuppression is 1st line of treatment ○ Response observed in < 50% of patients ○ Remission often not sustained • Antiviral therapy, such as ganciclovir and acyclovir, reduces viral replication without altering clinical course • Rituximab, combination chemotherapy, and adoptive immunotherapy also options for PTLD

Risk Factors

Prognosis

• EBV status of donor and recipient ○ EBV-negative pediatric patients receiving EBV-positive organs are especially vulnerable – Unable to control primary EBV infection, limit EBV replication, or prevent PTLD ○ Transmission via direct exposure to infected tissue or contact with saliva ○ Rare transmission via aerosolized droplets or blood ○ Contracting primary EBV infection after transplant also important risk factor for PTLD • Pediatric liver transplant (LT) recipient • Intensity and duration of immunosuppression ○ Immunosuppression renders LT recipient susceptible to primary EBV infection, reactivation, or reinfection • Risk factors for development of PTLD specifically after LT ○ Chronic hepatitis C ○ Alcoholic cirrhosis ○ Age > 50

• PTLD in LT has up to 50% mortality rate

ETIOLOGY/PATHOGENESIS Infectious Agents

CLINICAL ISSUES Epidemiology • One of most prevalent viruses infecting humans, affecting 90-95% of world population • Incubation time of 4-6 weeks • Most common in combined liver-kidney transplantation followed by cardiac, liver, lung, and kidney transplants • PTLD occurs in up to 3% of adults and 15% of pediatric LT cases

Presentation • • • • •

Fever Lymphadenopathy Malaise Gastrointestinal and respiratory symptoms EBV-related diseases post transplant are EBV hepatitis and PTLD • Severe or fulminant liver failure rare ○ Most occur post transplant or in immunosuppressed patients with HIV or receiving chemotherapy • Most PTLD present within ~ 2 years after LT

Liver Transplantation

TERMINOLOGY

MICROSCOPIC Histologic Features • EBV hepatitis ○ Portal mononuclear cell inflammation – Subtle bile duct damage – Focal endotheliitis of portal and central veins ○ Sinusoidal lymphocytic infiltration in lined-up, bead-like fashion ○ Lobular disarray with focal hepatocyte necrosis ○ Scattered atypical lymphocytes ○ Granulomas can be found • PTLD ○ 4 major categories – Early lesions (plasmacytic hyperplasia and infectious mononucleosis-like PTLD) – Polymorphic PTLD – Monomorphic PTLD – Classic Hodgkin lymphoma-type PTLD ○ Most PTLD cases are of B cell origin – Only 5% of T-cell or T-/NK-cell origin ○ Not all PTLD cases associated with EBV – EBV negativity associated with longer time to presentation, monomorphic-type PTLD, and T-/NK-cell lymphoma – 60-70% of B-cell PTLD EBV related, while < 10% of Tcell cases are EBV related ○ Early lesions in liver characterized by – Mixed lymphoplasmacytic inflammation in predominantly portal areas – Polyclonal B cells – Preserved architecture ○ Polymorphic PTLD is polyclonal or monoclonal lymphoid infiltration in predominantly portal areas, effacing portal architecture – Lymphoid cells include immunoblasts, plasma cells, and small- and intermediate-sized cells (full range of Blymphocyte maturation) – May demonstrate κ- or λ-light chain class restriction 297

Liver Transplantation

Epstein-Barr Virus, Liver – Clonal immunoglobulin gene rearrangements can be demonstrated with genetic testing – Most common type in pediatric patients □ Related to primary EBV infection ○ Monomorphic PTLD is monoclonal proliferation meeting criteria for lymphomas in immunocompetent hosts – Mass forming with destruction of hepatic architecture – Most commonly diffuse large B-cell subtype – In B-cell PTLD, EBV positivity associated with MUM1/IRF4 expression and nongerminal center phenotype □ PTLD in LT more likely to have germinal-center phenotype (CD10[+/-], Bcl-6[+], MUM-1/IRF4[-], CD138[-]) – Although T-cell or T-/NK-monomorphic PTLD occur, vast majority are B cell, have detectable EBV, and clonal rearrangement of Ig genes ○ Other types include – Burkitt lymphoma – Plasma cell myeloma – Peripheral T-cell lymphoma not otherwise specified – Hepatosplenic T-cell lymphoma ○ Classic Hodgkin lymphoma type is least common form of PTLD – Must fulfill criteria for classic Hodgkin lymphoma – Reed-Sternberg-like cells found in other types of PTLD □ EBV-infected cells may show Reed-Sternberg-like features □ Lack CD15 expression □ Usually CD20(+) and CD45(+), whereas classic Hodgkin lymphoma is CD15(+) and CD45 mostly (-)

ANCILLARY TESTS Immunohistochemistry • For PTLD: Panel of antibodies, such as CD20, CD3, MUM1/IRF4, Bcl-6, CD10, CD138, Bcl-2, and immunoglobulin light chains κ and λ ○ CD15, CD30, pax-5, CD3, CD45 for Hodgkin lymphoma

In Situ Hybridization • EBV-encoded RNA confirms presence of virus

PCR • For immunoglobulin heavy chain or T-cell receptor to confirm clonality of lymphoid proliferation in PTLD cases

DIFFERENTIAL DIAGNOSIS T-Cell-Mediated Rejection • Can coincide with EBV infection • Endothelialitis and bile duct damage occurs in both rejection and EBV infection ○ EBV hepatitis has less duct damage ○ Rejection has more duct damage proportional to severity of inflammatory infiltrate • Mononuclear cell dominance with fewer eosinophils in EBV • EBER in situ hybridization negative or positive only in occasional cells in rejection, whereas many cells positive in EBV disease • Sinusoidal lymphocytic infiltrates more prominent in EBV infection 298

Recurrent Hepatitis C Virus • Portal fibrosis • Less sinusoidal inflammation • EBER in situ hybridization is negative or positive in only occasional cells

Cytomegalovirus • Both occur post transplant • Cytomegalovirus (CMV) associated with microabscesses, intranuclear and cytoplasmic viral inclusions • CMV immunohistochemistry positive

Herpes Simplex Virus • Viral inclusions in hepatocyte nuclei can be at edge of necrosis • Herpes simplex virus (HSV) immunohistochemistry positive

Varicella-Zoster Virus • Skin lesions • Liver biopsy findings similar to HSV

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • PTLD masses in porta hepatis of liver allograft can present with features of biliary obstruction

Pathologic Interpretation Pearls • Sinusoidal lymphocytic inflammation in EBV hepatitis • In situ hybridization for EBV-encoded RNA • PTLD: Early lesions show preserved hepatic architecture ○ Polymorphic PTLD is polyclonal or monoclonal ○ Monomorphic PTLD effaces hepatic architecture – Most commonly diffuse large B-cell subtype

SELECTED REFERENCES 1.

Soriano-López DP et al: A scheduled program of molecular screening for Epstein-Barr Virus decreases the incidence of post-transplantation lymphoproliferative disease in pediatric liver transplantation. Transplant Proc. 48(2):654-7, 2016 2. Al-Mansour Z et al: Post-transplant lymphoproliferative disease (PTLD): risk factors, diagnosis, and current treatment strategies. Curr Hematol Malig Rep. 8(3):173-83, 2013 3. Lo RC et al: Post-transplant lymphoproliferative disorders in liver transplant recipients: a clinicopathological study. J Clin Pathol. 66(5):392-8, 2013 4. Centers for Disease Control. http://www.cdc.gov/ncidod/diseases/ebv.htm. Accessed August 21, 2012 5. Izadi M et al: Hepatic involvement by lymphoproliferative disorders post liver transplantation: PTLD.Int. Survey. Hepatol Int. 5(3):759-66, 2011 6. Kamdar KY et al: Posttransplant lymphoproliferative disease following liver transplantation. Curr Opin Organ Transplant. 16(3):274-80, 2011 7. Kim RD et al: Adult post-transplant lymphoproliferative disease in the liver graft in patients with recurrent hepatitis C. Eur J Gastroenterol Hepatol. 23(7):559-65, 2011 8. Végso G et al: Lymphoproliferative disorders after solid organ transplantation-classification, incidence, risk factors, early detection and treatment options. Pathol Oncol Res. 17(3):443-54, 2011 9. Swerdlow et al. WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. Lyon, France: IARC Press, 2008 10. Crum NF: Epstein Barr virus hepatitis: case series and review. South Med J. 99(5):544-7, 2006 11. Johnson LR et al: Impact of Epstein-Barr virus in monomorphic B-cell posttransplant lymphoproliferative disorders: a histogenetic study. Am J Surg Pathol. 30(12):1604-12, 2006 12. Randhawa PS et al: Epstein-Barr virus-associated syndromes in immunosuppressed liver transplant recipients. Clinical profile and recognition on routine allograft biopsy. Am J Surg Pathol. 14(6):538-47, 1990

Epstein-Barr Virus, Liver

EBER In Situ Hybridization in EBV Hepatitis (Left) This case of EBV hepatitis shows sinusoidal lymphocytes ﬈ and a small granuloma ﬊. (Right) EBER in situ hybridization shows scattered positive lymphocytes st in a portal tract.

Monomorphic PTLD

Liver Transplantation

EBV Hepatitis With Granuloma

CD20 Stain in EBV-Associated Monomorphic PTLD (Left) Liver biopsy shows a case of EBV-associated monomorphic PTLD classified as Burkitt lymphoma. There is an atypical monomorphic population of lymphoid cells with tingible body macrophages in the background imparting a starry sky pattern. (Right) CD20 highlights neoplastic B cells. EBER in situ hybridization (not shown) was positive and the Ki-67 defined proliferation index was > 95%. (Courtesy J. LaPointe, MD.)

EBV-Associated PTLD

EBER In Situ Hybridization (Left) This case of EBVassociated PTLD shows a portal tract expanded by atypical lymphoid cells spilling out into surrounding parenchyma. (Right) EBER in situ hybridization is positive in numerous infiltrative lymphocytes ﬉. (Courtesy J. LaPointe, MD.)

299

Liver Transplantation

Fungal Infections, Liver KEY FACTS

ETIOLOGY/PATHOGENESIS • Candidiasis most common cause of fungal infection in transplant patients ○ Followed by aspergillosis (2nd) and cryptococcosis (3rd) • Non-albicans species emerging as predominant in some studies • Colonization of gastrointestinal tract and translocation to extraluminal areas

CLINICAL ISSUES • Risk factors include ○ Retransplantation ○ Cytomegalovirus infection ○ Multiple blood transfusions ○ Colonization with Candida • High mortality often due to delayed diagnosis and treatment • Prophylaxis may reduce incidence of fungal infections ○ Has not improved overall mortality

• Histologic identification of fungal forms on tissue or PCR performed on tissue block • Most common in first 30 days

MICROSCOPIC • Liver necrosis • Mixture of budding yeasts, pseudohyphae, and hyphae in candidiasis • Branching septate hyphae in aspergillosis • GMS, PAS positivity

ANCILLARY TESTS • PCR to confirm type of organism or detect fungal DNA from clinical specimen

Liver Biopsy With Fungal Infection

GMS Special Stain Liver Biopsy

Histoplasmosis

GMS Special Stain in Histoplasmosis Case

(Left) Candida albicans budding yeast forms ﬇ are present in this liver biopsy from a liver transplant patient. In this case, minimal inflammation is present. (Right) Special stain demonstrates numerous budding yeast forms st.

(Left) In this case of histoplasmosis, a portal tract demonstrates a compact, noncaseating granuloma with surrounding mild lymphocytic inflammation. (Courtesy J. Lin, MD.) (Right) Gomori methenamine silver stain highlights small round yeast forms st, measuring 3-5 μm in diameter, which is consistent with histoplasmosis.

300

Fungal Infections, Liver

Abbreviations • Invasive fungal infection (IFI)

ETIOLOGY/PATHOGENESIS Infectious Agents • Candida species most common (60-80%) ○ Candida albicans > Candida glabrata > Candida tropicalis • Aspergillus species (10-15%) • Cryptococcus neoformans (10-15%) • Histoplasma capsulatum (< 5%)

Risk Factors • Immunosuppression renders transplant recipients susceptible to primary IFI • Cytomegalovirus infection, colonization with Candida • Prolonged preoperative hospitalization and prolonged operative time

Mechanisms of Infection • Colonization of gastrointestinal tract and translocation to extraluminal areas • Inhalation of aerosolized spores from environment

CLINICAL ISSUES Epidemiology • 5-42% of patients develop fungal infection after transplantation • Most common in first 30 days ○ Recent data suggests shift toward IFI occurring > 90 days after transplantation • Modern surgical and medical techniques and practices have reduced risk of Candida infection but not Aspergillus

Presentation • Fatigue and malaise, along with subtle nonspecific symptoms, often delay diagnosis • Nonspecific liver enzyme elevations can be noted

Treatment • Prophylaxis of high-risk patients with antifungal therapy may be considered ○ Primary prophylaxis with fluconazole or other antifungal agents may shift infections toward non-Candida species ○ May reduce incidence of fungal infections – Has not improved mortality – Some reports do not show reduction in fungal infection even with prophylaxis • Aspergillus, Cryptococcus, and Histoplasma treated with extended duration of amphotericin &/or itraconazole or caspofungin

Prognosis • Mortality of 25-70% • Aspergillosis has mortality approaching 100% in late diagnosis or untreated patients

Diagnosis

• • •

•

Liver Transplantation

○ Difficult to distinguish between infection and colonization, especially if sample obtained from nonsterile sites Culture and microscopic analysis of Candida and Cryptococcus Cryptococcal serum antigen and Histoplasma urine antigen or serum have limited sensitivity Cross-sectional imaging, such as CT of chest and abdomen, help identify Aspergillus infection ○ Also Aspergillus PCR, galactomannan, or β-glucan test Histologic identification of fungal forms on tissue or PCR performed on tissue block or other clinical specimen

TERMINOLOGY

MICROSCOPIC Histologic Features • Candidiasis ○ Candida albicans most common ○ Can present as – Intraabdominal abscess – Biliary strictures with recurrent cholangitis – Peritonitis – Hepatic artery thrombosis ○ Budding yeast, hyphae, and pseudohyphae (but C. glabrata is yeast only) ○ Liver necrosis associated with fungal forms ○ Cholestasis may be present ○ Bilomas subsequent to bile infarcts from hepatic artery stenoses may be infected with Candida • Aspergillosis ○ 2nd most common cause of fungal infection in transplant patients ○ Aspergilloma characterized by central necrosis surrounded by giant cell reaction and fibrotic wall ○ Branching septate fungal hyphae • Cryptococcosis ○ Cryptococcus is 3rd most common cause of fungal infection in transplant patients ○ Round yeasts with narrow-based buds exhibiting great variation in size (2-20 μm)

ANCILLARY TESTS PCR • To confirm type of organism or detect fungal DNA from clinical specimen

Special Stains • GMS and PAS positivity

SELECTED REFERENCES 1.

2. 3. 4.

Raghuram A et al: Invasive fungal infections following liver transplantation: incidence, risk factors, survival, and impact of fluconazole-resistant Candida parapsilosis (2003-2007). Liver Transpl. 18(9):1100-9, 2012 Liu X et al: Invasive fungal infections in liver transplantation. Int J Infect Dis. 15(5):e298-304, 2011 Rosenhagen M et al: A risk profile for invasive aspergillosis in liver transplant recipients. Infection. 37(4):313-9, 2009 Fung JJ: Fungal infection in liver transplantation. Transpl Infect Dis. 4 Suppl 3:18-23, 2002

• Difficult laboratory isolation of fungi due to slow growth of pathogens and contamination of fungal organisms in environment 301

Liver Transplantation

Plasma Cell-Rich Rejection KEY FACTS

TERMINOLOGY • Form of immune-mediated graft injury ○ With prominent plasma cell infiltrates ○ In patients with no history of autoimmune hepatitis in native liver • Includes entities formerly described as ○ "De novo autoimmune hepatitis" ○ "Posttransplant plasma cell hepatitis" • By definition, affected patients have no history of autoimmune hepatitis in native liver

CLINICAL ISSUES • Treated with heightened immunosuppression

MICROSCOPIC • Plasma cell-rich portal &/or perivenular inflammatory cell infiltrates ○ Estimated > 30% plasma cells

• Easily recognizable periportal/interface &/or perivenular necroinflammatory activity

ANCILLARY TESTS • Autoantibodies reported in 20-74% of pediatric recipients and 60-70% of adult recipients

DIAGNOSTIC CHECKLIST • (1) Portal &/or perivenular plasma cell-rich (estimated > 30%) infiltrates ○ With easily recognizable periportal/interface activity &/or perivenular necroinflammatory activity – Usually involving majority of portal tracts &/or central veins • (2) Lymphocytic cholangitis often present • (3) Original liver disease other than autoimmune hepatitis • Criteria 1 and 3 above required for diagnosis

Plasma Cell-Rich Portal Infiltrates

Perivenular Inflammation and Injury

Plasma Cell-Rich Portal Infiltrates

Plasma Cells and Hepatocyte Necrosis

(Left) H&E shows dense portal inflammatory cell infiltrates with numerous plasma cells and interface activity in plasma cell-rich rejection. (Right) H&E shows centrilobular perivenular hepatocyte injury ſt and plasma cell-rich ﬉ perivenular inflammation in this patient with plasma cell-rich rejection.

(Left) H&E demonstrates plasma cell-rich portal infiltrates associated with periportal interface activity ﬉ in a liver transplant recipient with plasma cell-rich rejection. (Right) H&E shows parenchymal necrosis ﬊ associated with clusters of plasma cells ﬈ in this liver transplant recipient with plasma cell-rich rejection.

302

Plasma Cell-Rich Rejection

Synonyms

• Easily recognizable periportal/interface &/or perivenular necroinflammatory activity • Lymphocytic cholangitis usually present

• Posttransplant plasma cell hepatitis • De novo autoimmune hepatitis

ANCILLARY TESTS

Definitions

Autoantibody Titers

• Includes entities formerly described as ○ "De novo autoimmune hepatitis" ○ "Posttransplant plasma cell hepatitis" • Form of immune-mediated graft injury with prominent plasma cell infiltrates • Affected patients have no history of autoimmune hepatitis in native liver

• • • •

ETIOLOGY/PATHOGENESIS Immune Mediated • Considered pattern of allograft rejection • Pathogenesis not well understood • Exhibits features of acute T-cell-mediated rejection, antibody-mediated rejection, and autoimmunity ○ Patients may exhibit donor-specific antibodies and portal microvascular C4d deposition • Associated with antibodies to glutathione S-transferase theta 1

CLINICAL ISSUES Epidemiology • Incidence ○ 3-5% of liver transplant recipients

Presentation • Usually late (> 6 months) after transplant • Often with biochemical evidence of graft dysfunction ○ Elevated transaminases • May exhibit features similar to T-cell-mediated rejection ○ Fever ○ Fatigue ○ Graft tenderness ○ Hepatomegaly • Some patients asymptomatic

Treatment • Treated with heightened immunosuppression ○ Corticosteroids ± azathioprine

Prognosis • Prognosis good, if treated • Untreated patients may develop picture of chronic rejection ○ Can lead to portal hypertension, graft failure, and death

MICROSCOPIC

20-74% of pediatric recipients 60-70% of adult recipients Prevalence increases with posttransplant time interval Anti-smooth muscle antibodies most common ○ Others may also be present – Antinuclear antibodies – Antimitochondrial antibodies – Antiliver kidney microsome antibodies

Other Tests • Hypergammaglobulinemia commonly present

DIFFERENTIAL DIAGNOSIS Recurrent Autoimmune Hepatitis • Distinction based on history of autoimmune hepatitis in native liver

T-Cell-Mediated Rejection • More mixed portal infiltrates with fewer plasma cells • Tends to occur earlier post transplant (first 6 months)

Chronic Hepatitis • Recurrent or de novo hepatitis B and hepatitis C virus infection should be excluded

Other Infections • Hepatitis A virus and cytomegalovirus infection can also exhibit prominent plasma cell infiltrates

DIAGNOSTIC CHECKLIST Criteria for Diagnosis • (1) Portal &/or perivenular plasma cell-rich (estimated > 30%) infiltrates ○ With easily recognizable periportal/interface activity &/or perivenular necroinflammatory activity – Usually involving majority of portal tracts &/or central veins • (2) Lymphocytic cholangitis ○ Desirable but not absolutely required for diagnosis • (3) Original liver disease other than autoimmune hepatitis • Criteria 1 and 3 above required for diagnosis

SELECTED REFERENCES 1.

2.

Histologic Features • Plasma cell-rich portal &/or perivenular inflammatory cell infiltrates ○ Plasma cells estimated to comprise > 30% of inflammatory cells ○ Involves majority of portal tracts &/or central veins

Liver Transplantation

TERMINOLOGY

3.

4.

Demetris AJ et al: 2016 comprehensive update of the Banff working group on liver allograft pathology: introduction of antibody-mediated rejection. Am J Transplant. 16(10):2816-2835, 2016 Castillo-Rama M et al: "Plasma cell hepatitis" in liver allografts: identification and characterization of an IgG4-rich cohort. Am J Transplant. 13(11):296677, 2013 Fiel MI et al: Plasma cell hepatitis (de-novo autoimmune hepatitis) developing post liver transplantation. Curr Opin Organ Transplant. 17(3):287-92, 2012 Ward SC et al: Plasma cell hepatitis in hepatitis C virus patients post-liver transplantation: case-control study showing poor outcome and predictive features in the liver explant. Liver Transpl. 15(12):1826-33, 2009

303

Liver Transplantation

Graft-vs.-Host Disease, Liver KEY FACTS

TERMINOLOGY

• Immunosuppressed recipient cannot destroy donor cells

• Attack of immunocompetent, donor-derived T lymphocytes against recipient tissues • Usually occurs in hematopoietic cell transplant recipients ○ Infrequent after solid organ transplant and rare after blood transfusion • Acute and chronic forms of graft-vs.-host disease (GVHD) defined clinically • "Late-onset/persistent acute" GVHD occurs > 100 days after transplant in absence of chronic GVHD • "Classic chronic" GVHD: > 100 days after transplant • "Overlap syndrome": Coexisting acute and chronic GVHD features • No clear dichotomy between acute and chronic disease based on liver histology

CLINICAL ISSUES

ETIOLOGY/PATHOGENESIS • Donor-derived T-lymphocyte response against immunocompromised host epithelium

• Represents major hepatic complication after stem cell transplant

MICROSCOPIC • Bile duct epithelial cell damage ○ Cytoplasmic vacuolization and attenuation ○ Nuclear enlargement and pleomorphism ○ Sparse lymphocytic infiltration ○ Duct loss occurs with progression to chronic disease • Endotheliitis in some cases • Nonspecific changes including portal or lobular inflammation, cholestasis, or hepatocyte swelling • Acute hepatitis pattern seen in up to 25% of patients ○ Prominent lobular inflammation and denser portal infiltrates • Fibrosis with chronic disease

Bile Duct Damage

Bile Duct Abnormalities

Portal Inflammation

Cholestasis

(Left) H&E shows sparse portal inflammatory cell infiltration ﬈ and bile duct damage in graft-vs.-host disease (GVHD). The epithelial cells are irregular, unevenly spaced ſt, and show vacuolization ﬊. The duct lumen is irregular. (Right) H&E demonstrates portal features of GVHD. The portal inflammation is mild. One bile duct shows biliary epithelial cell abnormalities, including nuclear pleomorphism, loss of polarity, and uneven spacing ﬈.

(Left) H&E shows mild portal inflammation and biliary epithelial cell injury with uneven spacing of nuclei ﬊ and vacuolization ﬉. Iron overload ﬈ is often seen in stem cell transplant recipients. (Right) H&E shows nonspecific lobular changes seen in GVHD, including cholestasis ﬈, mild inflammation, and hepatocyte swelling.

304

Graft-vs.-Host Disease, Liver

Abbreviations • Graft-vs.-host disease (GVHD) • Hematopoietic cell transplantation (HCT)

Synonyms • Vanishing bile duct syndrome ○ Refers to loss of bile ducts in chronic GVHD

Prognosis • 50% of patients respond to treatment, although fewer achieve complete response • Mortality rates range from 20-75% • Duct loss is poor prognostic factor

MICROSCOPIC

Definitions

Histologic Features

• Attack of immunocompetent, donor-derived T lymphocytes against recipient tissues ○ Usually occurs in HCT recipients ○ Infrequent after solid organ transplant ○ Rarely occurs after blood transfusion • Acute and chronic forms of GVHD defined clinically ○ No clear dichotomy exists in liver histology ○ Classic acute GVHD occurs ○ Persistent or late-onset acute GVHD – Occurs > 100 days after transplant – Lacks features of chronic GVHD ○ "Classic chronic" GVHD can occur at any time ○ "Overlap syndrome" exhibits coexisting features of acute and chronic GVHD • Diagnostic categories have been defined by consensus ○ Not GVHD ○ Possible GVHD ○ Likely GVHD

• Bile duct epithelial cell damage ○ Nuclear enlargement and pleomorphism ○ Cytoplasmic vacuolization and attenuation ○ Sparse lymphocytic infiltration ○ Duct loss with progression to chronic disease • Endotheliitis described but less common • Nonspecific changes also seen ○ Portal or lobular inflammation, typically mild ○ Cholestasis, apoptosis, or hepatocyte swelling ○ Fibrosis with chronic disease • Acute hepatitis pattern in up to 25% of patients ○ Prominent lobular inflammation and denser portal infiltrates

ETIOLOGY/PATHOGENESIS Immune Mediated • Due to antigen incompatibility between host and recipient ○ Can occur with autologous grafts – Usually self-limited and responds well to treatment • Donor-derived T-lymphocyte response against host tissues, resulting in tissue injury • Immunosuppressed recipient cannot destroy donor cells

CLINICAL ISSUES Epidemiology • Major hepatic complication after HCT ○ Affects up to 70% of HCT recipients

Presentation • Jaundice • Elevated liver enzymes ○ Elevated serum alkaline phosphatase and bilirubin ○ Isolated elevated transaminases may also be seen • May exhibit manifestations of skin or gastrointestinal tract involvement ○ Rash, diarrhea, weight loss • Chronic GVHD often presents with widespread wasting disease ○ Salivary gland, oral, ocular, and musculoskeletal involvement • Hepatic failure and coagulopathy with advanced disease

Treatment

Liver Transplantation

○ Systemic corticosteroids are mainstay therapy

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Drug-Induced Liver Injury • Cyclosporine causes mild hyperbilirubinemia by inhibiting canalicular bile transport • Can cause elevated bilirubin but not usually biopsied

Cholangitis Lenta • Hyperbilirubinemia occurs in patients with neutropenia and fever • Attributed to hepatocyte retention of conjugated bilirubin • Affected patients may also be septic or have localized infections

Infections (Fungal, Bacterial, Viral) • In clinical differential but usually distinct histologically • Exclude with serologic testing, immunohistochemistry, or in situ hybridization

Biliary Obstruction • Excluded with imaging studies • Portal edema, neutrophilic infiltrate, and bile infarcts favor obstruction • Bile duct proliferation is not usually prominent in GVHD

Posttransplant Lymphoproliferative Disorder • More prominent lymphocytic infiltrates

SELECTED REFERENCES 1. 2. 3.

4.

Matsukuma KE et al: Diagnosis and differential diagnosis of hepatic graft versus host disease (GVHD). J Gastrointest Oncol. 7(Suppl 1):S21-31, 2016 Salomao M et al: Histopathology of graft-vs-host disease of gastrointestinal tract and liver: an update. Am J Clin Pathol. 145(5):591-603, 2016 Shulman HM et al: NIH Consensus development project on criteria for clinical trials in chronic graft-versus-host disease: II. The 2014 Pathology Working Group Report. Biol Blood Marrow Transplant. 21(4):589-603, 2015 McDonald GB: Hepatobiliary complications of hematopoietic cell transplantation, 40 years on. Hepatology. 51(4):1450-60, 2010

• Drugs 305

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SECTION 6

Heart Transplantation

Pathologic Classification of Cardiac Allograft Diseases Evaluation of Failed Native and Transplanted Heart History of Heart Transplantation

308 310 312

Evaluation of Explanted Heart Ischemic Heart Disease Dilated Cardiomyopathy Hypertrophic Cardiomyopathy Congenital Heart Disease Sarcoidosis, Heart Arrhythmogenic Right Ventricular Cardiomyopathy Other Causes of End-Stage Heart Disease

314 318 322 326 330 332 336

Allograft Rejection Acute Cellular Rejection, Heart Antibody-Mediated Rejection, Heart Chronic Allograft Vasculopathy, Heart

342 346 348

Noninfectious Lesions 350 352

Quilty Lesions Site of Previous Biopsy

Infections Myocarditis

354

Heart Transplantation

Pathologic Classification of Cardiac Allograft Diseases

TERMINOLOGY Pathologic Classification • According to main etiology, postheart transplant diseases can be broadly classified into following groups and their respective differential diagnoses ○ Postoperative complications ○ Rejection ○ Infection ○ Posttransplant lymphoproliferative disorders (PTLD)

Abbreviations • • • •

Acute cellular rejection (ACR) Antibody-mediated rejection (AMR) Cardiac allograft vasculopathy (CAV) Quilty lesion ○ Noninvasive (QA) ○ Invasive (QB)

Definitions • ACR ○ Rejection of transplanted heart mediated by activated lymphocytes ○ May occur within days to years after transplantation • AMR ○ Rejection primarily targeting microvasculature and capillaries – Due to antibody formation or preexisting antibodies against graft • CAV ○ Rejection resulting in accelerated form of coronary artery disease ○ Characterized by uniform, circumferential intimal fibrosis • Quilty lesion ○ Nodular endocardial infiltrate of small, mature lymphocytes – May extend into underlying myocardium • Site of previous biopsy ○ Histologic changes in endomyocardium due to prior biopsy at same site • PTLD ○ Lymphoid proliferation that, in many cases, manifests as frank lymphoma arising in immunocompromised hosts

CLINICAL ISSUES Posttransplant Complications • Well-established endomyocardial biopsy protocols for surveillance after heart transplantation ○ ~ 13 biopsies done in 1st year – Most in first 3 months ○ Noninvasive approaches to diagnose both acute and chronic rejections being evaluated • Most common treatment issue is ACR • AMR diagnosed in < 5% of recipients ○ Clinical criteria include – Allograft function – Serum antibodies ○ Pathologic criteria still being defined

○ C4d &/or C3d staining by IHC or immunofluorescence appears most diagnostically useful based on current literature ○ Increases risk of developing chronic rejection (CAV) ○ Associated with decreased survival • CAV is most significant long-term complication ○ Diagnosed by imaging coronary arteries ○ Since pathologic changes of fibrosis are uniform and circumferential, diagnosis is difficult ○ Clinical presentation varies – Acute myocardial infarction – Recent onset of cardiac arrhythmias – Heart failure – Sudden death • PTLDs only rarely involve heart allograft ○ Occur more often in other parts of body (i.e., gastrointestinal tract, lymph nodes) ○ May be monoclonal or polyclonal ○ Almost all are EBV driven • Opportunistic infections rare in allograft; most commonly occur in lung ○ Bacterial – Hospital acquired – Community acquired – Opportunistic organisms (e.g., Nocardia) ○ Fungal – Pneumocystis jiroveci pneumonia □ Rare due to prophylaxis – Aspergillus pneumonia – Candidiasis – Mucormycosis – Disseminated histoplasmosis ○ Protozoal – Toxoplasmosis □ Rare, can involve transplanted heart ○ Viral – CMV □ Pneumonia □ Gastrointestinal infection – Herpes infection, rare – Human papilloma viral infections □ Verruca vulgaris; multiple skin lesions may occur □ Cervical infection; increased risk of dysplasia and carcinoma □ Oral and upper respiratory tract infections

Surgical Complications • Rare due to improvement in surgical techniques ○ Postoperative hemorrhage ○ Acute graft failure – More likely due to poor donor management ○ Wound infections ○ Mediastinitis ○ Anastomotic dehiscence – Early complication ○ Stenosis – Late complication

Recurrence of Native Disease • Rare in cardiac allografts

308

Pathologic Classification of Cardiac Allograft Diseases

PATHOLOGIC ISSUES Examination of Native Heart • Gross and microscopic findings vary according to primary disease process ○ Coronary artery disease ○ Dilated cardiomyopathy ○ Hypertrophic cardiomyopathy ○ Congenital heart disease – Typically, multiple surgeries performed before transplantation ○ Arrhythmogenic right ventricular cardiomyopathy ○ Sarcoidosis • Superimposed changes of prior therapy, such as ○ Coronary artery bypass grafts ○ Ventricular assist device placement

• •

• •

Examination of Posttransplant Biopsy • Most process all tissue for light microscopy ○ 5 levels with multiple sections on each slide ○ Slides 1, 3, and 5 stained by H&E ○ Slides 2 and 4 (on charged slides) saved for special stains, as needed – Some transplant centers routinely perform C4d IHC stain to evaluate for AMR – Others perform both C4d and C3d IHC • 1 or 2 fragments may be submitted for immunofluorescence, according to clinical indications ○ C4d &/or C3d by IF is very sensitive for AMR

SELECTED REFERENCES 1.

2. 3.

4. 5.

Pathologic Diagnoses on Posttransplant Biopsies • ~ 70% of surveillance biopsies show no diagnostic pathologic changes • Common pathologic findings include ○ ACR – Most common in first 6 months after transplantation but may occur at any time – Typically asymptomatic – Grading of rejection based on amount of lymphocytic infiltration and if there is associated myocyte damage – International Society of Heart and Lung Transplantation □ Most widely used grading system □ 1990 version has grades 0, 1A, 1B, 2, 3A, 3B, and 4 □ 2005 version has fewer grades to improve interobserver variability: Grades 0, 1R, 2R, and 3R (where R denotes "revised") □ Many prefer 1990 version as it provides more information and is easily translated into 2005 grades ○ Quilty lesion – Idiopathic lesion □ Currently thought to be clinically insignificant – In 1990 classification, divided into □ Noninvasive (Quilty A) □ Invasive (Quilty B)

Heart Transplantation

– In 2005 classification, this distinction omitted ○ Site of previous biopsy – Due to biopsy protocols, transplanted heart biopsied several times in first 3 months – Leads to same area being sampled – Changes associated with previous biopsy site include □ Fibrin □ Granulation tissue □ Minimal inflammation – Main differential diagnosis is ACR IHC for C4d helpful since H&E findings of AMR are nonspecific Infections and PTLD are exceedingly rare in transplanted heart ○ Involves other organs, such as – Lung (infection) – Lymphoid tissue (PTLD) Drug reaction may be seen as eosinophilic infiltrates in endomyocardium Chronic rejection cannot be diagnosed on endomyocardial biopsy ○ Mainly involves epicardial coronary arteries and their branches

○ Giant cell myocarditis – Often fatal ○ Amyloidosis ○ Sarcoidosis

6.

7.

8.

9.

10. 11.

12. 13.

Mavrogeni SI et al: Cardiac transplantation: towards a new noninvasive approach of cardiac allograft rejection. Expert Rev Cardiovasc Ther. 15(4):307-313, 2017 Thiene G et al: Diagnostic use of the endomyocardial biopsy: a consensus statement. Virchows Arch. 463(1):1-5, 2013 Kirk R et al: The Registry of the International Society for Heart and Lung Transplantation: fifteenth pediatric heart transplantation report--2012. J Heart Lung Transplant. 31(10):1065-72, 2012 Labarrere CA, Jaeger BR. Biomarkers of heart transplant rejection: the good, the bad, and the ugly! Transl Res. 159(4):238-51, 2012 Leone O et al: 2011 consensus statement on endomyocardial biopsy from the Association for European Cardiovascular Pathology and the Society for Cardiovascular Pathology. Cardiovasc Pathol. 21(4):245-74, 2012 Stehlik J et al: The Registry of the International Society for Heart and Lung Transplantation: 29th official adult heart transplant report--2012. J Heart Lung Transplant. 31(10):1052-64, 2012 Angelini A et al: A web-based pilot study of inter-pathologist reproducibility using the ISHLT 2004 working formulation for biopsy diagnosis of cardiac allograft rejection: the European experience. J Heart Lung Transplant. 30(11):1214-20, 2011 Berry GJ et al: The ISHLT working formulation for pathologic diagnosis of antibody-mediated rejection in heart transplantation: evolution and current status (2005-2011). J Heart Lung Transplant. 30(6):601-11, 2011 Mehra MR et al: International Society for Heart and Lung Transplantation working formulation of a standardized nomenclature for cardiac allograft vasculopathy-2010. J Heart Lung Transplant. 2010 Jul;29(7):717-27. Erratum in: J Heart Lung Transplant. 30(3):360, 2011 Crespo-Leiro MG et al: Heart transplantation. Curr Opin Organ Transplant. 15(5):633-8, 2010 Stewart S et al: Revision of the 1990 working formulation for the standardization of nomenclature in the diagnosis of heart rejection. J Heart Lung Transplant. 24(11):1710-20, 2005 Gallo P et al: Causes of late failure after heart transplantation: a ten-year survey. J Heart Lung Transplant. 16(11):1113-21, 1997 Billingham ME et al: A working formulation for the standardization of nomenclature in the diagnosis of heart and lung rejection: Heart Rejection Study Group. The International Society for Heart Transplantation. J Heart Transplant. 9(6):587-93, 1990

309

Heart Transplantation

Evaluation of Failed Native and Transplanted Heart

CLINICAL IMPLICATIONS Common Indications for Transplantation • Ischemic heart disease • Dilated cardiomyopathy • Chronic rejection is most common indication for retransplantation ○ Acute graft failure rare cause for retransplantation

Indications for Endomyocardial Biopsy • Consensus statement from American Heart Association, American College of Cardiology, and European Society of Cardiology recommends endomyocardial biopsy in limited number of clinical scenarios whereby results might affect heart failure treatment ○ Unexplained new onset heart failure of < 2 weeks duration with normal or dilated left ventricle ○ Unexplained new onset heart failure of 2 weeks to 3 months duration with dilated left ventricle and new arrhythmias or heart block or failure to respond to usual care • Endomyocardial biopsy considered "reasonable" in additional clinical scenarios ○ Unexplained heart failure > 3 months with dilated left ventricle and new arrhythmias or heart block or failure to respond to usual care – Relatively low sensitivity, but if sarcoid is identified, it may respond to steroids ○ Unexplained heart failure with suspected allergic reaction and eosinophilia ○ Unexplained heart failure associated with suspected anthracycline toxicity – Requires electron microscopy – Biopsy to decide on further chemotherapy; patients not in complete remission are not transplant candidates ○ Unexplained heart failure with restrictive cardiomyopathy – Assessment for amyloidosis, hemochromatosis, sarcoidosis, metabolic diseases, hypertrophic cardiomyopathy

○ Unexplained cardiomyopathy in children • Endomyocardial biopsy may be considered in following scenarios ○ Unexplained heart failure with dilated left ventricle for > 2 weeks without associated heart block or arrhythmias ○ Heart failure associated with unexplained hypertrophic cardiomyopathy ○ Suspected arrhythmogenic right ventricular dysplasia ○ Unexplained ventricular arrhythmias • Following guidelines, biopsy yields diagnosis and change in treatment course in 20-25% of patients

Evaluation of Explants • Excluding patients with ischemic heart disease, up to 30% of patients may be misdiagnosed prior to transplant without endomyocardial biopsy ○ Since sensitivity of biopsy for some diseases is quite low (e.g., lymphocytic myocarditis, sarcoidosis), even with biopsy, pretransplant diagnosis may be incorrect ○ Identification of alternative diagnoses beyond idiopathic dilated cardiomyopathy may have implications for posttransplant surveillance, treatment, prognosis, and evaluation of family members

MACROSCOPIC General Features • • • • •

Obtain clinical and procedural history Photograph heart while intact and after sectioning Weigh heart, which will be missing majority of atria Examine valves for thickening, calcifications, or vegetations Section and describe occlusion of any bypass grafts and left anterior descending, left circumflex, and right coronary arteries ○ Describe location and patency of any stents • Describe epicardial fat and any evidence of previous procedures ○ Normal epicardial fat runs along atrioventricular grooves and along course of coronary arteries • For ischemic heart disease, "bread-loaf" heart from apex toward base, stopping 1-2 cm below atrioventricular valves

Ischemic Heart Disease Cross Sections (Left) Cross sectioning ("breadloafing") the ventricles in ischemic heart disease shows the distribution of lesions, in this case a large, healed infarction in the left ventricular lateral and posterior wall st. (Right) This 4-chamber cut of an explanted heart is from a patient who was retransplanted for accelerated graft vasculopathy. The previous suture line ﬉ can be identified in the left atrium.

310

4-Chamber Sectioning Approach

Evaluation of Failed Native and Transplanted Heart ○ Disarray may be present in normal hearts at apex, in trabeculae, and at junction of septum with ventricular free wall – Disarray does not necessarily indicate hypertrophic cardiomyopathy Size of myocytes is best assessed by nuclear size Assess presence or absence of inflammation ○ Type of inflammatory cells present ○ Presence or absence of associated myocyte damage Presence or absence of fibrosis ○ Reported as interstitial or replacement type ○ May be highlighted by trichrome Infiltrative diseases should be assessed with appropriate special stains Any presence of epicardium should be reported because of risk for cardiac tamponade ○ Fat may normally be present throughout right ventricle wall – Large nerves in fat are suggestive but not diagnostic of epicardium ○ Mesothelial cells are diagnostic of epicardium and positive by immunohistochemistry for – Nuclear WT1 – Calretinin – Cytokeratins

• •

•

• •

Explant of Transplanted Hearts

Evaluation of Explanted Heart

• For patients undergoing retransplant, usual cause of heart failure is accelerated vasculopathy ○ Additional sections of coronary arteries should be submitted to document extent of disease • For retransplanted heart or those with previous procedures, evaluate integrity of suture lines

• If biopsy was not performed, many of same special stains (amyloid, iron) may be performed if indicated

Specimen Handling • Biopsy ○ Distribution of tissue in appropriate fixatives is performed by cardiologist – Depending on indication, ≥ 3 but ideally > 5 fragments submitted in formalin for light microscopy – For possible infiltrative or metabolic diseases, ≥ 1 piece submitted in glutaraldehyde for EM – Additional tissue may be frozen or placed in Zeus solution for immunofluorescence or frozen for viral genomic testing, but these are less common – Genomic testing for some viruses can be performed on paraffin-embedded tissue ○ ≥ 3 H&E levels should be evaluated – Deeper levels may be ordered to evaluate patchy disease processes such as myocarditis ○ Additional stains are added as needed for – Amyloid (Congo red, sulfated Alcian blue) – Iron (Prussian blue) – Glycogen (PAS) – Collagen (trichrome) – Inflammatory cells (CD3, CD68)

Heart Transplantation

○ Remainder of heart opened along lines of blood flow • For cardiomyopathy, after determining that coronary arteries are not involved, heart may be opened in 4chamber cut • Measure thickness of left ventricular (LV) and right ventricular (RV) free wall and septum ○ Exclude epicardial fat and trabecular muscle • Describe chambers as normal, dilated, or restricted ○ Measure maximum LV chamber diameter excluding papillary muscles • Measure circumference of all valves • Measure and describe location and consistency of any myocardial lesions • Describe any endocardial fibrosis or thrombi including atrial appendages • Sections submitted ○ Representative sections of most occluded areas of each main coronary artery, often following decalcification ○ Any focal lesions (such as infarcts) as well as representative sections of uninvolved heart ○ For cardiomyopathy, ≥ 3 (many recommend 8-10) sections of left ventricle including anterior, posterior, lateral, and septal sections from both near base and apex ○ ≥ 2 sections each of RV and septum ○ Representative sections of any abnormal valves

SELECTED REFERENCES 1.

Rossano JW et al: The Registry of the International Society for Heart and Lung Transplantation: Nineteenth Pediatric Heart Transplantation Report2016; Focus Theme: Primary Diagnostic Indications for Transplant. J Heart Lung Transplant. 35(10):1185-1195, 2016 2. Bennett MK et al: Evaluation of the role of endomyocardial biopsy in 851 patients with unexplained heart failure from 2000-2009. Circ Heart Fail. 6(4):676-84, 2013 3. Mehra LM et al: Preponderance and implications of etiologic misclassification in advanced heart failure: a clinical-pathologic investigation. J Heart Lung Transplant. 32(2):268-9, 2013 4. Thiene G et al: Diagnostic use of the endomyocardial biopsy: a consensus statement. Virchows Arch. 463(1):1-5, 2013 5. Stone JR et al: Recommendations for processing cardiovascular surgical pathology specimens: a consensus statement from the Standards and Definitions Committee of the Society for Cardiovascular Pathology and the Association for European Cardiovascular Pathology. Cardiovasc Pathol. 21(1):2-16, 2012 6. From AM et al: Current status of endomyocardial biopsy. Mayo Clin Proc. 86(11):1095-102, 2011 7. Luk A et al: Do clinical diagnoses correlate with pathological diagnoses in cardiac transplant patients? The importance of endomyocardial biopsy. Can J Cardiol. 25(2):e48-54, 2009 8. Cooper LT et al: The role of endomyocardial biopsy in the management of cardiovascular disease: a scientific statement from the American Heart Association, the American College of Cardiology, and the European Society of Cardiology. Circulation. 116(19):2216-33, 2007 9. Ardehali H et al: Diagnostic approach to the patient with cardiomyopathy: whom to biopsy. Am Heart J. 149(1):7-12, 2005 10. Society for Cardiovascular Pathology Heart Dissection videos. Published 2011. Reviewed February 14, 2018. Accessed April 16, 2018.

MICROSCOPIC Evaluation of Biopsy • Arrangement of cardiac myocytes ○ Normally arranged in parallel arrays 311

Heart Transplantation

History of Heart Transplantation

TERMINOLOGY Definitions • Cardiac allograft: Transplanting heart from donor of same species as recipient but with different genotype • Cardiac xenograft: Transplanting heart from donor of one species into recipient of another species (cross species)

CHRONOLOGY AND EVOLUTION Timeline • Spans ~ 100 years • Remarkably rapid progress in research of surgical techniques and development of immunosuppressive drugs and their clinical application

Research Era (~ 1900-1960s) • Late 1890s ○ Dr. Alexis Carrel (Chicago, Illinois) – Known as father of vascular and transplant surgery – Developed technique of vascular anastomosis without constricting lumen or causing thrombosis □ Crucial advance that paved way for solid organ transplantation • 1905 ○ Dr. Alexis Carrel – Together with Charles Guthrie performed 1st experimental cardiac transplantation in dogs – Recognized difference in survival times between autografts and allografts (but did not attribute this to rejection) – Developed tissue preservation techniques and usage of vein grafts in arterial system – Recognized with Nobel Prize in Physiology and Medicine in 1912 • 1933 ○ Dr. Frank Mann (Mayo Clinic) – First to describe pathologic changes of rejection in transplanted hearts as impressive leukocytic infiltration of myocardium – Attributed changes to biological incompatibility between donor and recipient, thus proposing concept of cardiac allograft rejection to explain graft failure • 1956 ○ Dr. Peter Medawar (Birmingham, UK) – First to describe immune system as responsible for rejection of transplanted organs – Inoculation of mouse embryos with cells of another mouse strain introduced concept of acquired immunologic tolerance • 1960 ○ Drs. Richard Lower and Norman Shumway (Stanford, California) – Developed biatrial heart transplant technique in dogs □ Rapidly adapted to humans and used successfully until 1990s – Described 1st strategy in providing graft protection by deep topical hypothermia and recipient protection through cardiopulmonary bypass with surface cooling • 1964 ○ Dr. James Hardy (Jackson, Mississippi) 312

– Performed 1st cardiac xenograft by transplanting chimp heart into 68-year-old man □ Patient survived for 90 minutes

Early Clinical Era (1960s-1980) • 1966 ○ Dr. Michael DeBakey (Houston, Texas) – 1st successful implant of booster pump as temporary assist device • 1967 ○ Dr. Christiaan Barnard (Cape Town, South Africa) – 1st successful human cardiac allograft; patient lived for 18 days □ 3 days later, Dr. Adrian Kantrowitz successfully transplanted anencephalic donor heart into 3week-old baby with tricuspid atresia and atrial communication • 1969 ○ Dr. Denton Cooley (Houston, Texas) – 1st total artificial heart implant in 47-year-old man □ Artificial heart used as bridge until donor heart was found 3 days later • 1973 ○ Dr. Philip Caves (Stanford, California) – Introduced 1st percutaneous transvenous endomyocardial biopsy technique □ Provided reliable means for monitoring allograft rejection □ However, throughout 1980s, pathologic classification and grading of rejection varied with each transplant center

Modern Era (1980-Present) • 1980 ○ Introduction of cyclosporine A for treating cardiac transplant rejection • 1981 ○ Dr. Bruce Reitz (Stanford, California) – 1st successful heart-lung allograft – Cyclosporine A used experimentally to combat rejection • 1982 ○ Dr. William DeVries (Salt Lake City, Utah) – Permanent total artificial heart (Jarvik-7) implanted into 61-year-old man □ Patient lived for 112 days • 1984 ○ Dr. Denton Cooley – Performed 1st successful pediatric heart transplant on 8-month-old girl who survived until 13 years of age ○ Surgeons at Columbia University performed cardiac allograft on 4-year-old boy – Received 2nd transplant in 1989 and continues to lead productive life ○ Dr. Leonard Bailey (Loma Linda, California) – Performed 1st infant cardiac xenograft by transplanting baboon heart into 12-day-old girl, who survived for nearly 1 month • 1986 ○ Dr. Leonard Bailey

History of Heart Transplantation

Year

Landmark Event

Pioneers

1905

1st experimental heart transplant in dogs

Dr. Alexis Carrel

1960

Biatrial heart transplantation technique adapted in humans; used until 1990s

Drs. Richard Lower and Norman Shumway

1964

1st cardiac xenograft: Chimp heart into human who survived for 90 minutes

Dr. James Hardy

1966

Successfully implant of booster pump as temporary assist device

Dr. Michael DeBakey

1967

1st successful human cardiac allograft

Dr. Christiaan Barnard

1973

Introduced transvenous endomyocardial biopsy for monitoring allograft rejection

Dr. Philip Caves

1982

1st permanent total artificial heart (Jarvik-7) implant

Dr. William DeVries

1990

Introduction of new grading system for cellular rejection and refining therapies

Dr. Margaret Billingham/ISHLT

1991

Introduction of bicaval technique of heart transplantation

Dr. Hans H. Sievers

2000

1st use of Jarvik-2000, a left ventricular assist device; patient lived for nearly 7 years

Dr. Robert Jarvik

2010

Gene-expression profiling of peripheral-blood specimens shown to correlate with the results of endomyocardial biopsy

Dr. Hannah Valantine

Heart Transplantation

Major Milestones in Heart Transplantation

ISHLT = International Society for Heart and Lung Transplantation.

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– 1st series of successful heart transplants in infants with hypoplastic left heart syndrome 1989 ○ 1st multiorgan (heart-liver-kidney) transplant into 26year-old woman who survived for 4 months 1990 ○ International Society for Heart and Lung Transplantation (ISHLT) – Developed grading system for cellular rejection – Established uniform criteria of various transplant pathologies – Refined communication and comparison of treatment regimens and outcomes between transplant centers 1991 ○ Dr. Hans H. Sievers – Introduction of bicaval technique for heart transplantation; offered significant short-term advantages over biatrial technique 2000 ○ Dr. Robert Jarvik (New York, New York) – 1st use of Jarvik-2000, left ventricular assist device, in 63-year-old man who led active life for nearly 7 years after implant 2004 ○ New grading scale commissioned by ISHLT to address challenges and inconsistencies in old grading system ○ Antibody-mediated rejection recognized as important factor in up to 10% of cardiac recipients 2010 ○ Stanford scientists demonstrate that gene-expression profiling of peripheral-blood specimens correlates with results of endomyocardial biopsy ISHLT Consensus Guidelines and Statements ○ 2000: Consensus statement on xenotransplantation in continued effort to address issue of donor shortage ○ 2014: Consensus statement on diagnosis and care of patients with primary graft dysfunction ○ 2016: Updates on listing criteria for heart transplantation

– Heart failure prognosis scores in combination with cardiopulmonary exercise test to determine prognosis and guide listing for transplantation – Retransplantation is indicated for those patients who develop significant coronary allograft vasculopathy (CAV) with refractory cardiac allograft dysfunction, without evidence of ongoing rejection ○ 2016: Consensus statement on definition of, diagnostic criteria for, and management of antibody-mediated rejection

TREATMENT ISSUES Ongoing Challenges • Growing population of patients with end-stage heart disease • National donor shortage: Mitigated by alternative surgical procedures for advanced disease, xenotransplantation, and use of suboptimal donor organs • Certain regulations regarding allograft nonutilization due to demographic factors do not necessarily predict inferior outcome

Surgical Techniques • With continuous advances in surgical techniques, postoperative complications have steadily decreased over last few decades • Similarly, better organ preservation and donor management also contribute to better outcomes

Immunosuppression • Multidrug therapies and more effective drugs have led to significant decline in incidence of clinically significant acute cellular rejection ○ Chronic rejection (CAV) remains most important limitation to long-term survival of heart transplant recipients

SELECTED REFERENCES 1.

Berry GJ et al: The ISHLT working formulation for pathologic diagnosis of antibody-mediated rejection in heart transplantation: evolution and current status (2005-2011). J Heart Lung Transplant. 30(6):601-11, 2011

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Heart Transplantation

Ischemic Heart Disease KEY FACTS

CLINICAL ISSUES • Most common cause of heart failure overall and 2nd most common etiology of heart failure in transplantation • 5-year survival post transplant (> 70%) is lower than nonischemic cardiomyopathy ○ Higher than congenital heart disease or valvular cardiomyopathy • Metabolic risk factors that increase risk for CAD are also independent risk factors for posttransplant mortality and decreased 5-year survival

MACROSCOPIC • Large areas of fibrosis often present and may be subendocardial or transmural, consistent with healed myocardial infarctions ○ Distribution corresponds to that of coronary artery lesions • Usually multifocal obstruction of multiple coronary arteries

○ Both gross and histologic examinations typically overestimate extent of stenosis due to remodeling of entire arterial wall and luminal collapse

MICROSCOPIC • In areas of old myocardial infarction, fibrosis replaces myocytes • Myocytes immediately subjacent to endocardium are typically spared due to oxygen supply received from ventricular blood • Patchy interstitial or replacement fibrosis and myocyte hypertrophy are often seen but not specific

TOP DIFFERENTIAL DIAGNOSES • Healed inflammation in other types of heart disease (e.g., myocarditis, sarcoidosis, etc.)

Subendocardial and Interventricular Septal Fibrosis

Healed Transmural Infarction

Ventricular Aneurysm

Ventricular Hypertrophy With Old Infarction

(Left) These transverse sections demonstrate endocardial fibrosis ﬉ at the apex and patchy subendocardial fibrosis in the anterior left ventricle ſt and interventricular septum ﬇. (Right) Cross section through the ventricles of an explanted heart demonstrates the resolution of a transmural myocardial infarction. There is loss of myocardium and marked thinning of the anterior left ventricle ſt.

(Left) One relatively common long-term complication of transmural myocardial infarction is an aneurysm ſt, which develops at the site of the infarction due to the intraventricular pressure against the weakened ventricular wall. (Right) Gross image demonstrates left ventricular and septal hypertrophy ﬈. There is an area with patchy subendocardial fibrosis centered on the interventricular septum, likely the site of a prior infarction ﬉.

314

Ischemic Heart Disease

Synonyms • Ischemic cardiomyopathy, coronary artery disease (CAD), coronary heart disease

Definitions • Heart failure due to CAD ○ Almost always due to atherosclerotic coronary arteries ○ Rarely due to anomalous coronary arteries or fibromuscular dysplasia • Includes stable and unstable angina, myocardial infarction, and sudden cardiac death

ETIOLOGY/PATHOGENESIS Clinical Risk Factors • Hypertension, diabetes mellitus, obesity, dyslipidemia, and physical inactivity • Family history of CAD is strong risk factor ○ Specific mechanisms for this polygenic trait not elucidated • Tobacco smoking

CLINICAL ISSUES Epidemiology • Incidence ○ Most common cause of heart failure overall ○ 2nd most common etiology in transplantation ○ Leading cause of death worldwide • Age ○ Typically presents in middle-aged to older adults ○ Incidence increases with age • Sex ○ More common in males overall, but risk increases for women after menopause

Treatment • Surgical approaches ○ Once medical management, percutaneous coronary balloon angioplasty with stenting, and coronary bypass surgery have failed, patients are eligible for orthotopic heart transplant

Prognosis • 5-year survival post transplant (> 70%) lower than nonischemic cardiomyopathy but higher than congenital heart disease or valvular cardiomyopathy • Metabolic risk factors that increase risk for CAD are also independent risk factors for posttransplant mortality and decreased 5-year survival

MACROSCOPIC General Features • Hypertrophy with left ventricular dilation • Large areas of fibrosis often present and may be subendocardial or transmural, consistent with healed myocardial infarctions • Often thinning and aneurysm formation as well as patchy endocardial fibrosis in areas of scar • Usually multifocal obstruction of coronary arteries

Heart Transplantation

○ Degree of vessel &/or graft occlusion should be determined during gross examination ○ Significant ischemia occurs with 75% reduction in crosssectional area (50% reduction in diameter) ○ Both gross and histologic examinations typically overestimate extent of stenosis (compared to angiographic methods) due to remodeling of entire arterial wall and luminal collapse

TERMINOLOGY

Sections to Be Submitted • Left main, left circumflex, and right coronary arteries sectioned every 2 mm (ideally after decalcification) ○ Areas of most severe occlusion submitted ○ Bypass grafts &/or stents, if present, should be evaluated • Submit areas of fibrosis, discoloration, and grossly uninvolved myocardium

MICROSCOPIC Histologic Features • Coronary arteries ○ Eccentric luminal obstruction due to atherosclerotic plaques (often calcified) – Simple plaques have overlying fibrous cap with subjacent extracellular lipid (cholesterol crystals), intracellular lipid (foamy macrophages), and inflammatory cells – Complex plaques may have multiple cores or solid fibrosis and be concentric ○ Recanalized thrombi may be present in either native coronary arteries or grafted vessels ○ Acute plaque changes (thrombosis, hemorrhage) typically not present in explanted specimens • In areas of old infarction, fibrosis replaces myocytes ○ Replacement fibrosis may be subendocardial or transmural ○ Myocytes immediately subjacent to endocardium typically spared due to oxygen supply received from ventricular blood • Cytoplasmic vacuolization ("myocytolysis") is sublethal ischemic change that can be either adjacent to infarcts or in subendocardium • Nonspecific changes in noninfarcted areas include interstitial fibrosis and myocyte hypertrophy

DIFFERENTIAL DIAGNOSIS Healed Inflammation in Other Types of Heart Disease (e.g., Myocarditis, Sarcoidosis, etc.) • Absence of prominent coronary artery occlusion • Location of fibrosis does not correspond to vascular distribution patterns

SELECTED REFERENCES 1. 2.

Kilic A et al: Orthotopic heart transplantation in patients with metabolic risk factors. Ann Thorac Surg. 93(3):718-24, 2012 Minicucci MF et al: Heart failure after myocardial infarction: clinical implications and treatment. Clin Cardiol. 34(7):410-4, 2011

315

Heart Transplantation

Ischemic Heart Disease

Coronary Artery Occlusion

Atherosclerotic Artery

Bypass Graft Organized Thrombus

Coronary Artery Plaque

Healed Myocardial Infarction

Cholesterol Clefts

(Left) Gross photograph of coronary arteries demonstrates complete occlusion in 1 part of the vessel ſt. The other section st has severe obstruction but a small, compressed lumen remains. (Right) Graphic of a coronary artery demonstrates the eccentric nature of atherosclerotic plaques. The lipid-laden central core ﬈ is filled with cholesterol crystals and inflammatory cells and is covered by a fibrous cap ﬊. The lumen ﬉ is severely occluded.

(Left) Low-power view of an arterial bypass graft demonstrates the end result of thrombus organization. There are multiple recanalized channels ﬈. This is similar to the recanalization pattern seen in native vessels. (Right) Low-power view of a coronary artery demonstrates an eccentric plaque covered by a fibrous cap ﬊. There is thinning and remodeling of the muscular wall below the plaque ﬈. The lumen ﬉ is collapsed, making it difficult to estimate the extent of occlusion.

(Left) Low-power view demonstrates a subendocardial ﬉ band of dense fibrosis ﬊, consistent with a healed infarction. The myocytes ﬈ immediately subjacent to the endocardium are spared due to the supply of oxygen from ventricular blood. (Right) High-power view from the center of an atherosclerotic plaque shows typical needle-shaped cholesterol clefts ﬈ (the crystals get dissolved due to processing), which are interspersed with foamy macrophages ﬉ and scattered inflammatory cells.

316

Ischemic Heart Disease

Atherosclerotic Plaque (Left) Low-power view of a coronary artery shows a thickened wall with marked intimal fibrosis ﬊ and an atherosclerotic plaque containing cholesterol clefts ﬈ and scattered chronic inflammation. Note the fibrotic cap ﬉. (Right) This section of a coronary vessel wall shows the lipid-laden core of an atherosclerotic plaque filled with abundant foamy histiocytes ﬈. Lumen is at the top right.

Organizing Infarction

Heart Transplantation

Atherosclerotic Plaque

Myocytolysis (Left) High-power view of a section from an explanted heart shows a microscopic focus ﬊ of an organizing myocardial infarction with granulation tissue and few lymphocytes. (Right) Highpower view shows extensive cytoplasmic vacuolation ﬈ of myocytes adjacent to an area of replacement fibrosis ﬊. These viable cells demonstrating "myocytolysis" are a common feature in ischemic heart disease, either adjacent to areas of fibrosis or in the subendocardium.

Endo- and Subendocardial Fibrosis

Myocyte Hypertrophy (Left) Low-power view demonstrates subendocardial fibrosis ﬉ with sparing of the myocytes ﬊ immediately subjacent to the endocardium. There is also endocardial fibrosis ﬈, a common but nonspecific finding in ischemic heart disease. (Right) Highpower view from an explanted heart with ischemic heart disease demonstrates myocyte hypertrophy and interstitial fibrosis ﬈. These findings are nonspecific and are frequently found in heart disease of other etiologies.

317

Heart Transplantation

Dilated Cardiomyopathy KEY FACTS

TERMINOLOGY

MACROSCOPIC

• Increased left ventricular cavity diameter and global systolic dysfunction in absence of ○ Hypertension ○ Valve disease ○ Coronary artery disease

• Cardiomegaly and ventricular dilation result in overall globoid appearance of heart • Left ventricular dilation can be assessed by measuring ventricular diameter • Right ventricular dilation may also occur ○ But not always present • Although ventricular hypertrophy occurs, dilation results in wall thickness that is normal or thin

ETIOLOGY/PATHOGENESIS • Up to 30% of patients have family history of cardiomyopathy • ~ 50% of cases of dilated cardiomyopathy have no known cause

CLINICAL ISSUES

MICROSCOPIC • Changes are nonspecific and include myocyte hypertrophy and interstitial fibrosis

• Nonischemic cardiomyopathy accounts for > 50% of adult heart transplants • Posttransplant 5- and 10-year survival rates better for dilated than for ischemic cardiomyopathy or congenital heart disease

Dilated Cardiomyopathy

Cross Section of Dilated Cardiomyopathy

Explanted Dilated Heart

Microscopic Nonspecific Findings

(Left) This explanted heart has a globoid shape. The left ſt and right ﬇ ventricles are dilated without an increase in wall thickness. There is increased trabeculation in the left ventricular wall. (Right) This autopsied heart with dilated cardiomyopathy has mild left ventricular dilation ﬇ with a left ventricular diameter > 4 cm.

(Left) This explanted heart with dilated cardiomyopathy has an overall rounded shape with particularly enlarged left ventricle. (Right) The microscopic findings in dilated cardiomyopathy are not specific. Frequently, there is interstitial fibrosis ﬈ and myocyte hypertrophy; however, the findings may be mild, especially in biopsies.

318

Dilated Cardiomyopathy

Abbreviations

– DCM often occurs with muscular dystrophy syndromes ○ Metabolic and mitochondrial disorders

• Dilated cardiomyopathy (DCM)

Peripartum Cardiomyopathy

Synonyms • Idiopathic DCM • Nonischemic cardiomyopathy

• Unclear relationship between pregnancy and DCM ○ May involve hormonal, inflammatory, familial, or hemodynamic factors

Definitions

Idiopathic

• DCM ○ Increased left ventricular cavity diameter and global systolic dysfunction in absence of hypertension, valve disease, or coronary artery disease ○ Identical pathological findings seen in various systemic diseases – Should be excluded clinically ○ Familial DCM defined as DCM in two 1st-degree family members ○ Peripartum defined as DCM developing between last month of pregnancy and 5 months after delivery

• ~ 50% of DCM have no known cause

Heart Transplantation

TERMINOLOGY

CLINICAL ISSUES Epidemiology • Incidence ○ Nonischemic cardiomyopathy accounts for > 50% of adult heart transplants • Age ○ Most common in young to middle-aged adults – Can be seen in children and elderly

Presentation

ETIOLOGY/PATHOGENESIS Environmental Exposure • Many factors associated with DCM, but unclear role in pathogenesis, and most have no specific findings ○ Heavy alcohol use ○ Chemotherapeutic agents, especially anthracyclines (doxorubicin, daunorubicin) – Risk related to cumulative dose – Presents months to years after treatment ○ Trace mineral deficits (selenium) or accumulation (arsenic, cobalt) ○ Hemochromatosis

Infectious Agents • Parvovirus B19, human herpesvirus 6, coxsackievirus, influenza virus, adenovirus, echovirus, cytomegalovirus, human immunodeficiency virus, Borrelia burgdorferi, and Corynebacterium diphtheriae ○ All associated with myocarditis ○ Can cause postinflammatory DCM • Chagas disease is leading cause of DCM in South America

Inflammatory • Connective tissue diseases, such as ○ Systemic lupus erythematosus ○ Scleroderma

Genetics • Up to 30% have family history of cardiomyopathy ○ Variable degree of dysfunction • Most commonly autosomal dominant transmission but any pattern may occur ○ Expressivity and penetrance highly variable • Mutations in > 50 different genes, and most are specific to given family ○ Mutations in sarcomere genes most common (35-40%) – Titin (TTN) gene most commonly mutated □ ~ 25% of familial cases ○ Mutations in sarcolemmal, cytoskeletal, and nuclear envelop proteins (lamin A/C) also common

• Heart failure most common • Arrhythmias common in some genetic subtypes (LMNA mutation)

Prognosis • Pretransplant survival depends on functional status and ejection fraction • Up to 45% of pediatric patients with DCM may regain normal function • Patients transplanted for peripartum DCM have higher rates of rejection and both shorter graft and overall survival

Complications • Common complications include arrhythmias and thromboembolic events due to hemostasis

IMAGING Echocardiography • Reduced or normal left ventricular wall thickness and left ventricular spherical dilatation, often with dilatation of other chambers • Impaired left ventricular contractile function

MACROSCOPIC General Features • Cardiomegaly present with ↑ heart weight relative to ageand size-matched controls • Ventricular dilation results in overall globoid appearance of heart (instead of pointed apex) • Left ventricular dilation can be assessed by measuring ventricular diameter (excluding papillary muscles) ○ Normal adult diameter < 4 cm • Right ventricular dilation may also occur ○ Normal adult circumference < 13 cm ○ Can be assessed by increase in tricuspid valve circumference • Although hypertrophy occurs, dilation of chambers results in normal or thin ventricular wall • Atria typically dilated 319

Heart Transplantation

Dilated Cardiomyopathy ○ Often absent in explanted specimens • Organizing thrombi can be seen due to hemostasis ○ May lead to patchy endocardial fibrosis • Patchy myocardial fibrosis may be present ○ Does not correspond to coronary artery obstruction

• Grossly visible ventricular septal and left ventricular scarring frequent in dilated stage of hypertrophic cardiomyopathy • Myofiber disarray present, particularly in ventricular septum if thoroughly sampled

Sections to Be Submitted

• Any iron deposition in heart is abnormal • Heaviest iron deposits seen in outer myocardium ○ May be absent in endomyocardial biopsies

• Coronary arteries to exclude ischemic heart disease • Left and right ventricles extensively sampled to assess hypertrophy, fibrosis, and inflammation • Interventricular septum well sampled to exclude end-stage hypertrophic cardiomyopathy

MICROSCOPIC Histologic Features • Biopsies typically to rule out other causes of cardiomyopathy, including myocarditis, amyloid, or hemochromatosis ○ Absence of inflammation, amyloid, and iron should be documented – Any iron accumulation is abnormal • Nonspecific changes include myocyte hypertrophy and interstitial fibrosis ○ Myocytes vary in size – Have enlarged, hyperchromatic nuclei without disarray – Multinucleation can be seen ○ Hypertrophy may be very mild or even absent in limited material – Does not correlate with prognosis • Increased lipofuscin pigment often present in myocytes • Small foci of lymphocytic infiltrates without myocyte necrosis can be seen

ANCILLARY TESTS Electron Microscopy • Performed to exclude other causes of DCM • Nonspecific changes associated with hypertrophy or degeneration often present, including ○ Myofibrillar disarray or loss ○ Dilated T-tubules ○ Z-band abnormalities ○ Mitochondrial abnormalities ○ Intercalated disc dehiscence • DCM due to anthracycline toxicity associated with ○ Vacuolation due to dilation of sarcoplasmic reticulum ○ Patchy disarray and dropout of myofibrils

DIFFERENTIAL DIAGNOSIS Ischemic Heart Disease • Also causes systolic dysfunction and left ventricular dilation ○ Has occlusion of coronary arteries, subendothelial fibrosis, or transmural infarctions • Fibrosis and infarctions in regions consistent with coronary artery distribution

Hypertrophic Cardiomyopathy • Patients can undergo dilated-hypokinetic evolution

320

Hemochromatosis

Myocarditis • Often clinical history of recent viral infection associated with imaging findings of DCM • Classically, foci of myocyte necrosis, but endomyocardial biopsy lacks sensitivity due to patchy nature ○ Viral pathogens detected by PCR in patients without histologic evidence of myocarditis ○ High interobserver variability even among expert pathologists in evaluating myocarditis

Arrhythmogenic Right Ventricular Cardiomyopathy • Often greater degree of arrhythmia than expected for DCM • Subepicardial distribution of fibrosis • Some left ventricle predominant or biventricular forms of arrhythmogenic right ventricular cardiomyopathy (ARVC) with more fibrosis than fat ○ Can mimic DCM • Definitive distinction between DCM and ARVC not always possible on morphologic basis ○ Differential diagnosis should be raised for appropriate genetic testing

SELECTED REFERENCES 1. 2.

Lee TM et al: Pediatric cardiomyopathies. Circ Res. 121(7):855-873, 2017 McNally EM et al: Dilated cardiomyopathy: genetic determinants and mechanisms. Circ Res. 121(7):731-748, 2017 3. Dipchand AI et al: The registry of the International Society for Heart and Lung Transplantation: eighteenth official pediatric heart transplantation report--2015; focus theme: early graft failure. J Heart Lung Transplant. 34(10):1233-43, 2015 4. Lund LH et al: The registry of the International Society for Heart and Lung Transplantation: thirty-second official adult heart transplantation report-2015; focus theme: early graft failure. J Heart Lung Transplant. 34(10):124454, 2015 5. McNally EM et al: Genetic mutations and mechanisms in dilated cardiomyopathy. J Clin Invest. 123(1):19-26, 2013 6. Herman DS et al: Truncations of titin causing dilated cardiomyopathy. N Engl J Med. 366(7):619-28, 2012 7. Pietra BA et al: Early predictors of survival to and after heart transplantation in children with dilated cardiomyopathy. Circulation. 126(9):1079-86, 2012 8. Rasmusson K et al: Peripartum cardiomyopathy: post-transplant outcomes from the United Network for Organ Sharing Database. J Heart Lung Transplant. 31(2):180-6, 2012 9. Matsumura Y et al: Long-term prognosis of dilated cardiomyopathy revisited: an improvement in survival over the past 20 years. Circ J. 70(4):37683, 2006 10. Felker GM et al: Underlying causes and long-term survival in patients with initially unexplained cardiomyopathy. N Engl J Med. 342(15):1077-84, 2000 11. Rose AG et al: Dilated (congestive) cardiomyopathy: a syndrome of severe cardiac dysfunction with remarkably few morphological features of myocardial damage. Histopathology. 9(4):367-79, 1985

Dilated Cardiomyopathy

Dilated Cardiomyopathy (Left) In this explanted heart, there is marked left ventricular dilation with extensive endocardial fibrosis ﬈ resulting in white, plaquelike thickening of the endocardial surface. (Right) The left ventricle has a thickened endocardium ﬈ with more prominent trabeculation ﬊ than normal.

Left Ventricular Thrombus

Heart Transplantation

Left Ventricular Endocardial Fibrosis

Dilated Cardiomyopathy With Fat (Left) On this close-up view of the left ventricular wall of a deceased patient with dilated cardiomyopathy, there is endocardial fibrosis ﬈ seen as a thickened white plaque, and a more recent adherent, organizing thrombus ﬇. (Right) High-power view of the left ventricle in a dilated cardiomyopathy case demonstrates the nonspecific changes typical of dilated cardiomyopathy with interstitial fibrosis ﬈ between myocytes, some of which have markedly enlarged nuclei ﬊. There is also focal fatty ﬉ infiltration and replacement.

Endocardial Fibrosis in Dilated Cardiomyopathy

Hemosiderin Deposition in Myocytes (Left) This section of left ventricular free wall from an explanted heart with dilated cardiomyopathy shows marked fibrosis of the endocardium ﬈ overlying the subjacent myocardium ﬊. (Right) In this section stained with Prussian blue, iron deposits ﬈ can be seen in some myocytes. Iron deposits are greatest in the outer section of the heart, and any amount of iron deposition is abnormal. Hemochromatosis should be ruled out in biopsies for dilated cardiomyopathy.

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Heart Transplantation

Hypertrophic Cardiomyopathy KEY FACTS

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• Most identified mutations involve genes coding for contractile proteins with cardiac myosin binding protein C (MYBPC3) and myosin heavy chain (MYH7) being most commonly affected

• Foci of myofiber disarray and myocyte hypertrophy required for diagnosis and may be quite focal (5% of section) or more extensive • Intramural coronary arteries often demonstrate intimal fibrosis or medial hypertrophy • ANCA

MACROSCOPIC • Left ventricular wall is thickened, often with septal prominence, but can be symmetric • Bulging of muscle into left ventricle below aortic valve may cause outflow obstruction ○ Abnormal contact of wall with anterior mitral valve leaflet leads to sharp, defined patch of endocardial fibrosis with mitral valve distortion and fibrosis • Multiple sections of ventricular septum should be taken perpendicular to long axis

TOP DIFFERENTIAL DIAGNOSES • Myocyte "disarray" occurs normally where cells meet at acute angles (junction of ventricular septum with anterior and posterior free walls and in trabeculations) and at edges of fibrosis • Multiple metabolic disorders may phenocopy hypertrophic cardiomyopathy and should be excluded, particularly in young patients

Gross Image Hypertrophic Cardiomyopathy

Myocyte Disarray

Intimal Fibrosis of Intramural Artery

Pediatric Hypertrophic Cardiomyopathy

(Left) In this explanted heart, the left ventricle (LV) wall is markedly thickened. Note the LV free wall ﬇ and anterior papillary muscle st. (Courtesy B.M. Shehata, MD.) (Right) In this microscopic section of hypertrophic cardiomyopathy, there is myocyte disarray, interstitial fibrosis, and myocyte hypertrophy with enlarged, hyperchromatic nuclei ﬈.

(Left) High-power view of an intramural artery demonstrates a markedly narrowed lumen due to intimal fibrosis ﬈. There is interstitial fibrosis ﬊ extending between the adjacent myocytes. (Right) Section of a heart from an infant with hypertrophic cardiomyopathy demonstrates myocyte disarray with intermixed bundles of cells running at diverse angles. Although small overall, the size of the myocytes is increased relative to that expected for an infant.

322

Hypertrophic Cardiomyopathy

Abbreviations • Hypertrophic cardiomyopathy (HCM)

Synonyms • Idiopathic hypertrophic subaortic stenosis

Definitions • Increased ventricular wall thickness ○ Not due to structural heart conditions (valvular or congenital heart disease), hypertension, or exercise ○ Some classifications also include metabolic disorders

ETIOLOGY/PATHOGENESIS

○ Ventricular septal myotomy or ethanol septal ablation can be performed in severe cases of outflow obstruction

Prognosis • Annual mortality rate < 1% due to sudden death, heart failure, or thromboembolic stroke

IMAGING MR Findings

Heart Transplantation

TERMINOLOGY

• Left ventricular hypertrophy and outflow abnormalities • May be used for familial screening

Echocardiography • Left ventricular hypertrophy (> 15 mm), often with small ventricular cavity

Genetic Basis • Mutations identified in ~ 50% of patients ○ Usually autosomal dominant but with variable expressivity and age-related penetrance • Most identified mutations involve genes coding for contractile proteins [cardiac troponins, myosin light chains, cardiac myosin binding protein-C (MYBPC3), cardiac myosin heavy chains, cardiac α-actin, α-tropomyosin, and titin] ○ MYBPC3 and β-myosin heavy chain (MYH7) most commonly affected genes ○ Unreliable correlation between genotype and phenotype

CLINICAL ISSUES Epidemiology • Incidence ○ Affects ~ 1 in 500 of general population but with variable severity ○ Most common cause of sudden death in children, young adults, and athletes • Age ○ Typically presents in adolescents or young adults but can be seen in infants or older adults

Presentation • Varies from chest pain, dyspnea, fatigue, or syncope to sudden cardiac death due to arrhythmias • Individuals identified by family screening who carry mutation may be asymptomatic with nonhypertrophic phenotype and absent or subtle changes on imaging

Natural History • Highly variable: Many patients have normal life spans ○ Most patients clinically stable for long periods • < 20% of patients demonstrate adverse remodeling, including systolic or diastolic dysfunction, aneurysms, or severe microvascular disease • ~ 5% progress to end-stage disease ○ Rapid deterioration ○ Features similar to dilated cardiomyopathy

MACROSCOPIC General Features • Thickened left ventricular wall ○ Often with septal prominence but can be symmetric ○ Left ventricular cavity may be small or normal ○ Wall thinning and left ventricular dilation can occur with late decompensation • Right ventricular hypertrophy uncommon • Bulging of septal muscle into left ventricle below aortic valve may cause outflow obstruction ○ Contact of septal wall with anterior mitral valve leaflet leads to sharply defined patch of endocardial fibrosis with mitral valve distortion and fibrosis – "Classic" feature seen in < 1/3 of cases

Size • Heart weight often significantly elevated

Sections to Be Submitted • Multiple sections of ventricular septum should be taken perpendicular to long axis • Routine sections of left ventricle and posterior right ventricle may also show disarray

MICROSCOPIC Histologic Features • Foci of myofiber disarray and myocyte hypertrophy required for diagnosis but not specific ○ May be quite focal (5% of section) or more extensive ○ Nuclear size and myocyte diameter highly variable in areas both with and without disarray ○ Type I disarray: Individual cells or bundles of myocytes arranged at oblique or perpendicular angles ○ Type II disarray: Transversely cut bundles of myocytes separated by thin, longitudinal bundles of myocytes • Fibrosis common ○ May be patchy and interstitial or transmural • Intramural coronary arteries often demonstrate intimal fibrosis or medial hypertrophy

Treatment • Options, risks, complications ○ Implantable defibrillators used to prevent sudden death in high-risk patients • Surgical approaches 323

Heart Transplantation

Hypertrophic Cardiomyopathy

ANCILLARY TESTS Genetic Testing • Testing for mutations in sarcomeric proteins may be indicated for genetic counseling but not required for diagnosis • ~ 50% of patients will not have identified mutation • Genetic testing for metabolic/storage disorders should be considered in young patients with severe manifestations

Electron Microscopy • May be helpful to evaluate storage diseases

Metabolic Testing • May be helpful to evaluate storage disease

DIFFERENTIAL DIAGNOSIS Secondary Forms of Hypertrophic Cardiomyopathy • More often show concentric hypertrophy and may have myocyte vacuolization • Extracardiac manifestations may be present • May require genetic, metabolic, or electron microscopy for diagnosis • Lysosomal storage diseases (Fabry disease, Hurler disease) ○ Fabry disease – X-linked deficiency of α-galactosidase A – Results in accumulation of glycosphingolipids in lysosomes – Electron microscopic lamellar bodies (Intracytoplasmic, concentric, electron-dense layered structures) but may be seen in other storage diseases • Glycogen storage disease (Pompe, PRKAG2, Danon) ○ LAMP-2 deficiency (Danon disease) – X-linked deficiency in lysosome-associated membrane protein-2 – Cardiomyopathy presents similarly to severe classic HCM – Electron microscopy shows autophagic vacuoles filled with glycogen, degenerating mitochondria and debris – Myopathy and intellectual disability often present ○ Protein kinase AMP-activated γ2 (PRKAG2) deficiency – Autosomal dominant inheritance of mutations in noncatalytic subunit – Usually accompanied by arrhythmias, particularly Wolff-Parkinson-White syndrome – Accumulation of cardiac glycogen as PAS-positive intracytoplasmic vacuoles – Absence of myofiber disarray or fibrosis – Accumulation of intracytoplasmic glycogen in vacuoles can be seen by electron microscopy • Mitochondrial cytopathies • Syndromic (Noonan syndrome, Leopard syndrome, Friedrich ataxia) • Fatty acid metabolic disorders

Hypertension-Associated Hypertrophy • Clinical history of hypertension • Typically presents with concentric left ventricular hypertrophy • Wall hypertrophy usually causes marked compromise of left ventricular cavity • May be some dilation of chamber if significant old infarcts or heart failure • Myocytes appear "disarrayed" in normal hearts at certain locations ○ Where cells meet at acute angles (junction of ventricular septum with anterior and posterior free walls) ○ Trabeculae ○ At edges of fibrosis

Dilated Cardiomyopathy • Subset of HCM patients may progress to dilated stage similar to dilated cardiomyopathy • Dilated HCM often has diffuse or transmural fibrosis instead of patchy, interstitial fibrosis typical of dilated cardiomyopathy • Dilated cardiomyopathy lacks myocyte disarray and involvement of intramural coronary arteries

Athlete's Heart • Within clinical differential diagnosis for hypertrophic changes but not seen in explanted hearts • More likely to have milder, concentric hypertrophy with mild left ventricular dilatation, but overlap exists • Myocytes may show "disarray" in normal hearts

Infants of Diabetic Mothers • Asymmetric hypertrophy and ventricular outflow obstruction may occur as part of generalized organomegaly • Clinically resolves during infancy

SELECTED REFERENCES 1. 2. 3.

4. 5. 6.

7. 8. 9. 10. 11.

Amyloidosis • Amyloid may deposit preferentially in base of ventricular septum • Interstitial amyloid surrounds individual myocytes • Congo red stain positive 324

12.

13.

Ellepola CD et al: Genetic testing in pediatric cardiomyopathy. Pediatr Cardiol. ePub, 2017 Lloyd DF et al: Cardiac manifestations of inherited metabolic disease in children. Pediatr Int. 59(5):525-529, 2017 Marian AJ et al: Hypertrophic cardiomyopathy: genetics, pathogenesis, clinical manifestations, diagnosis, and therapy. Circ Res. 121(7):749-770, 2017 Sen-Chowdhry S et al: Update on hypertrophic cardiomyopathy and a guide to the guidelines. Nat Rev Cardiol. 13(11):651-675, 2016 D'souza RS et al: Danon disease: clinical features, evaluation, and management. Circ Heart Fail. 7(5):843-9, 2014 Frustaci A et al: Diagnostic contribution of left ventricular endomyocardial biopsy in patients with clinical phenotype of hypertrophic cardiomyopathy. Hum Pathol. 44(1):133-41, 2013 Maron BJ et al: Genetics of hypertrophic cardiomyopathy after 20 years: clinical perspectives. J Am Coll Cardiol. 60(8):705-15, 2012 Frey N et al: Mechanisms of disease: hypertrophic cardiomyopathy. Nat Rev Cardiol. 9(2):91-100, 2011 Maron BJ et al: Clinical outcome and phenotypic expression in LAMP2 cardiomyopathy. JAMA. 301(12):1253-9, 2009 Hughes SE: The pathology of hypertrophic cardiomyopathy. Histopathology. 44(5):412-27, 2004 Lamke GT et al: Surgical pathology of subaortic septal myectomy associated with hypertrophic cardiomyopathy. A study of 204 cases (1996-2000). Cardiovasc Pathol. 12(3):149-58, 2003 Nishino I et al: Primary LAMP-2 deficiency causes X-linked vacuolar cardiomyopathy and myopathy (Danon disease). Nature. 406(6798):906-10, 2000 Davies MJ et al: Hypertrophic cardiomyopathy--pathology and pathogenesis. Histopathology. 26(6):493-500, 1995

Hypertrophic Cardiomyopathy

Alternating Myocyte Orientation (Left) Low-power view of hypertrophic cardiomyopathy demonstrates both myocyte disarray and interstitial fibrosis. (Right) High-power photomicrograph demonstrates myocyte disarray. Although the majority of the fibers are cut transversely ﬈, intervening myocytes are cut longitudinally ﬊.

Intimal Fibrosis and Myocyte Disarray

Heart Transplantation

Myocyte Disarray and Interstitial Fibrosis

Myocyte Disarray (Left) Low-power view demonstrates both myocyte disarray and microvascular changes. The intramural artery ﬈ has a thickened wall. There is interstitial fibrosis, and the myocytes are arranged in haphazard bundles with interstitial fibrosis. (Right) High-power view of a teenager with hypertrophic cardiomyopathy shows enlarged nuclei, mild interstitial fibrosis, and disarray ﬈

Myocyte Disarray

Danon Disease (Left) Microscopic section from a 9-week-old infant with sarcomeric gene mutations shows intersecting bundles of myocytes. Although cells appear small, the nuclei are enlarged compared to expected for an infant of this age. (Right) In this hypertrophic heart from a patient with Danon disease, enlarged myocyte nuclei ﬈ can be seen, and many of the myocytes are vacuolated.

325

Heart Transplantation

Congenital Heart Disease KEY FACTS

CLINICAL ISSUES • Indications for transplant include severe heart failure, growth failure or severe activity limitation due to heart failure, intractable arrhythmias, and risk of irreversible pulmonary vascular disease • Posttransplant outcomes are poorer than for cardiomyopathies or ischemic heart disease • Highest risk in 1st year after transplant • Most common types requiring transplantation are single ventricle lesions, D- or L-transposition of great arteries, and right ventricle (RV) outflow tract lesions (e.g., tetralogy of Fallot) • 56% of transplants in infants < 1 year • 3% of adult heart transplants • Most common types of CHD that require transplantation

• Transposition of great arteries: Aorta arises from morphologic RV and pulmonary artery (PA) from morphologic LV ○ Dextro-transposition of great arteries: Aorta arises anterior and right of PA • Congenitally corrected-transposition of great arteries: Aorta arises anterior and left of PA ○ Noncyanotic: Oxygenated blood flows from LA to leftsided RV to aorta • In explanted hearts, absence of majority of atria and great vessels as well as previous procedures may make certain abnormalities impossible to recognize without clinical history • Ventricular structure is determined, and AV valves are described as normal, stenotic, imperforate, or straddling

MICROSCOPIC

MACROSCOPIC • Tetralogy of Fallot: Pulmonary stenosis, large ventricular septal defect overriding aorta, RV hypertrophy

• Features are nonspecific and include myocyte hypertrophy, interstitial fibrosis, and endocardial fibrosis

Tetralogy of Fallot

CC-TGA

D-TGA

Mustard-Senning Repair of D-TGA

(Left) In this diagram showing tetralogy of Fallot, there is pulmonary stenosis ſt, a ventricular septal defect (VSD) ﬇ with an overriding aorta st, and consequent right ventricle (RV) hypertrophy ﬈. (Right) In congenitally corrected TGA, the aorta ſt is to the left of the pulmonary artery (PA) st and arises from the morphologic RV (coarse trabeculations) ﬊. Because the morphologic RV is on the left and connected to the left atrium (LA), this is acyanotic ("congenitally corrected"). This heart has an associated VSD ﬇.

(Left) In D-TGA, the aorta ſt arises anterior and to the right of the PA st, and the morphologic RV ﬉ is on the right. This is a cyanotic form of CHD: Mixing must occur through an ASD ﬇, VSD (not shown), or PDA ﬈ to allow survival. (Right) In this "atrial switch" operation, a baffle ﬉ diverts blood from the vena cava to the LA ﬊. Deoxygenated blood then flows to the PA ﬈ via the left ventricle (LV). The pulmonary veins are redirected by a baffle ﬇ to the right atrium (RA), where oxygenated blood flows to the RV and aorta ſt.

326

Congenital Heart Disease

Abbreviations • Congenital heart disease (CHD)

ETIOLOGY/PATHOGENESIS Developmental Anomaly • ~ 12% have chromosomal anomalies ○ Trisomies 13, 18, and 21; DiGeorge, Noonan, Turner, Marfan syndromes, CHARGE, VACTERL/VATER • Familial CHD is rare, may be autosomal dominant, recessive, or X-linked with variable penetrance and expressivity • > 30 individual genes identified ○ Cardiac transcription factors ○ Signaling molecules ○ Ciliary proteins ○ Chromatin modifying genes • Large copy number variants contribute to 10-15% of CHD, often with other developmental anomalies

Environmental Exposure

Prognosis

• Some forms of CHD linked to environmental toxins or maternal drugs, smoking diabetes, or obesity

• Posttransplant outcomes for adult CHD are poorer in 1st year than for cardiomyopathies • Infants have increased early mortality but best overall posttransplant survival (median > 20 years) ○ Slower progression of cardiac allograft vasculopathy

Infectious Agents • Associated with early congenital infections (rubella)

CLINICAL ISSUES Epidemiology • Incidence ○ 0.8% of infants, many surgically repaired without transplant ○ Most common types of CHD that require transplantation – Single ventricle lesions post palliative surgery – Dextro-transposition of great arteries (D-TGA) (12%) – Right ventricular outflow tract lesions [most commonly tetralogy of Fallot (TOF)] – Ventricular septal defect (VSD)/atrial septal defect (ASD) – Left ventricular outflow tract lesions – Congenitally corrected TGA (CC-TGA) – Complete atrioventricular canal defects

Presentation • Usually prenatally diagnosed • Cyanotic heart disease ○ Right-to-left shunts (TOF, pulmonary stenosis/atresia, tricuspid atresia, Ebstein anomaly) ○ D-TGA ○ Total anomalous pulmonary venous connection ○ Truncus arteriosus ○ Hypoplastic left heart syndrome • Acyanotic heart disease ○ Left-to-right shunts (ASD, VSD, atrioventricular septal defect, aortopulmonary window) ○ Aortic coarctation/stenosis/interrupted arch ○ CC-TGA: Blood flows from LA to left-sided RV to aorta

Treatment • Surgical approaches

Heart Transplantation

○ Staged hypoplastic left heart syndrome reconstruction – Norwood procedure (stage I) □ Aorta anastomosed to proximal pulmonary artery (PA) allowing right ventricle (RV) to pump blood to systemic circulation □ Blalock-Taussig shunt (or variant) connects systemic blood supply (innominate artery) to distal PA □ Atrial septectomy allows pulmonary venous blood to return to right atrium – Glenn/Hemi-Fontan (stage II) □ Blalock-Taussig shunt is divided □ Superior vena cava anastomosed to left PA – Fontan procedure (stage III) diverts venous blood directly to PA, bypassing ventricle □ Inferior vena cava connected to right PA via conduit □ Fenestrated conduit connects to RA ○ Mustard/Senning procedures for TGA – Redirect venous return toward opposite ventricle – RV functions as systemic ventricle – Now replaced by arterial switch

TERMINOLOGY

MACROSCOPIC General Features • • • •

Correlate with clinical history including previous surgeries Often multiple abnormalities in single heart Describe origin and course of major coronary arteries Can be described using sequential segmental analysis to evaluate morphology of atria, ventricles, and great vessels • Morphologically, right atrial appendage is triangular with broad base and left atrial appendage is smaller and fingerlike with narrow base • Ventricular structure is determined, and AV valves are described as normal, stenotic, imperforate, or straddling ○ Single ventricle should be described as morphologically left (fine trabeculations with smooth septal surface), right (coarse trabeculations), or indeterminate ○ Most common VSDs are membranous • Aorta should arise posterior and right of PA

Transposition of Arteries: Ventriculoarterial Discordance • Aorta arises from morphologic RV and PA from morphologic left ventricle ○ Associated ASD or VSD allows mixing • Coronary artery anatomy may be abnormal • May be outflow tract obstruction of either vessel • D-TGA: Aorta arises right and anterior to PA

MICROSCOPIC Histologic Features • Nonspecific myocyte hypertrophy and interstitial and endocardial fibrosis 327

Heart Transplantation

Congenital Heart Disease

Hypoplastic Left Heart Syndrome

Norwood Procedure (Stage I) HLHS Repair

Bidirectional Glenn Procedure (Stage II) HLHS Repair

Fontan Procedure (Stage III) HLHS Repair

Tricuspid Atresia

Double Outlet Right Ventricle

(Left) Hypoplastic left heart syndrome (HLHS), the most common single ventricle defect, has atresia, stenosis, or imperforate mitral &/or aortic valves with a small LV ſt and hypoplastic ascending aorta st. Survival depends on a PDA ﬇ and ASD ﬊. (Right) The proximal PA ſt is transected and anastomosed to the hypoplastic aorta augmented with graft ﬇. Blood flows to the ligated distal PA ﬈ via a Blalock-Taussig shunt ﬊. Atrial septectomy ﬊ ensures return of oxygenated blood to the single ventricle.

(Left) In the Glenn procedure (also called hemi-Fontan), the Blalock-Taussig shunt is divided ſt. Blood flow to the lungs is via anastomosis of the superior vena cava to the right PA ﬇. Deoxygenated blood from the inferior vena cava continues to the RV st. (Right) The inferior vena cava is connected to the right PA ſt via a conduit, bypassing the right atrium. Fenestrations in the conduit ﬈ allow deoxygenated blood to return to the RV if pulmonary pressures are high.

(Left) Tricuspid atresia ſt is always associated with an ASD ﬇ and a hypoplastic RV ﬈, and it is almost always associated with a VSD ﬉ and pulmonary outflow tract obstruction. (Right) In double outlet RV, both the aorta ſt and PA st completely or partially arise from the RV. A VSD ﬇ is required for mixing of blood, and the location of the VSD determines whether oxygenated or deoxygenated blood is directed into each vessel.

328

Congenital Heart Disease

Muscular VSD (Left) VSD is the most common congenital heart lesion and may occur in isolation or in association with other malformations. Membranous defects, the most common type, occur just below the aortic valve ﬇. Small defects may close spontaneously. (Right) This muscular VSD st is less common than a membranous defect. There is an intact membranous portion ſt. Muscular defects often close spontaneously.

Ebstein Anomaly

Heart Transplantation

Membranous VSD

Ebstein Anomaly (Left) In Ebstein anomaly, the tricuspid valve is malformed with part of the valve attached to the normal annulus ſt and part attached to the endocardium of the RV (downward displacement) ﬇, resulting in a small distal (functional) RV st and large proximal (atrialized) RV ﬈. (Right) This section from the right ventricle of an explanted heart with Ebstein anomaly shows interstitial fibrosis and myocyte hypertrophy. These findings are nonspecific and are seen in all forms of congenital heart disease.

Truncus Arteriosus

Pulmonary Atresia (Left) In truncus arteriosus, a single proximal great vessel ﬇ above the defect in the ventricular septum gives rise to both the aorta st and pulmonary arteries ſt. The aorta is often abnormal. (Right) In pulmonary atresia ﬇ with an intact ventricular septum ﬈, there is a complete obstruction to RV outflow, with RV hypoplasia and often tricuspid hypoplasia. RV hypoplasia is less marked with a VSD. A patent foramen ovale or ASD ﬊ allows blood flow between the atria. Pulmonary artery flow is dependent on a PDA st.

329

Heart Transplantation

Sarcoidosis, Heart KEY FACTS

• Systemic disease of unclear etiology characterized by noncaseating granulomas

• Lesions are most common in left ventricular free wall and interventricular septum near base of heart • Dilated cardiomyopathy: Common

CLINICAL ISSUES

MICROSCOPIC

• Angiotensin-converting enzyme levels often elevated but are neither sensitive nor specific • 1,25-dihydroxyvitamin D overproduction may cause symptomatic hypercalcemia • Corticosteroids are 1st-line therapy • Pacemaker placement for conduction abnormalities

• Well-formed, noncaseating granulomas with epithelioid histiocytes, usually with giant cells and relative paucity of lymphocytes • Lesions are most common in myocardium • Absence of infectious organisms on special stains (PAS, GMS) • Absence of necrosis and acute inflammatory cells • Late lesions may have more prominent fibrosis (in random distribution) and few granulomas

TERMINOLOGY

IMAGING • Echocardiogram shows left ventricular dilatation or dysfunction with regional wall motion abnormalities

MACROSCOPIC • Areas of fibrosis do not follow typical ischemic patterns, and coronary arteries are patent

TOP DIFFERENTIAL DIAGNOSES • Giant cell myocarditis • Infectious myocarditis • Ischemic cardiomyopathy

Granulomas in Epicardium and Myocardium

Multinucleated Giant Cells

Late-Stage Fibrosis

Cardiac Sarcoidosis (Gross)

(Left) Low-power view demonstrates myocardium and epicardium ﬈ with multiple granulomas ﬊. Granulomas are most commonly found in the myocardium but may also be seen in the epicardium. (Right) Medium-power view of a heart section shows multiple sarcoid granulomas that are typically tightly cohesive with multinucleated giant cells ﬈ and relatively few lymphocytes.

(Left) Low-power view of a section from an explanted heart demonstrates an older area of sarcoid with replacement fibrosis ﬊. There is an adjacent residual granuloma ﬈. (Right) A 4chamber cut of this explanted heart shows patchy, pale tan areas involved by sarcoid in the left ventricle st, interventricular septum ſt, and papillary muscle ﬊.

330

Sarcoidosis, Heart

IMAGING

Definitions

Ultrasonographic Findings

• Systemic disease of unclear etiology characterized by noncaseating granulomas

• Left ventricular dilatation or dysfunction with regional wall motion abnormalities

ETIOLOGY/PATHOGENESIS

MACROSCOPIC

Environmental Exposure

General Features

• Multiple environmental triggers proposed, including ○ Pollen ○ Insecticide ○ Dust

• Dilated cardiomyopathy may be seen • Noncaseating granulomas may be grossly visible as yellow or tan tumor-like infiltrates or only microscopic • Areas of fibrosis do not follow typical ischemic patterns • Patent coronary arteries

Infectious Agents • Prior infection with mycobacteria or propionibacteria suggested as triggers

Genetic Background • Despite abundance of theories concerning etiology, nearly all attribute portion of pathogenesis to genetic predisposition • Familial cases may be associated with specific HLA class II alleles or polymorphisms in TNF-α

CLINICAL ISSUES Epidemiology • Incidence ○ Cardiac involvement in 25% of patients at autopsy (3rd most frequent organ involved) but often clinically silent • Age ○ Young to middle-aged adults • Sex ○ More common in females • Ethnicity ○ Highest incidence in African Americans and northern Europeans ○ Highest rate of cardiac involvement in Japanese

Presentation • Symptoms depend on site of granulomas ○ Conduction abnormalities and heart block are most common • Additional presentations include systolic or diastolic heart failure, sudden cardiac death, and (rarely) pericarditis

Laboratory Tests • Angiotensin-converting enzyme levels often elevated but neither sensitive nor specific • Uninhibited 1,25-dihydroxyvitamin D production by macrophages (theorized to counter-regulate granuloma formation) may cause symptomatic hypercalcemia

Treatment • Corticosteroids are 1st-line therapy • Pacemaker placement for conduction abnormalities • Transplant for cardiac sarcoidosis uncommon

Heart Transplantation

TERMINOLOGY

Sections to Be Submitted • Lesions most common in left ventricular free wall and interventricular septum near base of heart • Endomyocardial biopsy has low diagnostic yield (sensitivity of ~ 30%) due to patchy nature of granulomas and relative lack of right ventricular involvement

MICROSCOPIC Histologic Features • Well-formed, noncaseating granulomas with epithelioid histiocytes and usually giant cells ○ Granulomas have relatively few lymphocytes • Lesions are most common in myocardium but can be in endocardium or epicardium • Schaumann bodies (lamellated, calcified protein) or asteroid bodies (stellate-shaped filamentous aggregates in multinucleated giant cells) suggestive but neither sensitive nor specific for sarcoid • Late lesions may have more prominent fibrosis (in random distribution) and few granulomas • Absence of necrosis and acute inflammatory cells • Absence of infectious organisms on special stains (PAS, GMS, AFB)

DIFFERENTIAL DIAGNOSIS Giant Cell Myocarditis • Absence of well-formed granulomas and fibrosis • Giant cells with adjacent myocyte necrosis • Eosinophils may be prominent, less frequently neutrophils and plasma cells

Infectious Myocarditis • Granulomas often necrotizing and less cohesive • Positive special stains for organisms

Ischemic Cardiomyopathy • Fibrosis typically subendocardial rather than randomly distributed • Coronary artery atherosclerosis present

SELECTED REFERENCES

Prognosis

1.

• Dependent on severity of heart failure

2. 3.

Hulten E et al: Cardiac sarcoidosis-state of the art review. Cardiovasc Diagn Ther. 6(1):50-63, 2016 Lagana SM et al: Cardiac sarcoidosis: a pathology-focused review. Arch Pathol Lab Med. 134(7):1039-46, 2010 Zaidi AR et al: Outcome of heart transplantation in patients with sarcoid cardiomyopathy. J Heart Lung Transplant. 26(7):714-7, 2007

331

Heart Transplantation

Arrhythmogenic Right Ventricular Cardiomyopathy KEY FACTS ○ Standard sections

ETIOLOGY/PATHOGENESIS • Mutations in genes related to desmosomes are common and affect cell-cell adhesion

CLINICAL ISSUES • Diagnosis made on combination of clinical, imaging, genetic, and pathologic criteria

MACROSCOPIC • Classically, large segments of right ventricular wall replaced by fibroadipose tissue ○ Result in ventricular dilation and translucent wall • Areas most commonly involved by dysplasia include ○ Apex ○ Inferior/diaphragmatic side of right ventricle ○ Anterior surface of pulmonary infundibulum • Generous sampling of right ventricle should include ○ Outflow tract ○ Posterior wall

MICROSCOPIC • Infiltration of ventricular wall by fat ○ With focal fibrosis and hypertrophy of remaining myocytes • Fibrosis and fatty infiltration begin in subepicardial region and extend toward endocardium

TOP DIFFERENTIAL DIAGNOSES • Normal right ventricular fat ○ Most prominent in anterior and lateral aspects – Unusual in posterior wall ○ Adipose tissue infiltrates in even streaks between myocytes so that original outer border of myocardium can be distinguished ○ Unaccompanied by fibrosis, inflammation, or myocyte necrosis

Explanted Heart

Right Ventricle, Low Power

Right Ventricle Endocardium, High Power

Residual Myocytes, High Power

(Left) Transillumination of this explanted heart with right ventricular cardiomyopathy is possible due to replacement of the majority of the right ventricular myocardium by fibroadipose tissue. (Right) The right ventricle (RV) free wall demonstrates replacement of the myocardium with fibroadipose tissue extending from the subepicardium ﬈ toward the endocardium ﬊, where residual myocytes remain.

(Left) High-power trichrome stain highlights the fibrosis ﬈ entrapping the residual subendocardial myocytes ﬇ subjacent to the endocardium ﬉. (Right) High-power view demonstrates residual myocytes ﬈ entrapped in surrounding paler pink fibrous tissue ﬊ with scattered small lymphocytes within the fat, which has replaced the RV free wall.

332

Arrhythmogenic Right Ventricular Cardiomyopathy

Abbreviations • Arrhythmogenic right ventricular cardiomyopathy (ARVC)

Synonyms • Arrhythmogenic cardiomyopathy • Arrhythmogenic right ventricular dysplasia (ARVD)

Definitions • Fibrofatty infiltration of myocardium, typically involving right ventricle (RV) or both ventricles • Diagnosis based on combination of clinical, EKG, imaging, genetic, and pathologic features ○ Major and minor criteria established by international task force

ETIOLOGY/PATHOGENESIS Genetics • Mutations in desmosomal genes common and affect cellcell adhesion ○ Detected in ~ 50% of patients ○ Most commonly autosomal dominant with variable expressivity ○ Plakophilin 2 (PKP2) most common mutation (up to 45%) ○ Desmoplakin (DSP) mutated in 10-15% – Left ventricular changes may be particularly prominent – Carvajal syndrome □ Autosomal recessive form of ARVC with DSP mutation □ ARVC, woolly hair, and epidermolytic palmoplantar keratoderma ○ Naxos syndrome – Autosomal recessive plakoglobin (JUP) mutations – ARVC, woolly hair, and palmoplantar keratoderma ○ Desmoglein 2 (DSG2) and desmocollin 2 (DSC2) are additional desmosomal genes that may have mutations ○ Some have > 1 mutation – Associated with more severe phenotype • Mutations in nondesmosomal proteins rare and include ○ Human ryanodine receptor 2 ○ Transforming growth factor β-3 ○ Transmembrane protein 43 encoding gene ○ Lamin A/C ○ Phospholamin ○ Desmin ○ Titin ○ αT-catenin ○ Some of these genes more commonly associated with other cardiomyopathy phenotypes (dilated or hypertrophic)

Pathophysiology • Hypothesis include those related to cell death and inflammation and those related to transdifferentiation of mesenchymal cells ○ Defects in cell-cell adhesion may lead to detachment and cell death with replacement by fibroadipose tissue

○ Missense mutations in desmosomal proteins may lead to aberrant protein localization with effects on intracellular signaling pathways, leading to transdifferentiation of myocytes into adipocytes • Arrhythmias thought to arise due to altered conduction at sites of abnormal desmosomal connections or at sites of fibroadipose tissue

CLINICAL ISSUES

Heart Transplantation

TERMINOLOGY

Epidemiology • Incidence ○ ~ 1 in 5,000 – Higher if mild/clinically undetected cases included ○ In regions of high prevalence (e.g., Italy), may account for up to 20% of sudden death in young adults • Age ○ Typically presents in young adults but case reports in elderly ○ Cases more difficult to diagnose in children since it is progressive disease • Sex ○ More common in males despite autosomal inheritance • Ethnicity ○ Most common in subregions of Europe but occurs worldwide

Presentation • Palpitations, syncope, or sudden death • Almost always accompanied by ventricular arrhythmias • May progress to heart failure

Laboratory Tests • Genetic testing can confirm diagnosis in proband ○ Helpful in counseling of family members ○ Large numbers of variants of uncertain significance ○ Frequent mutations in healthy controls can make genetic interpretation challenging

Natural History • Asymptomatic phase when patient at risk for sudden cardiac death ○ May be followed by symptomatic arrhythmias, right ventricular failure, or biventricular failure • Strenuous physical activity promotes progression

Treatment • Sports restriction recommended to prevent clinical disease in asymptomatic genetic carriers and slow progression in symptomatic patients • Arrhythmias may be treated with antiarrhythmogenic drugs, catheter ablation of the scar, or implantable cardioverter-defibrillator • Transplantation may be required for progression to heart failure mimicking dilated cardiomyopathy

IMAGING MR Findings • MR preferred modality for imaging of RV • Increased right ventricular adipose tissue and fibrosis • Right ventricular wall motion abnormalities and dilation 333

Heart Transplantation

Arrhythmogenic Right Ventricular Cardiomyopathy

MACROSCOPIC General Features • Large segments of right ventricular wall replaced by fibroadipose tissue, classically ○ Result in ventricular dilation and translucent wall – Aneurysm may result ○ Areas most commonly involved by "dysplasia" include – Apex – Inferior/diaphragmatic side of RV – Anterior surface of pulmonary infundibulum • RV alone or both ventricles may be involved ○ Rare left ventricle-predominant disease ○ Left ventricular disease may be identified grossly as patchy subendocardial fibrosis

• Dilated cardiomyopathy does not typically have subepicardial distribution of fibrosis • Definitive distinction not always possible on morphologic basis ○ Differential diagnosis should be raised for genetic testing

Transmural Infarction of Right Ventricle • Thinning of right ventricular wall is focal • Thick band of replacement fibrosis • Corresponding coronary artery disease

SELECTED REFERENCES 1. 2.

Sections to Be Submitted • Generous sampling of RV to include outflow tract and posterior wall in addition to standard sections ○ Fat not usually present in posterior of normal RV

MICROSCOPIC Histologic Features • Infiltration of ventricular wall by fat with focal fibrosis and hypertrophy of remaining myocytes ○ Fatty infiltration begins in subepicardial region and progresses toward endocardium – Infiltration may be transmural in RV – Left ventricle involvement typically focal and subendocardial □ Consists of fibrosis ± fat – Infiltrating adipose tissue is irregular – Generous sampling needed to identify fibrosis • Individual myocyte necrosis and mild lymphocytic inflammation may be present • Estimated myocyte loss of > 50% with fibrous replacement of RV free wall (± fatty replacement) is major diagnostic criterion ○ Endomyocardial biopsy may have low sensitivity due to sampling of subendocardial region

3.

4.

5. 6.

7. 8.

9. 10.

11.

12. 13.

14. 15.

DIFFERENTIAL DIAGNOSIS Normal Right Ventricular Fat • Right ventricular adipose tissue increases with age and weight • More prominent in women • Most common in lateral and anterior walls • Usually relatively little fat in posterior wall or outflow tract, even with morbid obesity • Usually subepicardial but may be transmural ○ Infiltrates in even streaks between myocytes ("marbling") ○ Unaccompanied by fibrosis, inflammation, or myocyte necrosis

Dilated Cardiomyopathy • Some left ventricular predominant or biventricular forms of ARVC can mimic dilated cardiomyopathy • ARVC accompanied by more arrhythmias than expected for dilated cardiomyopathy 334

16.

17.

Corrado D et al: Arrhythmogenic right ventricular cardiomyopathy. N Engl J Med. 376(15):1489-90, 2017 Pilichou K et al: Large genomic rearrangements of desmosomal genes in Italian arrhythmogenic cardiomyopathy patients. Circ Arrhythm Electrophysiol. 10(10), 2017 Bhonsale A et al: Impact of genotype on clinical course in arrhythmogenic right ventricular dysplasia/cardiomyopathy-associated mutation carriers. Eur Heart J. 36(14):847-55, 2015 Corrado D et al: Treatment of arrhythmogenic right ventricular cardiomyopathy/dysplasia: an international task force consensus statement. Eur Heart J. 36(46):3227-37, 2015 Lazzarini E et al: The ARVD/C genetic variants database: 2014 update. Hum Mutat. 36(4):403-10, 2015 McGregor SM et al: A brief review and update of the clinicopathologic diagnosis of arrhythmogenic cardiomyopathy. Arch Pathol Lab Med. 139(9):1181-6, 2015 Rizzo S et al: The changing spectrum of arrhythmogenic (right ventricular) cardiomyopathy. Cell Tissue Res. 348(2):319-23, 2012 Azaouagh A et al: Arrhythmogenic right ventricular cardiomyopathy/dysplasia: a review and update. Clin Res Cardiol. 100(5):38394, 2011 Basso C et al: Pathophysiology of arrhythmogenic cardiomyopathy. Nat Rev Cardiol. 9(4):223-33, 2011 Quarta G et al: Familial evaluation in arrhythmogenic right ventricular cardiomyopathy: impact of genetics and revised task force criteria. Circulation. 123(23):2701-9, 2011 Marcus FI et al: Diagnosis of arrhythmogenic right ventricular cardiomyopathy/dysplasia: proposed modification of the Task Force Criteria. Eur Heart J. 31(7):806-14, 2010 Sen-Chowdhry S et al: Arrhythmogenic cardiomyopathy: etiology, diagnosis, and treatment. Annu Rev Med. 61:233-53, 2010 El Demellawy D et al: An updated review on the clinicopathologic aspects of arrhythmogenic right ventricular cardiomyopathy. Am J Forensic Med Pathol. 30(1):78-83, 2009 Tansey DK et al: Fat in the right ventricle of the normal heart. Histopathology. 46(1):98-104, 2005 d'Amati G et al: Arrhythmogenic right ventricular cardiomyopathy: clinicopathologic correlation based on a revised definition of pathologic patterns. Hum Pathol. 32(10):1078-86, 2001 Burke AP et al: Arrhythmogenic right ventricular cardiomyopathy and fatty replacement of the right ventricular myocardium: are they different diseases? Circulation. 97(16):1571-80, 1998 McKenna WJ et al: Diagnosis of arrhythmogenic right ventricular dysplasia/cardiomyopathy. Task Force of the Working Group Myocardial and Pericardial Disease of the European Society of Cardiology and of the Scientific Council on Cardiomyopathies of the International Society and Federation of Cardiology. Br Heart J. 71(3):215-8, 1994

Arrhythmogenic Right Ventricular Cardiomyopathy

Left Ventricle Subendocardial Fibrosis (Left) Close-up view of the left ventricular free wall of an explanted heart with arrhythmogenic cardiomyopathy demonstrates small, pale, yellow-tan foci of fibrofatty replacement at the subepicardial aspect ſt. (Right) Low-power view of the left ventricle shows typical subendocardial involvement with superficial fatty ﬉ infiltration and pale pink fibrosis ﬊ in the subepicardial region. There is relative sparing of the subendocardial myocardium ﬈.

Left Ventricle Subendocardium, High Power

Heart Transplantation

Left Ventricle Subendocardial Fibrosis

Normal Fat Distribution in Obese Patient (Left) Microscopic section of the left ventricular free wall demonstrates abundant pale, dense fibrosis ﬊ below the epicardium ﬈. There is relative sparing of the myocardium deeper ﬉ in the wall. (Right) Normal RV from an obese patient demonstrates that the majority of the normal fat is distributed around the anterior st and lateral ſt aspects of the RV with sparing of the posterior ﬇ surface.

Normal Right Ventricle Fat Distribution

Normal Fat Distribution, High Power (Left) Right ventricular free wall from the autopsied heart of an obese woman demonstrates extensive fatty infiltration of the wall extending from the epicardium ﬉ to the endocardium with even strands of fat pushing apart bundles of myocytes. There is no fibrosis. (Right) Higher power view of fat within a normal RV demonstrates alternating strands of fat and myocardial cells in relatively even bands without fibrosis or damage to the myocardium.

335

Heart Transplantation

Other Causes of End-Stage Heart Disease KEY FACTS

CLINICAL ISSUES

MICROSCOPIC

• Deposition of β-amyloid in heart usually presents as restrictive heart disease ○ Hereditary amyloidosis most often due to transthyretin mutations ○ Transplantation is controversial and depends on amyloid type • Iron deposition may be primary (hereditary hemochromatosis) or secondary to iron overload ○ Usually presents as restrictive cardiomyopathy progressing to dilated cardiomyopathy • Chemotherapy-induced cardiomyopathy is most commonly due to anthracyclines ○ May present during treatment or decades later as dilated or restrictive cardiomyopathy ○ Transplant is option in long-term survivors with complete remission

• Deposition of amyloid is usually diffuse and interstitial, and biopsy is nearly 100% sensitive ○ Amyloid should stain red with Congo red and polarize as apple green ○ Immunohistochemistry has relatively low sensitivity for identification of amyloid type ○ Laser-capture microdissection and mass spectrometry have extremely high sensitivity for identification of amyloid types • In hemochromatosis, iron accumulation is greatest in subepicardial region, and biopsy is not sensitive ○ Any iron deposition is abnormal • In drug-induced cardiomyopathy, by time of explant, there is chronic damage with interstitial or replacement fibrosis and myocyte hypertrophy ○ May be no diagnostic findings on electron microscopy

Systemic Amyloidosis

Cardiac Amyloidosis

Cardiac Amyloid

Amyloidosis: Congo Red

(Left) In this cross section of the cardiac ventricles from an autopsy of a patient with systemic amyloidosis, both the left ﬇ and right ſt ventricular walls are markedly thickened with small ventricular cavities. The heart has a rubbery consistency. (Right) In this high-power view of a cardiac biopsy with amyloidosis, amyloid is seen in a typical, relatively diffuse interstitial ﬈ pattern surrounding individual myocytes.

(Left) This explanted heart shows an unusual, relatively circumscribed collection of amyloid ﬉ focally replacing myocytes. There is an absence of the usual interstitial amyloid between myocytes ﬈. (Right) Congo red stain of this large biopsy taken at the time of LVAD insertion shows deposition of red-staining amyloid in the vessels ﬉ as well as interstitially between adipocytes ﬈ and within fibrous tissue ﬊.

336

Other Causes of End-Stage Heart Disease

Amyloidosis • Deposition of β-amyloid in heart usually presents as restrictive heart disease • Echocardiogram shows biventricular hypertrophy and "granular" echogenicity ○ Usually concentric but can occasionally have septal prominence • Posttransplant survival worse than other restrictive cardiomyopathies ○ Outcome may depend on amyloid type • Subcutaneous fat aspiration can confirm diagnosis of amyloidosis without cardiac biopsy in appropriate clinical setting • Subtyping amyloid in heart is important to determine prognosis and treatment ○ Serum and urine testing for paraproteins may give falsepositive or false-negative results – Low levels of circulating light chains may not be detected • Light chain (primary systemic) amyloidosis ○ Associated with underlying plasma cell dyscrasia – Treatment with chemotherapy and sometimes stem cell transplant ○ Usually other organ systems also affected ○ Median survival < 1 year without treatment ○ Heart transplant is rare and controversial option • Hereditary amyloidosis ○ Mutations in many different proteins synthesized by liver – Transthyretin mutations are most common ○ Affected organs often include heart, liver, and peripheral nerves ○ Patients may be candidates for liver or combined heartliver transplant • Wild-type transthyretin amyloidosis ○ Formerly "senile systemic amyloidosis" ○ Elderly male patients (usually > 70 years) ○ Predominantly affects heart • Secondary amyloidosis ○ Amyloid-associated protein, acute phase reactant ○ Seen in chronic inflammatory disease ○ Rarely affects heart • Isolated atrial amyloidosis ○ Atrial natriuretic peptide ○ Predominantly in elderly women ○ Usually identified in atrial appendage resections

Hemochromatosis • Iron deposition may be primary (hereditary hemochromatosis) or secondary to iron overload • Usually presents as restrictive cardiomyopathy progressing to dilated cardiomyopathy • In secondary iron overload, diagnosis usually known, and biopsy or transplant not indicated • Treatment for hereditary hemochromatosis is phlebotomy ○ Combined heart/liver transplant reported but extremely rare

○ Most had previous surgical repairs ○ Transplants most commonly performed for chronic rheumatic heart disease

Drug-Induced Cardiomyopathy • Commonly due to anthracyclines but also alkylating agents, antimicrotubule agents, trastuzumab, and tyrosine kinase inhibitors ○ Dependent on cumulative dose, use of multiple agents, and dosing schedule ○ May also be affected by local radiation treatment ○ End-stage heart disease in 1-5% of patients ○ May present during treatment or decades later as dilated or restrictive cardiomyopathy • Transplant is option in long-term survivors with complete remission of primary disease ○ Most common for breast cancer, leukemia, or sarcoma

Heart Transplantation

CLINICAL ISSUES

Endocardial Fibroelastosis • Prevalence markedly decreased and now extremely rare ○ Historically may have been due to in utero viral infections • Presents in infants and children ○ Originally described as primary change or secondary to congenital heart disease (most commonly hypoplastic left heart syndrome) ○ No longer included in most recent American Heart Association classification of cardiomyopathies – Similar focal findings in many other types of heart disease – Rarely secondary to other defined myocardial disease including viral infections and metabolic diseases ○ Usually sporadic (10% familial) • Death due to heart failure usually within weeks to months but occasionally more chronic

Inborn Errors of Metabolism • > 40 inborn errors in metabolism may have cardiomyopathy as component ○ Includes general categories of mitochondrial diseases, glycogen or fatty acid metabolism disorders, and lysosomal storage diseases • Most autosomal recessive, X-linked, or have maternal inheritance due to mitochondrial mutations • Most with multiorgan involvement, which limits appropriateness for transplant ○ Cardiac manifestations may be most important for prognosis – Present with arrhythmias, valvular disease, or as hypertrophic, dilated, or restrictive cardiomyopathy ○ Although transplants relatively rare, may be considered if limited disease outside heart or considered for combined heart/liver transplant • Diagnosis usually made based on clinical findings and tests for metabolites and enzyme activity

MACROSCOPIC Amyloidosis • Usually biventricular wall thickening • Consistency is waxy or rubbery

Valvular Heart Disease

Hemochromatosis

• Relatively rare indication for transplantation

• May be dilated or restrictive 337

Heart Transplantation

Other Causes of End-Stage Heart Disease • Iron may cause myocardium to have darker brown color than normal

Chronic Valvular Disease • Valves with healed endocarditis are rubbery and thickened with thick chordae tendineae

Drug-Induced Cardiomyopathy • May be dilated or restrictive • Features are not specific

Primary Endocardial Fibroelastosis • May be associated with dilated (more common) or contracted left ventricle • Usually left ventricle and atrium most affected but often extends to right side • Endocardium diffusely thickened and white • Mitral and aortic valves may be involved and distorted ○ Often thickening of chordae tendineae ○ Papillary muscles are small and attached higher in left ventricle wall than normal

Inborn Errors of Metabolism • May present as either hypertrophic or dilated cardiomyopathy • Defects in mitochondria may present as histiocytoid cardiomyopathy with yellowish patchy thickening of endocardium

MICROSCOPIC Amyloidosis • Deposition of amyloid is usually uniform, resulting in high sensitivity for biopsy (near 100%) ○ Amyloid deposits are diffuse and interstitial • Amyloid should stain red with Congo red and polarize as apple green ○ Interstitial collagen does not stain red and is white when polarized ○ Most but not all amyloid will polarize • Amyloid stains green with sulfated Alcian blue • Rarely, deposition may be too scant to be detected by special stains but can be identified by electron microscopy ○ Amyloid fibrils are haphazardly arranged, unbranched filaments 8-10 nm in width ○ Often surround and constrict myocytes • Immunohistochemistry has relatively low sensitivity for identification of amyloid type • Laser-capture microdissection of amyloid from paraffinembedded tissue followed by mass spectrometry has extremely high sensitivity for identification of amyloid types ○ Allows identification of any type of amyloid, whether or not immunohistochemistry is successful

Hemochromatosis • Iron accumulation is greatest in subepicardial region ○ Endomyocardial biopsy has limited sensitivity in this setting • Any iron deposition in myocytes is abnormal • Iron stains blue by Prussian blue stain

Chronic Valvular Disease • Neovascularization seen in healed valvulitis 338

○ Vessels absent in normal heart valves • Typical features of acute rheumatic heart disease (Aschoff nodules) are not present

Drug-Induced Cardiomyopathy • In acute anthracycline damage, electron microscopy may be used to grade anthracycline toxicity ○ Vacuolization ○ Myofibrillar dropout ○ Necrosis • By time of explant, there is chronic damage with interstitial or replacement fibrosis and myocyte hypertrophy ○ Electron microscopy findings not well described

Endocardial Fibroelastosis • Endocardium thickened with parallel arrays of elastic fibers and collagen

Inborn Errors of Metabolism • Light microscopy often shows nonspecific degenerative changes including hypertrophy and vacuolization ○ Rarely PAS staining shows glycogen-positive vacuoles • Electron microscopy may show increased ○ Abnormal mitochondria ○ Vacuolization ○ Inclusions depending on metabolic abnormality ○ EM findings rarely pathognomonic of specific genetic defect • Histiocytoid cardiomyopathy has histiocyte-like myocytes with foamy pale cytoplasm forming subendocardial nodules ○ Positive for myocyte markers (desmin) and negative for histiocytic markers

DIFFERENTIAL DIAGNOSIS Endocardial Fibroelastosis vs. Focal Endocardial Fibrosis in Dilated Cardiomyopathy • Endocardial fibrosis thicker and more diffuse in endocardial fibroelastosis (EFE) • Interstitial fibrosis and myocytolysis may be present in both • Replacement fibrosis not identified in "primary" EFE

SELECTED REFERENCES 1.

Muchtar E et al: Restrictive cardiomyopathy: genetics, pathogenesis, clinical manifestations, diagnosis, and therapy. Circ Res. 121(7):819-837, 2017 2. El-Hattab AW et al: Mitochondrial cardiomyopathies. Front Cardiovasc Med. 3:25, 2016 3. Gertz MA et al: Pathophysiology and treatment of cardiac amyloidosis. Nat Rev Cardiol. 12(2):91-102, 2015 4. Shah S et al: Advanced heart failure due to cancer therapy. Curr Cardiol Rep. 17(4):16, 2015 5. Shehata BM et al: Exome sequencing of patients with histiocytoid cardiomyopathy reveals a de novo NDUFB11 mutation that plays a role in the pathogenesis of histiocytoid cardiomyopathy. Am J Med Genet A. 167A(9):2114-21, 2015 6. Gulati V et al: Cardiac involvement in hemochromatosis. Cardiol Rev. 22(2):56-68, 2014 7. Seki A et al: Primary endocardial fibroelastosis: an underappreciated cause of cardiomyopathy in children. Cardiovasc Pathol. Epub ahead of print, 2013 8. Bernaba BN et al: Pathology of late-onset anthracycline cardiomyopathy. Cardiovasc Pathol. 19(5):308-11, 2010 9. Rotela Samaniego JA et al: Clinical evolution of heart transplantation in patients with previous valvular cardiomyopathy. Transplant Proc. 39(7):23556, 2007 10. Mackay B et al: Assessment of anthracycline cardiomyopathy by endomyocardial biopsy. Ultrastruct Pathol. 18(1-2):203-11, 1994

Other Causes of End-Stage Heart Disease

Amyloidosis: Electron Microscopy (Left) Viewing the Congo red stain with polarized light demonstrates the typical apple-green birefringence ﬈. Note that not all the redstained amyloid ﬊ is polarized. The collagen ﬇ also polarizes but is white and should not be mistaken for amyloid. (Right) Electron microscopy of a biopsy with cardiac amyloidosis demonstrates pale gray bands of amyloid ﬈ closely hugging the myocytes ﬊. At this low magnification, the fibrillar structure of amyloid is not apparent.

Electron Microscopy Amyloid

Heart Transplantation

Amyloidosis: Polarized Congo Red Stain

Hereditary Hemochromatosis (Left) Higher power view of amyloid by electron microscopy shows disorganized, unbranched fine fibrils ﬈ of the typical size (810 nm). (Right) This cross section of the ventricles of an autopsy heart from a patient with hereditary hemochromatosis shows a subtle brown discoloration. The left ventricle st has a small chamber consistent with restrictive cardiomyopathy, while the right ventricular wall is thinned and the chamber ﬇ is dilated.

Hemochromatosis

Hemochromatosis: Prussian Blue (Left) High-power view of hemochromatosis shows brown-yellow iron pigment within the myocytes ﬈. The differential diagnosis includes lipofuscin pigment. This finding is most pronounced in the outer myocardium, which is less likely to be biopsied. (Right) Prussian blue stain of the same case shows positive (blue) staining for iron, which in some cells in very pronounced ﬈ and in others is fainter ﬊. Any degree of iron accumulation in myocytes is abnormal.

339

Heart Transplantation

Other Causes of End-Stage Heart Disease

Prosthetic Mitral Valve

Healed Valvulitis

Rheumatic Heart Disease

Rheumatic Heart Disease

Doxorubicin Toxicity

Doxorubicin Toxicity

(Left) In this explanted heart (viewed from above), the residual left atrial tissue ﬇ is pulled back to reveal the suture lines ſt of a bioprosthetic mitral valve. Gross examination should include if suture line is intact and whether vegetations are present on the leaflets. (Right) In this example of healed valvulitis, small blood vessels ﬊ are present. These are muscularized arterioles which are not present in normal valves. There is no acute inflammation or vegetation.

(Left) This explanted heart with chronic rheumatic heart disease had a dilated mitral valve with a markedly thickened, rubbery leaflet ﬊ and thickened chordae tendineae ſt. (Right) In this section of mitral valve from a patient with chronic rheumatic heart disease, there is marked neovascularization ﬈ with small granulation tissue-type vessels but minimal inflammatory cells. These findings may be seen in healed valvulitis of any etiology.

(Left) This is an explanted heart from a patient with doxorubicin toxicity with restrictive cardiomyopathy. The left ventricle is small with a transmural defect ﬇ from placement of a ventricular assist device. (Right) In this section from a mildly dilated explanted heart from a patient with doxorubicin toxicity, there are nonspecific changes of interstitial fibrosis ﬈ and mild myocyte hypertrophy ﬉.

340

Other Causes of End-Stage Heart Disease

Endocardial Fibroelastosis (Left) In this explanted heart from a child with endocardial fibroelastosis, the entire endocardial surface of the left ventricle ﬈ is thickened and fibrotic. The papillary muscle ﬊ arises high in the wall of the ventricle, and the mitral valve and chordae tendineae are thickened. (Right) This section from the explanted pediatric heart with endocardial fibroelastosis shows a thickened, collagenous endocardium ﬈.

Endocardial Fibroelastosis

Heart Transplantation

Endocardial Fibroelastosis

Histiocytoid Cardiomyopathy (Left) A Verhoeff van Gieson elastic stain of the endocardial thickening in endocardial fibroelastosis shows multiple parallel thick elastic layers ﬈ above the collagenous ﬊ layers of the endocardium overlying the myocardium ﬉. The ventricular cavity is at the top. (Right) In histiocytoid cardiomyopathy, the myocytes have voluminous pale eosinophilic to clear cytoplasm, sometimes bubbly or vacuolated ﬈, and can be mistaken for macrophages.

Mitochondrial Myopathy

Mitochondrial Myopathy (Left) Explanted heart from a patient with the mitochondrial myopathy Kearns-Sayre syndrome shows dilated cardiomyopathy with a large left ventricular chamber ﬇ and thinned left ventricular wall st. (Right) High-power view of a heart section from a patient with Kearns-Sayre syndrome shows nonspecific degenerative changes and vacuolization ﬉ of the hypertrophic myocytes.

341

Heart Transplantation

Acute Cellular Rejection, Heart KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Rejection of transplanted heart mediated by activated lymphocytes; may occur within days to years after transplantation

• EMB performed for rejection can be graded per International Society of Lung and Heart Transplantation guidelines (2005) ○ Several centers prefer 1990 grading since it provides more detailed data, which can be translated to 2005 grading

CLINICAL ISSUES • Most common in 1st 6 months after transplantation but may occur at any time (late acute rejection is often due to noncompliance with immunosuppressive medications) • Many patients are asymptomatic, rejection being most commonly diagnosed at time of surveillance endomyocardial biopsy (EMB) • Only moderate to severe grades of rejection are treated ○ Mild rejection treated if symptomatic • Most patients respond well to increased immunosuppression • Repeated attacks of ACR predispose to accelerated graft vasculopathy (chronic rejection)

TOP DIFFERENTIAL DIAGNOSES • Quilty effect ○ Endocardial lymphocytic infiltrate that often extends into underlying myocardium ○ Mature small lymphocytes with several capillaries • Site of previous biopsy ○ Organizing fibrin and granulation tissue, reactive endothelial cells, hemosiderin-laden macrophages, rare inflammatory cells

Focal Moderate Rejection

Focal Moderate Rejection

Multifocal Moderate Rejection

Severe Acute Rejection

(Left) Endomyocardial biopsy shows a focus of lymphocytic infiltrate, which is obvious at low power (grade 2 rejection). There is separation and replacement of myocytes as well as myocyte damage ſt. (Right) High power of endomyocardial biopsy shows lymphocytes and myocyte damage (grade 2/1R acute cellular rejection). The lymphocytes are large and activated. Myocyte borders are frayed and irregular ſt.

(Left) Heart biopsy shows multiple foci of acute rejection. The lymphocytes are large and activated ﬉. Myocyte damage is present ſt. (Right) There is an extensive infiltrate of activated lymphocytes and eosinophils that extends into the overlying endocardium. Myocyte damage is readily evident ſt.

342

Acute Cellular Rejection, Heart

Abbreviations • Acute cellular rejection (ACR)

Synonyms • Acute rejection (AR)

Definitions • Rejection of transplanted heart mediated by activated lymphocytes; may occur within days to years after transplantation

CLINICAL ISSUES Presentation • Most common in 1st 6 months after transplantation but may occur any time ○ Late AR often due to noncompliance with immunosuppressive medications • Many patients are asymptomatic, rejection being most commonly diagnosed at time of surveillance endomyocardial biopsy (EMB) • ~ 5% of patients exhibit symptoms ○ Nonspecific symptoms (e.g., malaise, fever, fatigue, nausea) ○ May be specific symptoms related to right or left ventricular dysfunction

Treatment • Drugs ○ Only moderate to severe grades of rejection are treated – Mild rejection may be treated if patient symptomatic or if evidence of hemodynamic compromise ○ Increase immunosuppression above baseline triple drug combination – For higher grades of rejection, add additional immunosuppressive drug – Specific treatment regimens vary from center to center

Prognosis • Most respond well to increased immunosuppression • Repeated attacks of ACR predispose to risk for accelerated graft vasculopathy (chronic rejection)

• Focal moderate rejection: Single large focus of lymphocytic infiltrate (± eosinophils) often associated with myocyte damage • Multifocal moderate rejection: ≥ 2 foci of moderate rejection • Diffuse moderate rejection: Extensive interstitial infiltrates • Severe rejection: Diffuse infiltrates with myocyte necrosis/damage

Heart Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Quilty Effect • Endocardial lymphocytic infiltrate that often extends into underlying myocardium • Mature small lymphocytes with several capillaries • CD21 highlights follicular dendritic meshwork in larger Quilty lesions, which is diagnostic

Site of Previous Biopsy • Organizing fibrin and granulation tissue, reactive endothelial cells, hemosiderin-laden macrophages, rare inflammatory cells

Infection • Cytomegalovirus and toxoplasmosis; exceedingly rare

Perioperative Ischemic Injury • Myocyte damage and necrosis is out of proportion to cellular infiltrate; frank coagulation necrosis is present rather than myocytolysis

Posttransplant Lymphoproliferative Disorder • Large, atypical lymphocytes, often positive for Epstein-Barr virus

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Infiltrate of predominantly activated T lymphocytes and macrophages ± eosinophils • Myocytolysis is prominent feature of myocyte damage and different from coagulative necrosis • Multiple levels need to be examined for appropriate grading as well as to distinguish from mimics

SELECTED REFERENCES

MICROSCOPIC

1.

Histologic Features

2.

• EMB performed for rejection can be graded per International Society of Lung and Heart Transplantation guidelines (2005) • Original grading consisted of 7-tier system (1990), which was subsequently updated in 2005 to reduce inter- and intraobserver variability ○ Some prefer 1990 grading as it provides more data that can be translated to 2005 grading system • Focal mild rejection: Small perivascular lymphocytic infiltrates • Diffuse mild rejection: Few activated lymphocytes between myocytes

3. 4. 5.

6.

7.

Ishibashi-Ueda H et al: Significance and value of endomyocardial biopsy based on our own experience. Circ J. 81(4):417-426, 2017 Andrew J et al: Latest developments in heart transplantation: a review. Clin Ther. 37(10):2234-41, 2015 Patel JK et al: Cardiac allograft rejection. Surgeon. 9(3):160-7, 2011 Tan CD et al: Update on cardiac transplantation pathology. Arch Pathol Lab Med. 131(8):1169-91, 2007 Sattar HA et al: The presence of a CD21+ follicular dendritic cell network distinguishes invasive Quilty lesions from cardiac acute cellular rejection. Am J Surg Pathol. 30(8):1008-13, 2006 Stewart S et al: Revision of the 1990 working formulation for the standardization of nomenclature in the diagnosis of heart rejection. J Heart Lung Transplant. 24(11):1710-20, 2005 Billingham ME et al: A working formulation for the standardization of nomenclature in the diagnosis of heart and lung rejection: Heart Rejection Study Group. The International Society for Heart Transplantation. J Heart Transplant. 9(6):587-93, 1990

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Heart Transplantation

Acute Cellular Rejection, Heart

Heart Biopsy Without Rejection

Focal Mild Rejection

Diffuse Mild Rejection

Focal Moderate Rejection

Multifocal Moderate Rejection

Diffuse Moderate Rejection

(Left) This endomyocardial biopsy from a heart transplant recipient shows no acute rejection. There are some prominent endothelial cells ſt and rare lymphocytes. (Right) Focal mild acute cellular rejection (grade 1A/1R) as seen here is characterized by a relatively small infiltrate of lymphocytes with no associated myocyte damage. The infiltrate is not aggressive, i.e., it does not infiltrate between myocytes and disappears on deeper sections. It is perivascular in location.

(Left) Mild (diffuse) acute cellular rejection (grade 1B/1R) is characterized by a diffuse but mild infiltrate of lymphocytes, which are seen between myocytes ſt in all the biopsy fragments. There is no myocyte damage. (Right) Focal moderate acute cellular rejection (grade 2/1R) is seen in this heart transplant biopsy. One of the biopsy fragments shows an area of lymphocytic infiltrate, which was seen on all 5 levels. Myocyte injury/damage is evidenced by frayed edges ſt.

(Left) Multifocal moderate rejection is seen in this endomyocardial biopsy, which had distinct foci of inflammation associated with myocyte damage. The infiltrate also involves the endocardium ﬊, which does not mean that it is a Quilty lesion since the character of the inflammation is the same as seen in the areas of rejection. (Right) Diffuse moderate rejection (grade 3B/3R) shows an extensive infiltrate of activated, large lymphocytes and several eosinophils ſt.

344

Acute Cellular Rejection, Heart

Intralymphatic Lymphocytes (Left) Site of previous heart biopsy shows surface fibrin ﬈ on the endocardium. There is early organization with granulation tissue and fibrosis. Inflammation is minimal with only a few scattered mononuclear cells. (Right) Intralymphatic lymphocytes ſt are seen in this surveillance biopsy from a heart transplant patient. The lymphatic channel is difficult to see, but the well-defined collection of mature small lymphocytes differentiate it from acute cellular rejection.

Contraction Bands

Heart Transplantation

Site of Previous Biopsy

Eosinophilic Drug Reaction (Left) Contraction bands are often present in endomyocardial biopsies. These are thought to be secondary to the biopsy procedure. The presence of cross-striations and preservation of nuclei differentiate it from infarction. (Right) Although eosinophils are often present in higher grades of acute cellular rejection, when there is a linear infiltrate out of proportion to lymphocytes, without associated myocyte damage, as seen here, it is suggestive of a drug reaction.

Quilty Lesion

Quilty Lesion (Left) There are multiple foci of Quilty lesion composed of large dense lymphocytic infiltrates. Myocytes in the uninvolved areas appear unremarkable, which together with the localized infiltrates differentiate from acute rejection. (Right) Invasive Quilty lesion at high power is composed of mature small lymphocytes, which infiltrate into the underlying myocardium. There is focal myocyte damage ﬈, the presence of which does not make this rejection.

345

Heart Transplantation

Antibody-Mediated Rejection, Heart KEY FACTS

TERMINOLOGY • Rejection due either to de novo antibody formation, or preexisting antibodies against heart allograft

CLINICAL ISSUES • Significantly variable incidence in different medical centers ○ Depends on detection protocol and rigor of criteria utilized on surveillance biopsies • Most common within 1st month ○ May occur months to years later • AMR treatment includes modalities intended to suppress production and enhance removal of antibodies • C4d/C3d deposition correlates with increased risk of graft loss and cardiac allograft vasculopathy

MICROSCOPIC • Currently no consensus on grading system • Signs of tissue injury include ○ Capillary endothelial swelling or denudation

○ Interstitial edema, mild pericapillary neutrophilic infiltration, ± hemorrhage ○ Intravascular thrombi, neutrophils, or macrophages • Immunological evidence of AMR includes ○ Diffuse positivity for C4d &/or C3d in > 50% of capillary endothelium by immunofluorescence (IF) performed on frozen tissue ○ Diffuse and strong staining for C4d &/or C3d by IHC immunoperoxidase method on formalin-fixed, paraffinembedded tissues ○ Interstitial and serum staining is not considered to be positive

TOP DIFFERENTIAL DIAGNOSES • Diffuse mild cellular rejection

DIAGNOSTIC CHECKLIST • H&E findings are nonspecific, and diagnosis often requires IF or IHC staining

Histologic Features of AMR

C4d Immunohistochemistry

C4d Immunofluorescence

Background C4d Immunohistochemistry

(Left) Histologic changes seen in AMR are subtle and include endothelial swelling ﬈ and intravascular lymphocytes and macrophages ﬊. Myocyte necrosis and hemorrhage are not generally present. (Right) Immunohistochemical study for C4d shows strong and diffuse endothelial staining of all capillaries ﬊. Note the arteriole, which is also positive ﬈. The staining is linear and lines each vessel.

(Left) Immunofluorescence staining for C4d also shows strong endothelial staining in this heart biopsy ſt, a pattern similar to that seen in corresponding IHC staining. (Right) This immunohistochemical stain for C4d shows extensive background staining of interstitium as well as serum staining and should be interpreted as being negative.

346

Antibody-Mediated Rejection, Heart

Abbreviations • Antibody-mediated rejection (AMR)

Synonyms • Humoral rejection

Definitions • Rejection targeting microvasculature and capillaries due to either de novo antibody formation, or preexisting antibodies against heart allograft • Hyperacute rejection attributed to AMR

ETIOLOGY/PATHOGENESIS Proposed Mechanisms • Deposition of alloantibodies within graft coronary vasculature followed by complement activation • Alloantibodies directed against class I and II major histocompatibility antigens (MHC or HLA) ○ Target MHC molecules on capillary and larger vessel endothelium with subsequent complement activation, endothelial damage, and cell death ○ Can also activate coagulation cascade resulting in microvascular thrombi ○ May also involve other non-HLA antigens

CLINICAL ISSUES Epidemiology • Significantly variable incidence in different medical centers ○ Depends on detection protocol and rigor of criteria utilized on surveillance biopsies • More common in allosensitized individuals with high panel reactive antibodies, positive crossmatch, or presence of donor-specific antibodies • Risk factors for allosensitization include ○ Female gender (due to pregnancy, especially if multiparous) ○ Multiple blood transfusions ○ Left ventricular assist device (LVAD) placement ○ Previous organ transplantation ○ Cytomegalovirus seropositivity (triggers formation of antiendothelial antibodies)

Prognosis • C4d/C3d deposition correlates with increased risk of graft loss and cardiac allograft vasculopathy • AMR also associated with increased risk of mortality ○ Demonstrated only with strong endothelial staining for C4d or C3d

MACROSCOPIC

Heart Transplantation

TERMINOLOGY

General Features • Edematous, swollen heart with foci of myocardial discoloration and hemorrhage ○ Rare finding at autopsy or retransplant

MICROSCOPIC Histologic Features • No current consensus on grading system, but ISHLT moving toward stratification based on presence/absence of histologic &/or immunopathologic findings • Histologic signs of tissue injury include ○ Capillary endothelial swelling or denudation ○ Interstitial edema, mild pericapillary neutrophilic infiltration, ± hemorrhage ○ Intravascular thrombi, neutrophils, or macrophages • Immunological evidence of AMR includes ○ Diffuse positivity for C4d &/or C3d in > 50% of capillary endothelium by immunofluorescence (IF) performed on frozen tissue ○ Diffuse and strong staining for C4d &/or C3d by IHC immunoperoxidase method on formalin-fixed, paraffinembedded tissues ○ Interstitial and serum staining is not considered positive – Considered background staining – In 80-90% of posttransplant heart biopsies ○ After treatment, most follow-up biopsies stay positive, but weaker and focal, for up to 3 weeks • Ancillary evidence of anti-HLA antibodies at time of endomyocardial biopsy • AMR only rarely associated with acute cellular rejection

DIFFERENTIAL DIAGNOSIS Diffuse Mild Cellular Rejection • Negative for C4d and C3d staining

Presentation • Most common within 1st month; may occur within few hours (hyperacute rejection) to 7 days in presensitized individuals ○ Less commonly seen months to years after transplantation • May be asymptomatic (termed "subclinical AMR") • Often signs of right/left ventricular dysfunction &/or hemodynamic compromise

Treatment • Poor response to most immunosuppressive therapy, which primarily targets cell-mediated rejection • Suppression of B-cell and plasma cell antibody production ○ Steroids, rituximab, bortezomib, splenectomy • Removal/inhibition of antibodies ○ Apheresis, intravenous immunoglobulins (IVIg)

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • H&E findings are nonspecific, and diagnosis often requires IF or immunohistochemical (IHC) staining • Only strong diffuse endothelial staining should be interpreted as positive

SELECTED REFERENCES 1. 2.

3.

Husain AN et al: Routine C4d immunohistochemistry in cardiac allografts: long-term outcomes. J Heart Lung Transplant. 36(12):1329-1335, 2017 Colvin MM et al: Antibody-mediated rejection in cardiac transplantation: emerging knowledge in diagnosis and management: a scientific statement from the American Heart Association. Circulation. 131(18):1608-39, 2015 Berry GJ et al: The ISHLT working formulation for pathologic diagnosis of antibody-mediated rejection in heart transplantation: evolution and current status (2005-2011). J Heart Lung Transplant. 30(6):601-11, 2011

347

Heart Transplantation

Chronic Allograft Vasculopathy, Heart KEY FACTS

TERMINOLOGY • Accelerated form of coronary disease characterized by uniform, circumferential intimal fibrosis

ETIOLOGY/PATHOGENESIS • Incidence correlates with severity and frequency of acute cellular rejection episodes • Antibody-mediated rejection greatly increases risk ○ Especially in pediatric patients • Other risk factors are older age, dyslipidemia, or atherosclerosis in donor

CLINICAL ISSUES • Incidence increases with time after transplantation • Seen in both adult and pediatric population • Often late presentation as patients do not experience anginal symptoms due to denervation of transplanted heart

• Various treatment modalities exist, but definitive therapy is retransplantation

MICROSCOPIC • Affects larger epicardial and smaller epicardial and intramyocardial arteries • Involvement characterized by concentric intimal proliferation of smooth muscle cells and spindled myofibroblasts • Elastin stain demonstrates relative sparing of elastic lamina

TOP DIFFERENTIAL DIAGNOSES • Atherosclerotic coronary artery disease ○ CAV affects smaller arteries and veins in addition to larger arteries – More circumferential – Intact internal elastic lamina – Usually lacks cholesterol clefts and calcification

Intimal and Medial Sclerosis

Chronic Rejection (Elastin)

Coronary Artery Vasculopathy

Coronary Artery Vasculopathy

(Left) Section of epicardial coronary artery shows marked intimal and medial fibrosis in this 68-year-old patient who underwent retransplantation for chronic rejection. (Right) Elastin stain demonstrates marked intimal fibrosis ﬊ and an intact internal elastic lamina ﬉ in this smaller branch of an epicardial coronary artery, also from a specimen with chronic rejection.

(Left) Medium-power view of coronary artery vasculopathy shows dense fibrosis of intima ﬈ and thinning of media ﬊. Note the lack of inflammatory cells. (Right) This section from a patient who died of chronic rejection demonstrates intimal fibrosis with a chronic inflammatory infiltrate, which can be seen in some cases of CAV. The arterial lumen is toward top left corner ﬈.

348

Chronic Allograft Vasculopathy, Heart

Abbreviations • Cardiac allograft vasculopathy (CAV)

Synonyms

○ Still widely used • Intravascular ultrasound considered most sensitive tool for CAV diagnosis, but invasive and lacks ability to evaluate entire coronary vasculature • Dobutamine stress echocardiogram can be useful, sensitive, noninvasive diagnostic screening modality

• Chronic rejection (CR)

Definitions • Rejection that results in accelerated form of coronary artery disease characterized by uniform, circumferential intimal fibrosis and subsequent obliteration of lumina

ETIOLOGY/PATHOGENESIS Immunologic Risk Factors • Incidence correlates with severity and frequency of acute cellular rejection episodes • Anti-HLA and antiendothelial antibodies associated with increased risk for CAV • Antibody-mediated rejection greatly increases risk of CAV ○ Especially in pediatric patients

Nonimmunological Risk Factors • Age: Adults at higher risk compared to children ○ Infants have lowest risk • Older age, dyslipidemia, or atherosclerosis in donor

CLINICAL ISSUES

MICROSCOPIC Histologic Features • Affects larger epicardial and smaller epicardial and intramyocardial arteries • Veins also affected but to much lesser extent • Involvement characterized by concentric intimal proliferation of smooth muscle cells and spindled myofibroblasts within extracellular matrix ○ Large epicardial coronary arteries show uniform involvement along entire length ○ Concentric intimal fibrosis most marked in intramyocardial and epicardial branches • Variable amounts of inflammatory infiltrate consisting of lymphocytes and foamy macrophages ○ May be seen ± endotheliitis and vasculitis • Distal myocardium may show signs of ischemia or infarction (patchy areas of fibrosis, coagulative necrosis) • None of above features specific for CAV

ANCILLARY TESTS

Epidemiology

Immunohistochemistry

• Incidence increases with duration of transplantation ○ Pediatric incidence: 2%, 9%, and 17% at 1, 5, and 10 years after transplantation, respectively ○ Adult incidence: 8%, 30%, and 50% at 1, 5, and 10 years after transplantation, respectively

• Trichrome and elastin stains better demonstrate intimal fibrosis and expansion ○ Elastin also demonstrates relative sparing of elastic lamina

Presentation • Often late as patients do not experience anginal symptoms due to denervation of transplanted heart • Other manifestations include cardiac arrhythmias, congestive heart failure, silent myocardial infarction, and even sudden death

Treatment • Implementation of statins or vasodilators as prophylaxis against progression if detected in early stages • Antiviral control of concurrent CMV infection (triggers endothelial inflammatory recruitment) • Increased immunosuppression • Percutaneous coronary intervention or bypass graft surgery • Definitive treatment remains retransplantation, although results generally suboptimal

Prognosis • Poor outcome and major indication for retransplantation • Remains leading cause of mortality > 1 year after transplantation

IMAGING Radiographic Findings • Difficult to visualize on coronary angiography because lesions result in uniform narrowing of arterial lumen

Heart Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Atherosclerotic Coronary Artery Disease • Narrowing by eccentric plaques composed of fibrosis, cholesterol clefts, and foamy macrophages ± calcifications • Disrupted internal elastic lamina with thinning of media

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Main differential diagnosis is atherosclerosis • CAV affects smaller arteries and veins in addition to larger arteries ○ More circumferential ○ Intact internal elastic lamina ○ Usually lacks cholesterol clefts and calcification

SELECTED REFERENCES 1. 2.

3. 4.

Merola J et al: Recent advances in allograft vasculopathy. Curr Opin Organ Transplant. 22(1):1-7, 2017 Prada-Delgado O et al: Prevalence and prognostic value of cardiac allograft vasculopathy 1 year after heart transplantation according to the ISHLT recommended nomenclature. J Heart Lung Transplant. 31(3):332-3, 2012 Savasta M et al: Immunology insights into cardiac allograft rejection. Rev Cardiovasc Med. 12(2):e68-76, 2011 Mehra MR et al: International Society for Heart and Lung Transplantation working formulation of a standardized nomenclature for cardiac allograft vasculopathy-2010. J Heart Lung Transplant. 29(7):717-27, 2010

349

Heart Transplantation

Quilty Lesions KEY FACTS

TERMINOLOGY • Endocardial infiltrate of small, mature lymphocytes that may extend into underlying myocardium

ETIOLOGY/PATHOGENESIS

• Quilty type B: Infiltration through underlying myocardium in nodular arrangement or through myocytes • Subtle myocyte damage • Occasionally fibrosis, plasma cells, eosinophils

ANCILLARY TESTS

• Etiology unknown

CLINICAL ISSUES • Incidental finding observed in 10-20% of surveillance endomyocardial biopsies performed in 1st year after transplantation • Self-limited lesion generally disappears within weeks or months after diagnosis • Presents issue for pathologist as it closely resembles acute cellular rejection (ACR)

MICROSCOPIC • Quilty type A: Nodular collection of mature, small lymphocytes with multiple capillaries confined to endocardium

• In larger lesions, CD21 highlights central follicular dendritic cell meshwork • Mixture of CD20(+) B cells and CD3(+) T cells

TOP DIFFERENTIAL DIAGNOSES • ACR • Previous biopsy site • Cardiac allograft vasculopathy

DIAGNOSTIC CHECKLIST • Generally centered on endocardium with prominent capillaries • In difficult cases, multiple levels and immunostaining for CD21 may differentiate from ACR

Quilty Type A Lesion

Quilty Type B Lesion

Quilty Type B Lesion With Fibrosis

Quilty Type B Lesion (CD21)

(Left) This right ventricular biopsy shows a noninvasive Quilty lesion ﬈ in the endocardium. The lesion is composed of small, mature lymphocytes that do not invade the underlying myocardium. (Right) In an invasive Quilty lesion, the mature lymphocytes extend from the endocardium into the underlying myocardium. Note multiple intralesional capillaries ﬈ and small, damaged myocytes ſt.

(Left) High magnification of invasive Quilty lesion shows a nodular infiltrate composed of mature lymphocytes associated with fibrosis but no capillaries. (Right) CD21 immunohistochemical stain highlights the follicular dendritic cells that form a central network in an invasive Quilty lesion.

350

Quilty Lesions

Abbreviations • Noninvasive Quilty type A (QA) • Invasive Quilty type B (QB)

Synonyms • Quilty effect (QE)

Definitions • Nodular endocardial infiltrate of small, mature lymphocytes in transplanted heart ○ May extend into underlying myocardium • Incidental finding that creates challenge differentiating from morphologic changes associated with allograft rejection

ETIOLOGY/PATHOGENESIS Unknown Mechanism • Quilty lesions have distinct molecular phenotype and correlate with impaired left ventricular ejection fraction ○ Suggests they may represent form of acute rejection ○ No proven link to infection, drug reaction, or rejection ○ Further studies required

CLINICAL ISSUES Presentation • Incidental finding observed in 10-20% of surveillance endomyocardial biopsies performed in 1st year after transplantation • Subsequent biopsies have persistent lesion for several weeks or months • Depending on site of biopsy, lesion may not be seen in subsequent biopsy but may reappear later

Treatment • No treatment or immunosuppression necessary

Prognosis • Self-limited lesion ○ Generally disappears within weeks or months after diagnosis • Some studies conclude that QE can herald episode of acute rejection

MICROSCOPIC Histologic Features • Nodular endocardial infiltrate of mature, small lymphocytes seen on both protocol-driven and symptomatic cardiac biopsies ○ QA when confined to endocardium without extension into myocardium ○ QB when lesion infiltrates underlying myocardium, ± myocyte damage – Invasive QB may be nodular or infiltrative between myocytes – May be associated with fibrosis ○ Obvious at low magnification and present on multiple levels

• Rare plasma cells and occasional eosinophils may be present • Usually contains several small capillaries • Subtle myocyte damage ○ Can be recognized by presence of irregular &/or small myocytes • In explanted transplants and autopsies, Quilty lesions can be in deep myocardium and epicardium ○ Can be single but usually multiple

Heart Transplantation

TERMINOLOGY

ANCILLARY TESTS Immunohistochemistry • Quilty lesion consists of mixture of CD20(+) B and CD3(+) T cells, respectively ○ Most B cells arranged centrally and T cells at periphery ○ Immunohistochemistry for B and T cells not helpful for distinguishing from mimics • In larger Quilty lesions, CD21 stain highlights central follicular dendritic cell meshwork, which confirms diagnosis

DIFFERENTIAL DIAGNOSIS Acute Cellular Rejection • • • •

Consists of larger activated lymphocytes More likely to be associated with eosinophils Fewer associated capillaries Immunostains show significant number of CD68(+) macrophages, CD3(+) T cells, and sparse CD20(+) B cells • Absence of CD21(+) follicular dendritic cell meshwork • QB lesions often confused with ACR ○ If tangential sectioning does not demonstrate relationship to endocardial surface ○ Deeper sections may demonstrate continuity with endocardium in QB • Both Quilty lesion and ACR may be present in same biopsy ○ In this case, Quilty lesion does not upgrade rejection

Previous Biopsy Site • Organizing fibrin and granulation tissue with few inflammatory cells and macrophages

Cardiac Allograft Vasculopathy • Lymphocytic infiltrates often associated with cardiac allograft vasculopathy in epicardial coronary arteries • Should not be confused with Quilty lesion

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Generally centered on endocardium with prominent capillaries • In difficult cases, multiple levels (to demonstrate continuity of invasive lesion with endocardial surface) and ○ Immunostain for CD21(+) dendritic cell meshwork may differentiate from ACR

SELECTED REFERENCES 1.

2.

Mozaffari K et al: Quilty effect: its definition and significance, in post-cardiac transplant endomyocardial biopsies with focus on our experience. Multidiscip Cardio Annal. 8(1):e9615, 2017 Sattar HA et al: The presence of a CD21+ follicular dendritic cell network distinguishes invasive Quilty lesions from cardiac acute cellular rejection. Am J Surg Pathol. 30(8):1008-13, 2006

351

Heart Transplantation

Site of Previous Biopsy KEY FACTS

• Changes due to prior biopsy at same site

• Deeper tissue sections may help reveal characteristic findings of previous biopsy site

ETIOLOGY/PATHOGENESIS

TOP DIFFERENTIAL DIAGNOSES

• Multiple endomyocardial biopsies are performed over time as part of posttransplant surveillance

• Acute cellular rejection ○ Should not have fibrin, granulation tissue, or hemosiderin ○ More lymphocytes in rejection that appear activated than at site of previous biopsy (SOPB) • Quilty lesion • Antibody-mediated rejection ○ C3d and C4d immunofluorescence or immunohistochemical studies also positively stain fibrin and necrotic myocytes • Postbiopsy perforation

TERMINOLOGY

CLINICAL ISSUES • Incidental finding on biopsy with no clinical implication

MICROSCOPIC • Findings depend on length of time since previous biopsy ○ Within 1 week of prior biopsy, fibrin commonly present on endocardial surface ○ Granulation tissue seen both in endocardium and for short distance into underlying myocardium ○ Prominent endothelial cells often seen in capillaries within granulation tissue • Special studies or stains are unnecessary ○ Since diagnosis based on histologic examination

DIAGNOSTIC CHECKLIST • Evaluate tissue away from SOPB for cellular rejection

Organizing Fibrin

Fibrin, Granulation Tissue and Fibrosis

Healing Biopsy Site

Healed Biopsy Site With Chronic Inflammation

(Left) Surveillance biopsy shows the typical histological appearance of site of previous biopsy with fibrin covering the endocardial surface ﬈ and granulation tissue between the fibrin and myocardium. (Right) The endocardial surface is shown on the right with a layer of fibrin. The underlying granulation tissue has only a few inflammatory cells. Note that fibrosis extends into the myocardium for a short distance.

(Left) Low-power view of an endomyocardial biopsy shows the site of previous biopsy older than those in previous images. Note lack of fibrin. Instead, there is fibrosis of endocardial surface. (Right) Higher power view shows dense fibrosis of the surface ﬈. A small cluster of mature lymphocytes and capillaries ſt is present, raising the differential of Quilty lesion.

352

Site of Previous Biopsy

Abbreviations • Site of previous biopsy (SOPB)

Synonyms

• Typically 1 biopsy fragment has previous biopsy site changes while other fragments do not ○ Changes seen in > 1 fragment occasionally • Nonspecific endomyocardial fibrosis common several months to years after transplantation ○ Cannot be distinguished from healed biopsy sites

• Biopsy site changes

Definitions • Histologic changes in endomyocardium due to prior biopsy at same site

ETIOLOGY/PATHOGENESIS

DIFFERENTIAL DIAGNOSIS Acute Cellular Rejection • Should not have fibrin, granulation tissue, or hemosiderin • More lymphocytes that appear activated than at SOPB • Eosinophils present in higher grades of rejection

Transplant Surveillance

Quilty Lesion

• Multiple endomyocardial biopsies performed over time as part of postheart transplant surveillance • Bioptome tends to follow same path toward right side of interventricular septum ○ Thus same site likely to be biopsied again • Findings of healing biopsy site commonly seen in 1st few weeks after transplantation when biopsies more frequently performed (often weekly)

• Endocardial collection of small mature lymphocytes often with small capillaries • May extend into underlying myocardium • Larger lesions usually have central dendritic cell network (positive for CD21)

Follow-Up Biopsy • After episode of significant rejection, follow-up biopsy often performed 1 week later and likely to show SOPB changes

CLINICAL ISSUES Presentation • Incidental finding on biopsy with no clinical implication

Treatment • None

Prognosis • Inconsequential finding

MICROSCOPIC Histologic Features • Findings depend on length of time since previous biopsy • Within 1 week of prior biopsy, fibrin commonly present on endocardial surface ○ Granulation tissue seen both in endocardium and for short distance into underlying myocardium ○ Prominent endothelial cells often seen in capillaries within granulation tissue ○ Macrophages and few inflammatory cells are present in association with granulation tissue – Usually few scattered lymphocytes seen – Rare presence of other cells, such as eosinophils or plasma cells • Older biopsy site changes include more fibrosis and less granulation tissue ○ Hemosiderin may also be present – Helps distinguish SOPB from acute cellular rejection • Special studies or stains are unnecessary ○ Diagnosis based on histologic examination • Deeper tissue sections may help reveal characteristic findings of previous biopsy site

Heart Transplantation

TERMINOLOGY

Antibody-Mediated Rejection • C4d capillary deposition ○ C3d and C4d immunofluorescence or immunohistochemical studies also positively stain fibrin and necrotic myocytes – Use caution when interpreting these stains – Do not confuse positive staining of previous biopsy site with strong endothelial staining as would be seen in antibody-mediated rejection

Postbiopsy Perforation • Reactive mesothelial cells and fibroblasts are seen when perforation has occurred during biopsy • If histology alone is insufficient to differentiate mesothelial cells from reparative changes of SOPB, immunohistochemical stains are helpful ○ Staining panel should include at least 1 mesothelial marker (calretinin, WT-1, or cytokeratin 5/6) and 1 endothelial marker (CD31 or CD34)

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Granulation tissue and fibrin are key findings indicating SOPB • Evaluate tissue away from SOPB for cellular rejection

SELECTED REFERENCES 1. 2. 3.

4.

5. 6.

Thiene G et al: Diagnostic use of the endomyocardial biopsy: a consensus statement. Virchows Arch. 463(1):1-5, 2013 From AM et al: Current status of endomyocardial biopsy. Mayo Clin Proc. 86(11):1095-102, 2011 Stewart S et al: Revision of the 1990 working formulation for the standardization of nomenclature in the diagnosis of heart rejection. J Heart Lung Transplant. 24(11):1710-20, 2005 Veinot JP: Diagnostic endomyocardial biopsy pathology--general biopsy considerations, and its use for myocarditis and cardiomyopathy: a review. Can J Cardiol. 18(1):55-65, 2002 Winters GL: The challenge of endomyocardial biopsy interpretation in assessing cardiac allograft rejection. Curr Opin Cardiol. 12(2):146-52, 1997 Rose AG et al: Endomyocardial biopsy site morphology. an experimental study in baboons. Arch Pathol Lab Med. 110(7):622-5, 1986

353

Heart Transplantation

Myocarditis KEY FACTS

TERMINOLOGY • Pathologically subclassified as ○ Lymphocytic ○ Eosinophilic ○ Giant cell ○ Granulomatous

ETIOLOGY/PATHOGENESIS • Viral agents are most common cause of lymphocytic myocarditis in developed countries

MICROSCOPIC • Low sensitivity of endomyocardial biopsy due to focal nature of infiltrates • Dallas criteria for lymphocytic myocarditis require identification of inflammation and associated myocyte injury ○ Borderline myocarditis has no myocyte injury and less prominent inflammation

• Eosinophilic myocarditis predominantly interstitial and perivascular with mixed infiltrate ○ Typically little myocyte damage or fibrosis in cases due to hypersensitivity • Giant cell myocarditis usually diffuse and multifocal with mixed infiltrate of giant cells, lymphocytes, and eosinophils ○ Giant cells usually at edge of inflammation ○ Eosinophils may be prominent ○ Prominent necrosis and tissue damage

TOP DIFFERENTIAL DIAGNOSES • Normal heart has < 5 lymphocytes per HPF • Cross sections of fibroblasts or endothelial cells should not be mistaken for inflammatory cells • Edema can be biopsy artifact and does not indicate myocyte damage • Sarcoidosis has well-formed, noncaseating granulomas with limited inflammatory infiltrates

Explanted Heart With Dilated Cardiomyopathy

Lymphocytic Myocarditis

Giant Cell Myocarditis

Recurrent Giant Cell Myocarditis

(Left) Explanted heart from a patient with lymphocytic myocarditis shows a markedly dilated left ventricle with a thin ventricular free wall ﬇. This patient was transplanted for a clinical diagnosis of dilated cardiomyopathy. (Right) High-power view of an explanted heart with lymphocytic myocarditis demonstrates a prominent lymphocytic infiltrate with focal encroachment ﬊ on cardiac myocytes.

(Left) Endomyocardial biopsy from a young adult with giant cell myocarditis shows extensive destruction of the myocardium with a mixed inflammatory infiltrate, including giant cells ﬈ and lymphocytes. (Right) Highpower view of recurrent giant cell myocarditis after transplant shows multiple giant cells ﬈, eosinophils ﬊, and myocyte damage ﬉.

354

Myocarditis

Definitions • Inflammatory disease of myocardium • Pathologically subclassified as ○ Lymphocytic ○ Eosinophilic ○ Giant cell ○ Granulomatous • Different clinical associations with each myocarditis subtype

ETIOLOGY/PATHOGENESIS Infectious Agents • Viruses are most common cause of lymphocytic myocarditis in developed countries ○ Coxsackie B virus historically among most commonly implicated viruses ○ Parvovirus B19, HHV-6, and adenovirus are most common agents detected by PCR in recent case series ○ In new onset dilated cardiomyopathy without histologic evidence of myocarditis, PCR may detect viral genetic material, but significance is controversial ○ Neonates and immunosuppressed patients may be at higher risk of developing myocarditis with viral infection ○ Proposed mechanisms of damage include – Direct virus injury – Persistent viral replication – Autoimmune reactions after viral clearance • Chagas disease is common cause in South America • Toxoplasmosis may cause myocarditis in immunosuppressed patients or in seronegative heart transplant patients receiving heart from seropositive donor ○ Other infections in transplant patients usually affect other organ systems or have severe systemic disease with heart involvement and are not diagnosed on biopsy or explant • Bacterial and fungal myocarditis occur but are rare • Parasitic infections are associated with eosinophilic myocarditis

Connective Tissue Diseases • Associated most commonly with systemic lupus erythematosus, dermato- or polymyositis ○ Usually lymphocytic myocarditis ○ Can be giant cell myocarditis

Hypersensitivity • Classically associated with drugs, such as dopamine, dobutamine, sulfonamides, and clozapine ○ Often in heart failure patients on multiple drugs ○ Identification of single responsible drug may not be possible • Not related to drug dosage • Not seen in response to fungal antigens or other allergens implicated in hypersensitivity of other organs

Hypereosinophilic Syndromes • Associated with eosinophilic myocarditis and extensive myocyte damage

Idiopathic • Etiology of some cases may not be determined

CLINICAL ISSUES Epidemiology • Incidence ○ Lymphocytic myocarditis accounts for ~ 10% of biopsies from patients with new onset cardiac dysfunction – Cause of sudden cardiac death in 5-10% of adults – Many subclinical cases of viral myocarditis ○ Hypersensitivity myocarditis in < 7% of explanted hearts – Other causes of eosinophilic myocarditis (parasitic, hypereosinophilic syndromes, eosinophilic leukemia, Churg-Strauss syndrome) are extremely rare ○ Giant cell myocarditis is extremely rare – Usually affects young to middle-aged adults but reported in children and elderly

Heart Transplantation

TERMINOLOGY

Presentation • Viral myocarditis often subclinical ○ Acute heart failure is most common presentation ○ Epicardial involvement can cause chest pain and pericardial effusion ○ Varies from acute (several weeks) to chronic with dilated cardiomyopathy – Usually presents late in infectious disease course, often with autoimmune component ○ Fulminant lymphocytic myocarditis defined as presenting < 2 weeks of viral infection with severe hemodynamic compromise • Hypersensitivity myocarditis ○ May have temporal association with drug use – Days to months after initial drug use ○ Difficult to identify in patients already in heart failure ○ May be accompanied by fever, peripheral eosinophilia, or rash – Signs may be absent or transient • Giant cell myocarditis most commonly presents as heart failure, arrhythmias, or heart block ○ Rapid onset presentation – Diagnosis made within weeks of symptom onset

Treatment • Avoidance of exercise • Viral myocarditis ○ Primarily supportive therapy – Immunosuppressive therapy for viral myocarditis is controversial – Antiviral therapy typically not useful • Hypersensitivity myocarditis ○ Steroid therapy and discontinuation of offending drug may be helpful – Often not possible to determine single causative drug • Giant cell myocarditis and myocarditis associated with connective tissue disease ○ Immunosuppression is beneficial

Prognosis • Fulminant viral myocarditis ○ Majority survive and recover within weeks ○ Complete functional and histologic recovery 355

Heart Transplantation

Myocarditis • Acute viral myocarditis ○ Recovery depends on degree of hemodynamic compromise ○ May resolve or progress to dilated cardiomyopathy • Hypersensitivity myocarditis does not affect posttransplant survival ○ Hypersensitivity reaction often not seen in posttransplant biopsies despite continuation of drug – Possibly due to posttransplant immunosuppression • Giant cell myocarditis ○ < 6-month average survival if untreated ○ Average survival to transplant may be > 1 year with immunosuppression – 20% recurrence in heart transplant patients – Recurrence often asymptomatic and detected only in surveillance biopsy

IMAGING Ultrasonographic Findings • Echocardiography is nonspecific ○ Classically demonstrates global left ventricular dysfunction ± pericardial effusion

MR Findings • Myocardial edema and injury may manifest as focal or global increases in enhancement, T2 signal intensity, or late gadolinium enhancement

MACROSCOPIC

DIFFERENTIAL DIAGNOSIS

General Features

Normal Heart

• Usually nonspecific • End-stage myocarditis grossly similar to other causes of dilated cardiomyopathy

• < 5 lymphocytes per HPF ○ Cross sections of fibroblasts or endothelial cells should not be mistaken for inflammatory cells • Edema can be biopsy artifact ○ Does not indicate myocyte damage

MICROSCOPIC

356

○ Prominent eosinophilic infiltration, predominantly interstitial and perivascular – Often mixed with lymphocytes, plasma cells, and neutrophils – Varies from mild and focal to diffuse and prominent ○ Typically little myocyte damage or fibrosis in hypersensitivity myocarditis – When present, myocyte damage usually manifests as myocytolysis and is associated with more severe inflammation – Vasculitis without necrosis can be present ○ Myocyte necrosis and fibrosis may be prominent in eosinophilic myocarditis associated with infection or hypereosinophilic syndromes (necrotizing eosinophilic myocarditis) ○ If giant cells or marked necrosis are present with eosinophils, consider diagnosis of giant cell myocarditis • Giant cell myocarditis is usually diffuse and multifocal ○ Mixed infiltrate of lymphocytes and giant cells – Giant cells are usually at edge of inflammation – Eosinophils may be prominent ○ Necrosis and tissue damage are prominent ○ Very poorly formed granulomas may be present, but well-formed granulomas are not consistent with giant cell myocarditis ○ Fibrosis usually not present unless patient has been treated

Histologic Features

Myocardial Infarction

• Low sensitivity of endomyocardial biopsy due to focal nature of infiltrates ○ False-negative rate for lymphocytic myocarditis is 55% – Targeting biopsies to region of abnormality on MR may increase sensitivity – Negative biopsy does not exclude diagnosis ○ Biopsy for giant cell myocarditis has sensitivity of 68-85% – Sensitivity increased by rebiopsy if high suspicion • Dallas criteria for lymphocytic myocarditis require identification of inflammation and associated myocyte injury ○ Myocyte injury may include necrosis, myocytolysis (vacuolization), or irregular/scalloped cell borders due to lymphocytic infiltration ○ High degree of interobserver variability in interpretation of lymphocytic infiltrates – Normal myocardium averages < 5 lymphocytes per HPF – By immunohistochemistry, < 15 lymphocytes or macrophages per mm² is normal ○ Dallas criteria for borderline myocarditis defined as less prominent inflammation without myocyte destruction • Eosinophilic myocarditis

• Small biopsy from edge of organizing infarction may have scattered lymphocytes and damaged myocytes ○ Usually contains mixture of lymphocytes, histiocytes, plasma cells, and granulation tissue with hemosiderin • Usually spares endocardium and immediately subjacent myocardium

Prior Biopsy Site • Requires prior biopsy history • Inflammation is component of granulation tissue ○ Also includes fibrin, histiocytes, and fibrosis

Sarcoidosis (vs. Giant Cell Myocarditis) • Well-formed, noncaseating granulomas with limited inflammatory infiltrates in sarcoidosis ○ Sarcoid giant cells usually at granuloma center • More fibrosis and less necrosis in sarcoidosis • Eosinophils not prominent in sarcoidosis

Polyangiitis With Granulomatosis (Wegener) and Other Vasculitides • In addition to vasculitis, significant tissue necrosis and palisading granulomas should be present • Serological tests for p-ANCA or c-ANCA may be positive

Myocarditis

Viral

Bacteria

Fungal

Spirochete

Rickettsial

Helminthic

Protozoan

Adenovirus

Chlamydia

Actinomyces

Lyme disease

Q fever

Echinococcus

Leishmania

Coxsackievirus B

Cholera

Aspergillus

Leptospirosis

Rocky Mountain spotted fever

Paragonimus

Malaria

Echovirus

Diphtheria

Blastomyces

Relapsing fever

Typhus

Schistosoma

Toxoplasma

Epstein-Barr virus

Mycoplasma

Candida

Syphilis

Hepatitis C

Neisseria

Coccidioides

Trichinella spiralis

Human herpesvirus 6

Salmonella

Cryptococcus

Wuchereria

HIV

Staphylococcus

Histoplasma

Mumps virus

Streptococcus

Mucor

Parvovirus B19

Tetanus

Poliovirus

Tuberculosis

Taenia solium

Heart Transplantation

Infectious Causes of Myocarditis

Rabies virus Rubella virus Measles virus Varicella zoster virus Yellow fever virus

Noninfectious Causes of Myocarditis Hypersensitivity

Systemic Disorders

Toxins/Other

Antibiotics (penicillin, cephalosporins, sulfonamides, tetracycline)

Systemic lupus erythematosus

Alcohol

Clozapine

Polymyositis

Anthracyclines

Diuretics

Dermatomyositis

Arsenic

Insect/snake bites

Rheumatoid arthritis

Catecholamines

Lithium

Sarcoidosis

Heavy metals

Tetanus toxoid

Granulomatosis with polyangiitis (Wegener)

Radiation

Dopamine/dobutamine

Giant cell myocarditis Churg-Strauss syndrome Celiac disease Hypereosinophilia Inflammatory bowel disease Kawasaki disease Scleroderma Thyrotoxicosis

Acute Rheumatic Fever • Aschoff nodules ○ Perivascular and interstitial and composed of primarily collagen, macrophages, and lymphocytes ○ Multinucleated giant cells can be present ○ Well-formed granulomas and myocyte necrosis are not typical

SELECTED REFERENCES 1.

2. 3.

Lazaros G et al: Established and novel treatment options in acute myocarditis, with or without heart failure. Expert Rev Cardiovasc Ther. 15(1):25-34, 2017 Trachtenberg BH et al: Inflammatory cardiomyopathic syndromes. Circ Res. 121(7):803-818, 2017 Rose NR: Viral myocarditis. Curr Opin Rheumatol. 28(4):383-9, 2016

4.

Xu J et al: Giant cell myocarditis: a brief review. Arch Pathol Lab Med. 140(12):1429-1434, 2016 5. Kindermann I et al: Update on myocarditis. J Am Coll Cardiol. 59(9):779-92, 2012 6. Baughman KL: Diagnosis of myocarditis: death of Dallas criteria. Circulation. 113(4):593-5, 2006 7. Mills RM et al: Endomyocardial biopsy: a procedure in search of an indication. Am Heart J. 147(5):759-60, 2004 8. Takkenberg JJ et al: Eosinophilic myocarditis in patients awaiting heart transplantation. Crit Care Med. 32(3):714-21, 2004 9. Okura Y et al: A clinical and histopathologic comparison of cardiac sarcoidosis and idiopathic giant cell myocarditis. J Am Coll Cardiol. 41(2):322-9, 2003 10. Aretz HT et al: Myocarditis. a histopathologic definition and classification. Am J Cardiovasc Pathol. 1(1):3-14, 1987

357

Heart Transplantation

Myocarditis

Lymphocytic Myocarditis

Lymphocytic Myocarditis

Myocarditis CD3

Myocarditis CD8

Lymphocytic Myocarditis

Hypersensitivity Myocarditis

(Left) Low-power view of a section of the left ventricle from an explanted heart with lymphocytic myocarditis demonstrates a marked lymphocytic infiltrate ﬉ and fibrosis ﬈ disrupting bundles of myocytes. (Right) Highpower view of a section of explanted heart shows a more subtle lymphocytic infiltrate ﬈ between myocytes under a reactive endocardium ﬊. Definitive myocyte damage is not present in this focus, highlighting the difficulty of making the diagnosis on a small biopsy.

(Left) Immunohistochemistry for CD3 in this section of myocardium from an explanted heart demonstrates numerous positive T cells infiltrating between myocytes. (Right) Immunohistochemistry for CD8 shows that the majority of the T cells in this section from an explanted heart with lymphocytic myocarditis are cytotoxic T cells.

(Left) High-power view of an endomyocardial biopsy from a patient with recent onset heart failure demonstrates increased interstitial lymphocytes ﬈. Some lymphocytes may be encroaching on myocytes ﬉, but interobserver agreement for myocyte damage is low. (Right) Explanted heart from a patient with dilated cardiomyopathy treated with multiple drugs shows a predominantly interstitial pattern of inflammation ﬈ with mixed lymphocytes and eosinophils, consistent with hypersensitivity myocarditis.

358

Myocarditis

Hypersensitivity Myocarditis (Left) High-power view of hypersensitivity myocarditis demonstrates that the infiltrate is often perivascular ﬊ in addition to interstitial ﬈, and myocyte damage is usually not present. (Right) High-power view of hypersensitivity myocarditis shows increased eosinophils with fewer lymphocytes. The inflammation is predominantly interstitial with minimal myocyte damage ﬈ seen as encroachment on adjacent myocytes.

Giant Cell Myocarditis

Heart Transplantation

Hypersensitivity Myocarditis

Giant Cell Myocarditis (Left) Biopsy of giant cell myocarditis shows a marked infiltrate destroying the myocardium. There are rare giant cells ﬉ that are often at the infiltrate edge and do not form well-circumscribed granulomas. (Right) Highpower view of giant cell myocarditis shows the myocardium completely replaced by multiple giant cells ﬉ intermixed with lymphocytes and eosinophils. Other mononuclear cells are macrophages and few plasma cells.

Giant Cell Myocarditis

Toxoplasmosis (Left) In this focus of giant cell myocarditis, no giant cells are present. There is extensive myocyte injury ﬈. The numerous eosinophils intermixed with lymphocytes would be atypical for lymphocytic myocarditis. The presence of giant cells in other fragments of the biopsy confirms the diagnosis. (Right) This autopsied heart shows toxoplasmosis ﬈ within the myocardium in an immunocompromised patient (stem cell transplant). Immunohistochemistry can also be used to identify organisms in suspicious cases.

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SECTION 7

Lung Transplantation

Pathologic Classification of Lung Allograft Diseases Examination of Native and Transplanted Lungs History of Lung Transplantation

362 364 366

Evaluation of Failed Native Lung Emphysema Cystic Fibrosis Idiopathic Pulmonary Fibrosis Connective Tissue Disease-Associated Lung Disease Sarcoidosis, Lung Pulmonary Arterial Hypertension Other Causes of End-Stage Lung Disease

368 370 372 374 376 378 380

Surgical Complications Surgical Aspects and Complications, Lung

384

Allograft Rejection Pathologic Classification of Rejection Antibody-Mediated Rejection, Lung Acute Cellular Rejection, Grade A Acute Cellular Rejection, Grade B Chronic Allograft Dysfunction, Lung

386 388 390 392 394

Noninfectious Lesions 396 398

Organizing Pneumonia Microaspiration

Infections Bacterial Infections Viral Infections Fungal Infections, Lung

400 402 406

Lung Transplantation

Pathologic Classification of Lung Allograft Diseases

TERMINOLOGY Pathologic Classification • Based on pathogenesis, divided into categories of alloimmune and nonalloimmune

Abbreviations • • • •

Acute cellular rejection (ACR) Antibody-mediated rejection (AMR) Bronchiolitis obliterans (BO) Bronchiolitis obliterans syndrome (BOS)

• Having mixture of inflammatory cells is characteristic for rejection • Infection is most important clinical and pathological differential diagnosis ○ Infection has predominantly neutrophils • Bronchial-associated lymphoid tissue (BALT) often misinterpreted as rejection ○ BALT is nodular and is composed of mostly lymphocytes ○ CD21 highlights follicular dendritic cell network in BALT • In recipients who survive > 1 year after transplant, death most often associated with chronic rejection

Definitions

Chronic Rejection

• Single lung transplantation ○ Unilateral excision of diseased lung with least pulmonary reserve and replacement with allograft lung • Bilateral sequential lung transplantation ○ Excision of both diseased lungs and replacement with allograft lungs and separate perihilar airway anastomoses • Lobar lung transplantation ○ Unilateral excision of diseased lung and replacement with bilateral lower lobes from living donor • Heart-lung transplantation ○ En-bloc excision of diseased heart and bilateral lungs and replacement with allograft heart-lung bloc • Native lung ○ Recipient's lung removed for transplantation and examined as surgical specimen • Transplanted or allograft lung ○ Lung transplanted into patient for any length of time, which may be examined at retransplantation or postmortem • Explanted lung ○ Surgically removed native or allograft lung • ACR ○ T-cell-mediated rejection affecting parenchymal vasculature &/or small airways ○ Can occur few days to years after transplantation • Chronic rejection ○ Also called BO ○ T-cell-mediated rejection affecting larger airways or vessels ○ Seen as early as 3 weeks to many years • AMR ○ Alloantibodies targeting microvasculature and capillaries • BOS ○ Clinical correlate to chronic airway rejection

• Grade C: Chronic airway rejection (BO) ○ Associated with decrease in FEV1 ○ Usually clinical diagnosis ○ Transbronchial biopsy is insensitive; wedge biopsy necessary for diagnosis • Grade D: Chronic vascular rejection ○ Better diagnosed on wedge lung biopsy

ALLOIMMUNE RESPONSE Acute Cellular Rejection • Grade A: Perivascular inflammation ○ Composed of activated lymphocytes, eosinophils • Grade B: Small airways inflammation [lymphocytic bronchiolitis (LB)] ○ Controversial whether this can be distinguished from airway infection histologically ○ LB has band-like mononuclear cell infiltrate ○ Low (but improving) diagnostic reproducibility

Antibody-Mediated Rejection • Humoral rejection ○ Rare in lung transplant recipients • Histologic characteristics are varied but may include neutrophilic capillaritis • Clinical diagnosis of AMR based on graft dysfunction and donor-specific antibodies • C3d and C4d deposition ○ Thought to occur in lung as in other solid organ allografts ○ May be identified by immunohistochemistry or immunofluorescence ○ May not be clinically useful due to background staining and overall nonspecificity – Deposition can occur in infection, alveolar hemorrhage, septal damage, etc.

NONALLOIMMUNE DISEASES Primary Graft Dysfunction • Associated with ischemia-reperfusion injury, reactive oxygen species, and inflammation • Decreasing incidence due to improvements in allograft preservation techniques

Bronchial Anastomosis Stenosis • Fibrosis and narrowing at anastomotic site • Occurs within few months post transplant

Acute Graft Failure • Graft function loss of unknown etiology • Diagnosis of exclusion

Viral Infection (Partial List) • Cytomegalovirus (CMV) ○ Can precipitate ACR ○ CMV pneumonia has decreased due to better CMV status matching, prophylaxis, and viral load monitoring • Herpes simplex virus ○ Rare due to prophylaxis

Fungal Infection (Partial List) • Aspergillus

362

Pathologic Classification of Lung Allograft Diseases

Bacterial Infection (Partial List) • Vancomycin-resistant Enterococcus • Methicillin-resistant Staphylococcus aureus • About 70% of bacterial infections occur within 1st year after transplant • At least half of all lung transplant recipients die from infectious complications • Infectious complications are most common cause of death in patients with BO

Organizing Pneumonia • Reversible distal airway and alveolar proliferation of fibroblasts and myofibroblasts ○ Results in characteristic Masson bodies • Can be seen in resolution phase of ACR, infection, or post transplant ○ Nonspecific response to injury ○ Represents organization of any fibrinous exudate

Microaspiration • Microscopic aspiration of gastric contents usually associated with symptoms of gastroesophageal reflux disease ○ Can result in exogenous lipoid pneumonia • Multinucleated giant cells within alveoli or interstitium • Triggers episodes of ACR

Recurrent Primary Disease • Sarcoidosis ○ Commonly recurs and often asymptomatic ○ Well-formed nonnecrotizing granulomas are diagnostic • Lymphangioleiomyomatosis ○ Case reports of recurrence

Neoplasia • Posttransplant lymphoproliferative disorder ○ Most often EBV driven and B-cell derived ○ Increased risk associated with increasing levels of immunosuppression ○ > half of lung transplant recipients with PTLD have allograft involvement – Also may have gastrointestinal tract, liver, or lymph node involvement • Increased risk of nonmelanocytic skin malignancies ○ Squamous cell carcinoma more common than basal cell carcinoma ○ Poor prognosis compared to general population with similar type of cancer • Primary lung carcinoma ○ Incidence of ~ 6% in transplanted lungs

• Cystic fibrosis • Other ○ α-1-antitrypsin (A1AT) deficiency ○ Connective tissue disease-associated lung disease ○ Sarcoidosis ○ Pulmonary arterial hypertension • Rare ○ Lymphangioleiomyomatosis ○ Surfactant dysfunction disorder ○ Pulmonary alveolar proteinosis ○ Iatrogenic complications ○ Langerhans cell histiocytosis ○ BO due to hematopoietic stem cell transplantation ○ Primary or secondary carcinoma • Since 1996, stable number of single lung transplants per year in United States ○ Double lung transplants have steadily increased

Lung Transplantation

• Candida • Pneumocystis carinii pneumonia rare due to prophylaxis • Fungal infections occur in 15-35% of lung transplant recipients • High overall mortality rate of 40-80%

Common Pediatric Indications • Age: 6-17 years ○ Cystic fibrosis • Age: 1-5 years ○ Idiopathic pulmonary arterial hypertension ○ Idiopathic pulmonary fibrosis • Age: < 1 year ○ Surfactant protein B deficiency ○ Congenital heart disease ○ Idiopathic pulmonary arterial hypertension

SELECTED REFERENCES 1.

Husain AN et al: Lung transplantation: the state of the airways. Arch Pathol Lab Med. 140(3):241-4, 2016 2. Troxell ML et al: Practical applications in immunohistochemistry: evaluation of rejection and infection in organ transplantation. Arch Pathol Lab Med. ePub, 2016 3. Goldfarb SB et al: The Registry of the International Society for Heart and Lung Transplantation: eighteenth official pediatric lung and heart-lung transplantation report--2015; focus theme: early graft failure. J Heart Lung Transplant. 34(10):1255-63, 2015 4. Verleden GM et al: Current views on chronic rejection after lung transplantation. Transpl Int. 28(10):1131-9, 2015 5. Yusen RD et al: The Registry of the International Society for Heart and Lung Transplantation: thirty-second official adult lung and heart-lung transplantation report--2015; focus theme: early graft failure. J Heart Lung Transplant. 34(10):1264-77, 2015 6. Berry G et al: Pathology of pulmonary antibody-mediated rejection: 2012 update from the Pathology Council of the ISHLT. J Heart Lung Transplant. 32(1):14-21, 2013 7. Bhorade SM et al: Interobserver variability in grading transbronchial lung biopsy specimens after lung transplantation. Chest. 143(6):1717-24, 2013 8. Gordon IO et al: SaLUTaRy: survey of lung transplant rejection. J Heart Lung Transplant. 31(9):972-9, 2012 9. Stewart S et al: Revision of the 1996 working formulation for the standardization of nomenclature in the diagnosis of lung rejection. J Heart Lung Transplant. 26(12):1229-42, 2007 10. Yousem SA et al: Revision of the 1990 working formulation for the classification of pulmonary allograft rejection: Lung Rejection Study Group. J Heart Lung Transplant. 15(1 Pt 1):1-15, 1996

INDICATIONS FOR LUNG TRANSPLANTATION Common Adult Indications • Chronic obstructive pulmonary disease/emphysema (unrelated to α-1-deficiency) • Idiopathic pulmonary fibrosis 363

Lung Transplantation

Examination of Native and Transplanted Lungs KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Native lung: Patient's lung(s) removed for transplantation and examined as surgical specimen • Transplanted lung: Donor lung transplanted into patient for any length of time ○ Examined at retransplantation or postmortem

• Explanted native lung: Findings depend on primary pulmonary disease • May be present ○ Superimposed infection ○ Inflammation ○ Diffuse alveolar damage ○ Microaspiration ○ Organizing pneumonia • Transplanted lung: Findings depend whether cause of death was pulmonary or nonpulmonary • Recurrent primary disease rarely reported • Additional findings may be present depending on premortem course, similar to native lung above

CLINICAL ISSUES • Review clinical information and imaging

MACROSCOPIC • Adhesions likely present ○ Require careful dissection to preserve peripheral lung tissue and mediastinal structures • Examine anastomotic sites ○ Document whether intact or disrupted • Section in coronal plane • Collect lung tissue and blood for cultures • Minimum of 3 sections from each lobe

ANCILLARY TESTS • If infection present or suspected, appropriate stains for organisms should be done on multiple blocks

Pulmonary Fibrosis Post Transplantation

Pulmonary Fibrosis Post Transplantation

Sarcoidosis of lung

Cystic Fibrosis

(Left) Gross photograph of an explanted lung shows the medial (hilar) side st. Note marked cobblestone appearance of pleura ſt due to fibrosis. (Right) The same lung is shown here on cut section, after fixation. Note patchy but large areas of fibrosis ﬊ and traction bronchiectasis ﬈. Sections should be taken from center and periphery.

(Left) Lung from a patient with end-stage sarcoidosis shows irregular scarring and dilated air spaces ﬊ on cut section. Extensive sampling is needed to find granulomas. (Right) Explanted lung from a cystic fibrosis patient shows markedly dilated bronchi ﬇, which are filled with yellowgreen purulent material.

364

Examination of Native and Transplanted Lungs

Definitions • Native lung: Patient's lung(s) removed for transplantation and examined as surgical specimen • Transplanted lung: Donor lung transplanted into patient for any length of time ○ Examined at retransplantation or postmortem • Allograft lung: Synonymous with transplanted lung • Explanted lung: Surgically removed native or allograft lung

CLINICAL ISSUES Presentation • Review clinical information and imaging from medical record ○ Any stent placement ○ Focal or mass lesions ○ Suspicion of pulmonary embolism ○ Prior surgery ○ Any history of infection

Prognosis • Early mortality after lung transplantation due to ○ Undiagnosed acute rejection ○ Infection ○ Pulmonary thromboembolism ○ Surgical complications ○ Diffuse alveolar damage • Late mortality after lung transplantation ○ Most often due to infection ○ Less often due to complications of acute or chronic rejection or other medical conditions

MACROSCOPIC General Features • Adhesions likely present ○ Require careful dissection to preserve peripheral lung tissue and mediastinal structures • Document any stents • Examine anastomotic sites and document whether intact or disrupted • Examine pleura and hilum ○ Dissect lymph nodes • Open airways, artery, and vein, if clinically indicated • Section in coronal plane • Photograph gross specimen • Collect frozen tissue for research as prescribed by institutional protocols and standards • Collect tissue for electron microscopy, if indicated, especially in pediatric patients • Collect lung tissue and blood for cultures ○ Viral ○ Fungal ○ Aerobic and anaerobic bacteria

Sections to Be Submitted • Minimum of 3 from each lobe ○ Central to include large airway ○ Middle to include medium-sized airway

○ Peripheral to include pleura • Any additional areas of concern • Adequate sampling of native lung from single lung transplant, especially if left since only 2 lobes • Bronchial and vascular margins at hilum • Sample lymph nodes in specimen, including hilar and peribronchial

Lung Transplantation

TERMINOLOGY

MICROSCOPIC Histologic Features • Explanted native lung: Findings will depend on primary pulmonary disease ○ Emphysema ○ Cystic fibrosis ○ Idiopathic pulmonary fibrosis ○ Connective tissue disease-associated lung disease ○ Sarcoidosis ○ Pulmonary arterial hypertension ○ Superimposed infection, inflammation, diffuse alveolar damage, microaspiration, or organizing pneumonia may be present • Transplanted lung: Findings depend on whether cause of death was pulmonary or nonpulmonary ○ Allograft rejection ○ Infections ○ Posttransplant lymphoproliferative disorder ○ Recurrent primary disease rarely reported – Sarcoidosis: Presence of granulomas does not correlate with graft dysfunction – Lymphangioleiomyomatosis: Abnormal nodular proliferation of smooth muscle cells with cyst formation in females of child-bearing age – Langerhans cell histiocytosis: Nodular proliferation of Langerhans cells in adult smokers ○ Additional findings may depend on premortem course, similar to native lung above

ANCILLARY TESTS Special Stains • If infection present or suspected, appropriate stains for organisms should be done on multiple blocks ○ GMS (for fungus) ○ AFB (Ziehl-Neelsen for mycobacteria, Fite for Nocardia) ○ Viral immunohistochemical stains as indicated by morphology

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Correlate pathologic, clinical, and radiologic findings • Submit many sections for adequate sampling

SELECTED REFERENCES 1.

2. 3.

Ofek E et al: Restrictive allograft syndrome post lung transplantation is characterized by pleuroparenchymal fibroelastosis. Mod Pathol. 26(3):350-6, 2013 Akindipe OA et al: Discrepancies between clinical and autopsy diagnoses in lung transplant recipients. Clin Transplant. 24(5):610-4, 2010 Burns KE et al: Pulmonary embolism on postmortem examination: an underrecognized complication in lung-transplant recipients? Transplantation. 77(5):692-8, 2004

365

Lung Transplantation

History of Lung Transplantation

TERMINOLOGY Definitions • Single lung transplantation: Unilateral excision of diseased lung with least pulmonary reserve and replacement with allograft lung • Bilateral sequential lung transplantation: Excision of both diseased lungs and replacement with allograft lungs with separate perihilar airway anastomoses • Heart-lung transplantation: En bloc excision of diseased heart and bilateral lungs and replacement with allograft heart-lung bloc • Lobar lung transplantation: Unilateral excision of diseased lung and replacement with bilateral right and left lower lobes from living donor

•

•

LANDMARK EVENTS IN LUNG TRANSPLANTATION Timeline • Late 1890s ○ Dr. Alexis Carrel (Chicago) – Developed vascular suture techniques allowing functional vascular anastomosis – 1912: Awarded Nobel Prize "in recognition of his work on vascular suture and transplantation of blood vessels and organs" • 1947 ○ Dr. Vladimir Demikov (Moscow) – 1st transplant of lungs alone in canine □ Died of bronchial dehiscence • 1963 ○ Dr. James D. Hardy (Mississippi) – 1st successful human lung transplant, donated after circulatory death □ Immunosuppression consisted of preoperative thymic radiation and postoperative azathioprine and prednisone □ Patient died of uremia after 18 days, but lung allograft was normal at autopsy • 1964-1965 ○ Drs. Otto Gago and William E. Adams (Chicago) – 1st successful experimental transplantation of pulmonary lobe in 30 unrelated canines • 1971 ○ Dr. F. Derom (Belgium) – 1st successful single lung transplant for idiopathic pulmonary fibrosis (IPF) – 1st lung transplant patient discharged from hospital □ Survived 10.5 months • 1971-1974 ○ Earliest descriptions of acute cellular rejection in lung allografts • 1972-1973 ○ Dr. Frank J. Veith (New York) – Diagnosis (with x-ray and bronchoscopic biopsy) and reversal (with corticosteroid use) of rejection in canine and human lung allografts • 1974 ○ Dr. John R. Benfield (Wisconsin) 366

•

•

•

•

•

•

•

– Established lung biopsy as gold standard to distinguish allograft rejection from infection Late 1970s ○ National Institutes of Health moratorium on lung transplantation – Leading causes of death at that time: Rejection, infection, bronchial stenosis, impaired healing of bronchial anastomosis (airway dehiscence) 1981 ○ Dr. Bruce Reitz (California) – 1st successful heart-lung transplant performed in human ○ International Society for Heart and Lung Transplantation (ISHLT) founded – Multidisciplinary organization dedicated to improving care of patients with advanced heart or lung disease through transplantation, mechanical support, and innovative therapies via research, education, and advocacy 1981-1983 ○ Dr. Joel Cooper (Toronto) – Demonstrated bronchial anastomosis complications associated with use of prednisone, which was decreased by transition to cyclosporine 1983 ○ Dr. Joel Cooper – 1st successful long-term single-lung transplant, in IPF patient who survived 6 years 1985 ○ Dr. Axel Haverich (California) – Introduced concept of chronic lung allograft rejection by describing airway fibrosis in lung allografts of primates 1986 ○ Dr. Joel Cooper – 1st successful double-lung transplant in patient with emphysema; survived 14 years – Single-lung transplantation in 2 patients with pulmonary fibrosis – Concerns over using single lung due to (1) presumed risk of hyperinflation of native lung in emphysema and (2) risk of infection in cystic fibrosis 1988 ○ Dr. Joel Cooper – 1st successful double-lung transplant for cystic fibrosis 1989 ○ Dr. Hervé Mal (France) – Anticipated ventilation-perfusion imbalance with native lung hyperinflation did not occur in single lungs transplanted into 2 patients with end-stage emphysema 1990 ○ Dr. Elbert Trulock (Missouri) – Bilateral sequential double-lung transplant 1st described □ Separate bronchial anastomoses reduces ischemic airway complications □ Eliminates need for total cardiopulmonary bypass ○ Dr. Vaughn Starnes (California) – 1st descriptions of living-donor lobar lung transplant □ Especially useful for pediatric patients

History of Lung Transplantation

Year

Landmark Event

Pioneer

1946-1947

1st experimental lung transplant in dogs

Dr. Vladimir Demikov

1963

1st lung transplant in human described

Dr. James D. Hardy

1971

1st successful single lung transplant

Dr. F. Derom

1981

1st successful heart-lung transplant

Dr. Bruce Reitz

1986

1st en bloc double-lung transplant

Dr. Joel Cooper

1987

1st pediatric lung transplant

Dr. Joel Cooper

1990

1st bilateral sequential double-lung transplant

Dr. Elbert Trulock

1990

1st living donor lobar lung transplant

Dr. Vaughn Starnes

1990

Introduction of pathologic grading system for rejection

International Society for Heart and Lung Transplantation

2001

1st lung transplant after ex vivo lung perfusion

Dr. Stig Steen

2005

Introduction of lung allocation score

Organ Procurement and Transplantation Network

2005

Introduction of ECMO for pre- and post transplant

--

•

•

•

•

•

•

○ Organ Procurement and Transplantation Network (OPTN) begins lung allocation based on wait time with appropriate size and blood group matching – No special status given for clinical urgency 1995 ○ Dr. Hiroshi Date (Missouri) – Reports improvement in airway complication rate in 229 patients: From 10.9% (1988) to 4.0% (1994) ○ Dr. Robert J. Keenan (Pennsylvania) – Reports superiority of tacrolimus compared to cyclosporine as primary immunosuppressive agent after lung transplantation due to association with decreased risk of bronchiolitis obliterans syndrome (BOS) 1999-2001 ○ Department of Health and Human Services issues final rule – Requires inclusion of medical urgency as primary determinant of organ allocation – Discourages use of waiting time 2000-2001 ○ Dr. Evan S. Garfein (New York) – Demonstrated superiority of end-to-end vs. telescoped bronchial anastomosis 2005 ○ Use of extracorporeal membrane oxygenation as bridge to and after transplantation ○ OPTN implements new lung allocation score – Incorporates measures of wait list urgency and posttransplant survival 2006 ○ Dr. Shaf Keshavjee (Toronto) – Implements Novalung interventional lung assist device as bridge to transplant 2011 ○ Use of acellular normothermic ex vivo lung perfusion demonstrated to improve allograft preservation after harvesting ○ Ongoing studies to determine efficacy of newer immunotherapy (e.g., belatacept) in lung transplantation for patients who have failed calcineurin inhibitors

Lung Transplantation

Timeline of Significant Events in Lung Transplantation

• Recipient issues ○ Most common primary indications for lung transplant from 1995-2013 were chronic obstructive pulmonary disease, followed by interstitial lung disease, then cystic fibrosis ○ Primary graft dysfunction (PGD): Most common cause of death (~ 25% rate) within 1st month post transplant, according to ISHLT – Similar outcomes between recipients with PGD receiving lungs ± extracorporeal perfusion ○ Antimicrobial prophylaxis improved – Pneumonia remains persistent problem ○ Functional BOS improvement with azithromycin therapy ○ Donor-recipient matching improved – Remains ongoing long-term issue • Donor issues ○ Major ongoing limitation is shortage of donors and organs ○ Organ recovery rate improved due to standardized protocols for donor management • Survival ○ Median survival progressively improving – 6.1 years (2004-2011) vs. 5.3 (1996-2003) and 3.9 (1988-1995) years ○ Current survival rate: 79% at 1 year; 53% at 5 years ○ Double-lung survival better than single lung – Median 6.7 years vs. 4.6 years, respectively ○ Younger patients survive ~ 3 years longer than older patients

SELECTED REFERENCES 1. 2.

3.

4.

Vigneswaran et al: Lung Transplantation: Principles and Practice, Vol. 243. 2nd edition CRC Press, 2015 Yusen RD et al: The registry of the International Society for Heart and Lung Transplantation: thirty-second official adult lung and heart-lung transplantation report--2015; focus theme: early graft failure. J Heart Lung Transplant. 34(10):1264-77, 2015 Yusen RD et al: The registry of the International Society for Heart and Lung Transplantation: thirteenth adult lung and heart-lung transplant report-2013; focus theme: age. J Heart Lung Transplant. 32(10):965-78, 2013 Kotloff RM et al: Lung transplantation. Am J Respir Crit Care Med. 184(2):159-71, 2011

367

Lung Transplantation

Emphysema KEY FACTS

TERMINOLOGY

CLINICAL ISSUES

• Chronic obstructive pulmonary disease (COPD) • Type of COPD with irreversible alveolar wall destruction and overinflation without gross fibrosis • Centriacinar emphysema affects proximal acini (respiratory bronchioles), spares distal acini (terminal bronchioles), and associated with smoking • Panacinar emphysema affects both respiratory and terminal bronchioles and associated with genetic α-1antitrypsin deficiency • Bullae seen in either type

• Survival rate for lung transplantation for COPD comparable to lung volume reduction surgery

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• Increased levels of proteases and decreased levels of antiprotease result in overall tissue destruction • Destruction of alveolar elastic tissue leads to decreased radial traction and collapse of respiratory bronchioles during expiration (functional airflow obstruction)

• • • • •

Enlarged airspaces Free-floating alveolar septa Nodular tips at ends of alveolar septa Patchy nonspecific interstitial fibrosis may be seen Secondary pulmonary hypertensive changes

Emphysema, Microscopic

Anthracosis in Emphysema

Emphysema With Peripheral Fibrosis

Secondary Pulmonary Arterial Hypertension

MACROSCOPIC • Centriacinar emphysema ○ Enlarged airspaces at centers of lobules; upper-lobe predominant • Panacinar emphysema ○ Enlarged airspaces throughout entire lobule; lower-lobe predominant

(Left) Low-power photomicrograph of emphysema shows large, irregular acini. Note intraalveolar, lightly pigmented "smoker's" macrophages ﬊, which are often present in emphysema, and septa with nodular tips ﬈. (Right) Medium-power photograph of emphysema demonstrates large airspaces. There is a significant amount of black anthracotic pigment and only minimal fibrosis. Note free-floating alveolar septa ﬉.

(Left) In some patients with emphysema, there may be focal fibrosis and chronic inflammation, especially in the periphery, as illustrated in this low-power image. (Right) This patient with emphysema also has pulmonary arterial hypertensive changes. There is irregular intimal fibrosis.

368

Emphysema

Definitions

Prognosis

• Emphysema ○ Type of COPD with irreversible alveolar wall destruction and overinflation without gross fibrosis • Centriacinar emphysema affects proximal acini (respiratory bronchioles), spares distal acini (terminal bronchioles), and associated with smoking • Panacinar emphysema affects both respiratory and terminal bronchioles and associated with genetic α-1antitrypsin deficiency

• Lung transplantation ○ 1-year survival in 78% of patients and 5-year survival in 50% of patients ○ Long-term survival is better for pretransplant diagnosis of α-1-antitrypsin deficiency than COPD ○ Survival for lung transplantation for COPD comparable to LVRS

Synonyms

ETIOLOGY/PATHOGENESIS Protease-Antiprotease Imbalance and Reactive Oxygen Species • Nicotine and reactive oxygen species (free radicals) in tobacco smoke lead to accumulation of alveolar macrophages • Alveolar macrophages secrete cytokines (e.g., TNF) and chemokines (e.g., IL-8), attracting neutrophils to alveolar spaces • Activated neutrophils release proteases (e.g., elastase and proteinase 3), damaging alveolar walls • Oxidative damage leads to inactivation of α-1-antitrypsin ○ Antiprotease secreted by neutrophils • Genetic deficiency of α-1-antitrypsin (homozygous PiZZ) presents in ~ 1% of emphysema patients • Matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs also imbalanced and contribute to tissue damage • Increased levels of proteases and decreased levels of antiprotease result in overall tissue destruction

Pathophysiology • Destruction of alveolar elastic tissue leads to ○ Decreased radial traction ○ Collapse of respiratory bronchioles during expiration (functional airflow obstruction)

CLINICAL ISSUES

IMAGING CT Findings • Centriacinar emphysema is upper-lobe predominant • Panacinar emphysema is predominant in lung bases

MACROSCOPIC General Features • Enlarged hyperinflated lung weighing less than normal • Apical blebs or bullae • Centriacinar emphysema: Visibly enlarged blackened airspaces at centers of lobules • Panacinar emphysema has enlarged airspaces throughout entire lobule

MICROSCOPIC Histologic Features • • • • • • • • •

Enlarged airspaces Free-floating alveolar septa Nodular tips at ends of alveolar septa Bronchial metaplasia Peribronchiolar pigmented "smoker's" macrophages Subpleural fibrosis may be present Patchy nonspecific interstitial fibrosis may be seen No significant inflammation Secondary pulmonary hypertensive changes

DIFFERENTIAL DIAGNOSIS

Presentation

Overinflation Artifact From Formalin Perfusion

• • • •

• Enlarged alveolar sacs but no free-floating septa (no alveolar damage)

Dyspnea on exertion occurs early Productive cough Dyspnea at rest and aversion to exertion occur late Acute exacerbation with wheezing, cough, dyspnea, and possibly fever • Evidence of airflow obstruction by pulmonary function testing • α-1-antitrypsin deficiency usually young to middle-aged adult patients ○ May have chronic liver disease and panniculitis • Hyperinflation and emphysema may occur in adult survivors of neonatal bronchopulmonary dysplasia

Treatment • Surgical approaches ○ Severe chronic disease or those hospitalized with severe acute exacerbation considered for lung transplantation

Lung Transplantation

• Chronic obstructive pulmonary disease (COPD)

○ Advantages of lung volume reduction surgery (LVRS) reported • Drugs ○ Bronchodilators and inhaled glucocorticoids

TERMINOLOGY

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Enlarged airspaces • Free-floating alveolar septa • Nodular tips at ends of alveolar septa

SELECTED REFERENCES 1.

2.

Hogg JC et al: The contribution of small airway obstruction to the pathogenesis of chronic obstructive pulmonary disease. Physiol Rev. 97(2):529-552, 2017 Berg K et al: The pathology of chronic obstructive pulmonary disease: progress in the 20th and 21st centuries. Arch Pathol Lab Med. 140(12):14231428, 2016

369

Lung Transplantation

Cystic Fibrosis KEY FACTS

TERMINOLOGY

MACROSCOPIC

• Multisystem disorder due to any of > 2,000 autosomal recessive mutations in CFTR gene

• Thick, purulent mucus in dilated airways

ETIOLOGY/PATHOGENESIS

• Dilated bronchi up to near pleural surface • Dense acute and chronic airway inflammation • Airway mucus usually contains neutrophils and inflammatory debris • Damage to airway epithelium leading to squamous metaplasia, ulceration, and necrosis with abscesses • Emphysematous change, airway fibrosis, and bronchiolitis obliterans in late stage

• Most common mutation is ΔF508 (~ 70% of patients) • Genetic mutations result in abnormal epithelial chloride channel • Viscid mucus impairs mucociliary action • Recurrent infections lead to airway inflammation, necrosis, fibrosis, and eventual irreversible dilatation (bronchiectasis)

CLINICAL ISSUES • Chronic productive cough and wheezing from obstruction • Persistent colonization and infection with atypical organisms • Overall survival in CF improved to mean of 50 years with recent advances in therapy, including new drugs that restore CFTR function

MICROSCOPIC

TOP DIFFERENTIAL DIAGNOSES • Primary ciliary dyskinesia (Kartagener syndrome)

Cystic Fibrosis

Bronchiectasis in Cystic Fibrosis

Bronchiectasis in Cystic Fibrosis

Bronchiolitis Obliterans in Cystic Fibrosis

(Left) Gross photograph of explanted lung shows thickened pleural surface on the left and parenchymal cut surface on the right. Bronchiectatic airways are filled with a large amount of greenish-yellow mucus. (Right) Low-power view of cystic fibrosis shows a dilated, inflamed bronchus. Compare to the adjacent pulmonary artery branch, which has irregular intimal fibrosis (pulmonary arterial hypertension).

(Left) High-power view of cystic fibrosis airway shows acute inflammation in the lumen ﬈, intact mucosa ﬊, and marked submucosal chronic inflammation ﬉ with peribronchial fibrosis. (Right) In older patients with cystic fibrosis, there is fibrous obliteration of bronchiole ﬈ (note residual smooth muscle ﬉) and emphysematous changes ﬊.

370

Cystic Fibrosis

IMAGING

Abbreviations

CT Findings

• Cystic fibrosis (CF)

• Hyperinflation, infiltrates, atelectasis, bronchiectasis

Synonyms • Mucoviscidosis

Definitions • Multisystem disorder due to autosomal recessive mutation in cystic fibrosis transmembrane conductance regulator (CFTR) gene

ETIOLOGY/PATHOGENESIS > 2,000 Known Gene Mutations • Wide variety of clinical expression in which non-CFTR factors play important role • Results in abnormal epithelial chloride channel and abnormal regulation of other ion transporters and cellular processes • Most common mutation is ΔF508 (~ 70% of patients) • Defective ion transport leads to dehydrated and viscid airway mucus ○ Viscid mucus impairs mucociliary action • Secretions build up and become infected • Recurrent infections lead to airway inflammation, necrosis, fibrosis, and eventual irreversible dilatation (bronchiectasis)

CLINICAL ISSUES Presentation • Highly variable due to different genotype-phenotype associations • Pulmonary symptoms ○ Chronic productive cough and wheezing from obstruction ○ Persistent colonization and infection with atypical organisms • Other presentations include meconium ileus and intestinal obstruction, pancreatic insufficiency and recurrent pancreatitis, biliary cirrhosis, malnutrition, obstructive azoospermia

Treatment • Bilateral lung transplantation for oxygen-dependent respiratory failure, hypercapnia, and pulmonary hypertension • Single lung transplants not performed due to infectious complications from native lung

MACROSCOPIC General Features • • • •

Airway dilatation extending into peripheral lung Thick mucus in airways Parenchymal abscess from infection Thickened pleura

Lung Transplantation

TERMINOLOGY

MICROSCOPIC Histologic Features • Dilated bronchi present near pleural surface • Dense acute and chronic airway submucosal inflammation • Airway mucus usually contains neutrophils and inflammatory debris • Bronchial gland and goblet cell hyperplasia • Damage to airway epithelium leading to squamous metaplasia, desquamation, ulceration, and necrosis with abscesses • Emphysematous change, airway fibrosis, and bronchiolitis obliterans develop in late stage • Secondary pulmonary arterial hypertension

DIFFERENTIAL DIAGNOSIS Bronchiectasis • Primary ciliary dyskinesia (Kartagener syndrome) • Postinfectious bronchiectasis usually reversible and localized • Intralobar sequestration, tumor, or foreign body-associated bronchiectasis is localized

Immunodeficiency Disorders • Recurrent infections

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Dilated, mucus-filled airways extending to pleural surface • Dense acute and chronic airway submucosal inflammation • Bronchiolitis obliterans

SELECTED REFERENCES 1.

Prognosis

2.

• Age at transplant increased > 10 years due to improved management • Longest median survival (6.4 years) after lung transplant of all pretransplant diagnoses • Overall survival in CF improved to mean of 50 years with recent advances in therapy, including new drugs that restore CFTR function • Infection and chronic rejection most significant posttransplant complications

3. 4. 5. 6. 7. 8.

Castellani C et al: Cystic fibrosis: a clinical view. Cell Mol Life Sci. 74(1):129140, 2017 Cribbs SK et al: Microbiome in the pathogenesis of cystic fibrosis and lung transplant-related disease. Transl Res. 179:84-96, 2017 Chaparro C et al: Lung transplantation for cystic fibrosis: an update. Expert Rev Respir Med. 10(12):1269-1280, 2016 Morrell MR et al: Lung transplantation for cystic fibrosis. Clin Chest Med. 37(1):127-38, 2016 Lobo LJ et al: Respiratory infections in patients with cystic fibrosis undergoing lung transplantation. Lancet Respir Med. 2(1):73-82, 2014 Cohen TS et al: Cystic fibrosis: a mucosal immunodeficiency syndrome. Nat Med. 18(4):509-19, 2012 Lobo J et al: Recent advances in cystic fibrosis. Clin Chest Med. 33(2):307-28, 2012 Lubamba B et al: Cystic fibrosis: Insight into CFTR pathophysiology and pharmacotherapy. Clin Biochem. Epub ahead of print, 2012

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Lung Transplantation

Idiopathic Pulmonary Fibrosis KEY FACTS

ETIOLOGY/PATHOGENESIS • Genetic predisposition of aging lung to peripheral tractional injury

CLINICAL ISSUES • Pirfenidone and Nintedanib (antifibrotic drugs) decrease rate of progression of idiopathic pulmonary fibrosis • Lung transplantation currently only definitive therapy • Median survival: 3 years after diagnosis (if not transplanted) ○ 4.5 years after transplant

IMAGING • Lower lobe-predominant patchy subpleural distribution of fibrosis • Bibasilar reticular opacities, ground-glass opacifications, honeycombing, and traction bronchiectasis

MICROSCOPIC • Geographic heterogeneity: Patchy lesional involvement

○ Lower lobe-predominant fibrosis that progresses to upper lobes ○ Fibrosis begins in subpleural and paraseptal areas and progresses toward center of lobe • Temporal heterogeneity: Dynamic process of constantly evolving fibroblastic foci laying down new collagen matrix that eventually becomes fibrotic ○ Fibroblastic foci at interface of fibrotic areas and less involved lung parenchyma ○ Established fibrosis with microscopic honeycombing • Absence of hyaline membranes, organizing pneumonia, granulomas, interstitial inflammatory infiltrate, or predominance of airway-centered changes

TOP DIFFERENTIAL DIAGNOSES • Chronic hypersensitivity pneumonitis • Connective tissue disease-associated interstitial lung disease

Usual Interstitial Pneumonia

Fibroblastic Focus in Usual Interstitial Pneumonia

Honeycombing in Usual Interstitial Pneumonia

Pulmonary Hypertension in Usual Interstitial Pneumonia

(Left) Low-power view shows the characteristic variation in intensity and time, i.e., more ﬉ and less ﬊ involved areas and older ſt and younger fibrosis (fibroblastic foci) ﬈, respectively. (Right) Highpower image of a fibroblastic focus ſt shows parallel arrangement of fibroblasts under the bronchiolar epithelium. Note abundant bluish matrix and lack of inflammation within the focus.

(Left) Within areas of honeycombing, there are collections of amorphous debris and mucus ﬊, which predispose patients with idiopathic pulmonary fibrosis (IPF) to develop superimposed infection. (Right) Secondary pulmonary arterial hypertension is often present in IPF/usual interstitial pneumonia, as seen in these arterioles with medial hypertrophy and adventitial collagen deposition ﬈.

372

Idiopathic Pulmonary Fibrosis

MACROSCOPIC

Abbreviations

General Features

• Idiopathic pulmonary fibrosis (IPF)

• Correlate well with HRCT findings • Subpleural interstitial fibrosis and honeycombing

Synonyms • Usual interstitial pneumonia (UIP) denotes array of histopathologic features in IPF • Cryptogenic fibrosing alveolitis

Definitions • Chronic progressive bilateral interstitial lung disease characterized by geographic and temporal heterogeneity with fibroblastic foci and microscopic honeycombing

ETIOLOGY/PATHOGENESIS Idiopathic • Etiology unknown, but pathogenesis likely involves environmental factors in patients with genetic predisposition • Majority of patients are smokers; also increased risk post exposure to variety of environmental substances

Tractional Injury • Genetic predisposition of aging lung to peripheral tractional injury ○ Shortened survival of type II pneumocytes ○ Prolonged survival of activated myofibroblasts • Recurrent damage to epithelial-mesenchymal interface resulting in fibroblastic foci • Fibrosis extends from periphery toward center of lung

Inflammation • Secondary contributor to pathogenesis ○ Abnormal wound healing involving IL-13 and M2 macrophage responses

CLINICAL ISSUES Presentation • Usually > 50 years of age • Dyspnea on exertion and nonproductive cough • Restrictive pattern on pulmonary function testing

Treatment • Pirfenidone and Nintedanib (antifibrotic drugs) decrease rate of progression of IPF • Lung transplantation currently only definitive therapy

Prognosis • Median survival: 3 years after diagnosis (if not transplanted) or 4.5 years after transplant • Survival is similar between bilateral and single lung transplants

IMAGING High-Resolution CT (HRCT) • Lower lobe-predominant patchy subpleural fibrosis • Bibasilar reticular opacities, ground-glass opacifications, honeycombing, and traction bronchiectasis • Above features considered highly specific such that lung biopsy is not essential for IPF diagnosis

MICROSCOPIC Histologic Features

Lung Transplantation

TERMINOLOGY

• Geographic heterogeneity: Patchy lesional involvement ○ Lower lobe-predominant fibrosis progressing to upper lobes ○ Fibrosis begins in subpleural and paraseptal areas and progresses toward center of lobe ○ Areas of preserved lung parenchyma • Temporal heterogeneity: Dynamic process of constantly evolving fibroblastic foci laying down new collagen matrix that eventually becomes fibrotic ○ Fibroblastic foci at interface of fibrotic areas and less involved lung parenchyma – Interstitial fibroblasts with gray/blue stroma streaming parallel to alveolar or bronchiolar wall just beneath epithelial lining ○ Established fibrosis with microscopic honeycombing – Loss of normal alveolar spaces – Replacement by cystic spaces lined by bronchiolar metaplasia and filled with mucus (mucostasis) • Absence of hyaline membranes, organizing pneumonia, granulomas, interstitial inflammatory infiltrate, or predominance of airway-centered changes (features suggesting alternate pathology) • Additional findings ○ Smooth muscle hyperplasia ○ Mild to moderate chronic inflammation ○ Secondary pulmonary hypertensive changes • Acute exacerbation ○ Alveolar damage with hyaline membranes ○ Organizing pneumonia ○ Features may mask underlying UIP

DIFFERENTIAL DIAGNOSIS Chronic Hypersensitivity Pneumonitis • Perilobular fibrosis, but presence of centrilobular and bridging fibrosis along with granulomas and localized organizing pneumonia is more specific for hypersensitivity pneumonitis

Connective Tissue Disease-Associated Interstitial Lung Disease • Usually mixed pattern of UIP and nonspecific interstitial pneumonia, often with germinal centers

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Patchy fibrosis with fibroblastic foci and microscopic honeycombing

SELECTED REFERENCES 1.

Noble PW et al: Pirfenidone for idiopathic pulmonary fibrosis: analysis of pooled data from three multinational phase 3 trials. Eur Respir J. 47(1):24353, 2016

373

Lung Transplantation

Connective Tissue Disease-Associated Lung Disease KEY FACTS

• ~ 40% of systemic sclerosis patients have interstitial lung disease (ILD) • ~ 10% of patients with systemic sclerosis (SSc) develop PAH (less common in other CTDs) • Lung transplantation for eligible patients with end-stage lung disease refractory to medical therapy

– Both within and away from areas of fibrosis ○ Fibroblastic foci present – Less frequent and smaller than in idiopathic ILD • CTD-pulmonary arterial hypertension (PAH) ○ Pulmonary hypertensive changes in absence of interstitial fibrosis • Other CTD-associated lung disease ○ Chronic aspiration ○ Alveolar hemorrhage • Look for viral or fungal infection ○ Patients immunosuppressed due to CTD therapy

MICROSCOPIC

TOP DIFFERENTIAL DIAGNOSES

• CTD-ILD ○ Often features of mixed UIP and nonspecific interstitial pneumonia (NSIP) patterns – Either may be seen alone, especially NSIP ○ May have intense lymphocytic and plasmacytic inflammation with germinal centers

• Idiopathic pulmonary fibrosis ○ UIP pattern unassociated with underlying CTD • PAH ○ Idiopathic PAH unassociated with underlying CTD

TERMINOLOGY • Parenchymal and vascular lung disease in connective tissue disease (CTD) patients

CLINICAL ISSUES

Nonspecific Interstitial Pneumonia

Usual Interstitial Pneumonia

Lymphoplasmacytic Inflammation

Pulmonary Arterial Hypertension

(Left) Many sections from this explanted lung show nonspecific interstitial pneumonia pattern. There is relatively uniform thickening of alveolar walls with mild fibrosis and chronic inflammation. (Right) Other sections from the same explanted lung have more extensive fibrosis with distortion of lung architecture and fibroblastic foci ﬈ (usual interstitial pneumonia pattern).

(Left) This wedge biopsy from a patient with systemic lupus erythematosus-associated lung disease shows chronic interstitial inflammation (lymphocytes and plasma cells) and lymphoid aggregates. (Right) Concentric intimal fibrosis and medial hypertrophy of pulmonary artery branches is present in this scleroderma-associated lung disease.

374

Connective Tissue Disease-Associated Lung Disease

MICROSCOPIC

Abbreviations

Histologic Features

• Connective tissue disease (CTD)

• CTD-ILD ○ Often features of mixed UIP and NSIP patterns – Either may be seen alone, especially NSIP ○ Fibroblastic foci less frequent and smaller than idiopathic ILD ○ May have intense lymphocytic and plasmacytic inflammation with germinal centers – Both within and away from areas of fibrosis ○ Pleural fibrosis and inflammation ○ Secondary pulmonary hypertensive changes often present – Medial and intimal hypertrophy – Mucinous degeneration of media • CTD-PAH ○ Pulmonary hypertensive changes in absence of interstitial fibrosis – Medial hypertrophy – Concentric intimal fibrosis – Plexiform lesions present but less frequent than idiopathic PAH • Other CTD-associated lung disease ○ Would be seen in addition to ILD or PAH ○ Chronic microaspiration – Foreign body giant cell granulomas containing aspirated material – Intraalveolar clusters of foamy macrophages ○ Alveolar hemorrhage – Hemosiderin-laden macrophages and red blood cells in alveolar spaces ○ Adenocarcinoma, especially in SSc ○ Pulmonary thromboemboli and infarction, especially systemic lupus erythematosus ○ Follicular bronchiolitis, especially in rheumatoid arthritis • Look for viral or fungal infection as patients are immunosuppressed due to CTD therapy

Synonyms • CTD-associated interstitial lung disease (CTD-ILD) • CTD-associated pulmonary arterial hypertension (CTD-PAH)

Definitions • Parenchymal and vascular lung disease occurring in CTD patients

ETIOLOGY/PATHOGENESIS Pathobiology of CTD-ILD • Aberrant homeostasis of circulatory and immune systems ○ Fibrosis with microvascular damage ○ Upregulated inflammatory response, including B lymphocytes

Pathobiology of CTD-PAH • Endothelial injury, angiogenesis, and autoimmunity play role

CLINICAL ISSUES Epidemiology • Incidence ○ ~ 40% of systemic sclerosis (SSc) patients have ILD ○ ~ 10% of SSc patients develop PAH (less common in other CTDs) ○ Most prevalent CTD-ILD is nonspecific interstitial pneumonia (NSIP) followed by usual interstitial pneumonia (UIP) ○ UIP pattern most common in rheumatoid arthritis

Presentation • Dyspnea on exertion and dry cough

Treatment • Surgical approaches ○ Lung transplantation for eligible patients with end-stage lung disease refractory to medical therapy • Drugs ○ Antiinflammatory agents ○ Antifibrotic agents

Prognosis • • • •

CTD-ILD has better prognosis than idiopathic ILD CTD-PAH has worse prognosis than idiopathic PAH ILD major cause of death in SSc patients Survival post transplant similar to patients with transplants due to idiopathic pulmonary fibrosis or idiopathic PAH

IMAGING CT Findings • High-resolution CT shows pulmonary fibrosis in reticulonodular pattern and ground-glass opacities

Lung Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Idiopathic Pulmonary Fibrosis • UIP pattern unassociated with underlying CTD • Inflammation confined to areas of fibrosis

Pulmonary Arterial Hypertension • Idiopathic PAH unassociated with underlying CTD

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • CTD-ILD has mixed UIP-NSIP pattern with germinal centers and subtle fibroblastic foci • CTD-PAH has similar histology as idiopathic PAH

SELECTED REFERENCES 1. 2. 3.

Rosenbaum JT et al: The microbiome: a revolution in treatment for rheumatic diseases? Curr Rheumatol Rep. 18(10):62, 2016 Vandecasteele EH et al: The heart and pulmonary arterial hypertension in systemic sclerosis. Acta Clin Belg. 71(1):1-18, 2016 Wallace B et al: Management of connective tissue diseases associated interstitial lung disease: a review of the published literature. Curr Opin Rheumatol. 28(3):236-45, 2016

375

Lung Transplantation

Sarcoidosis, Lung KEY FACTS

CLINICAL ISSUES • More common in African Americans • Remission often occurs with medical therapy • Asymptomatic recurrence in majority of transplanted lungs ○ Does not affect survival • Median survival is 6 years post transplant

• Granulomatous vasculitis involving adventitia and media without vascular necrosis • Schaumann or asteroid bodies may be present within giant cells but are not sensitive or specific for sarcoidosis

ANCILLARY TESTS • Negative acid-fast bacillus and silver stains

IMAGING

TOP DIFFERENTIAL DIAGNOSES

• Parenchymal changes including nodular opacities in perilymphatic distribution • Hilar lymphadenopathy

• • • •

MICROSCOPIC • Multiple submucosal and interstitial granulomas in bronchovascular and lymphatic distribution • Well-circumscribed, nonnecrotizing granulomas containing epithelioid histiocytes, multinucleated giant cells (often), and rim of lymphocytes

Mycobacterial or fungal infection Chronic hypersensitivity pneumonitis Granulomatosis with polyangiitis (Wegener) Chronic berylliosis

DIAGNOSTIC CHECKLIST • Granulomatous vasculitis and interstitial fibrosis lead to pulmonary hypertension • Well-formed nonnecrotizing submucosal granulomas along lymphovascular distribution, capably assessed by transbronchial biopsy

Pulmonary Sarcoidosis

Posttherapy Sarcoidosis

Pulmonary Sarcoidosis

Sarcoidosis in Lymph Node

(Left) Submucosal granuloma is composed of epithelioid histiocytes, multinucleated giant cells, and a few lymphocytes. Note small focus of bland necrosis ﬈, which can be occasionally present in sarcoidosis. (Right) Posttreatment sarcoid, as is often seen in explanted lungs, may show only residual giant cells as illustrated here. Note various inclusions in the giant cells, including a Schaumann body ﬉.

(Left) There is progressive fibrosis in pulmonary sarcoidosis st as seen in this high-power image. An asteroid body ﬈ is present within a multinucleated giant cell, which is characteristic but not diagnostic for sarcoidosis. (Right) Almost all patients undergoing lung transplantation for sarcoidosis have extensive involvement of hilar lymph nodes as seen here at low power. There is almost complete replacement of nodal architecture by granulomas.

376

Sarcoidosis, Lung

Definitions • Multiorgan granulomatous disease of unknown etiology frequently affecting lungs

ETIOLOGY/PATHOGENESIS Environmental Exposure • Slight increased risk with exposure to mold, mildew, pesticides, and agricultural employment • Decreased risk with exposure to tobacco and allergy to dust and feathers

Pathophysiology • Specific pathogenesis unknown; diagnosis of exclusion • Exposure of genetically susceptible host to unknown antigen leads to T-helper 1 (Th1) response with cytokine release and granuloma formation

CLINICAL ISSUES Epidemiology • More common in African Americans

Presentation • Asymptomatic (incidental findings on chest imaging or pulmonary function testing) in ~ 50% • Dyspnea or cough • Decreased diffusion capacity and restrictive or obstructive features on pulmonary function testing

Treatment • Drugs ○ Corticosteroids are 1st-line therapy for symptomatic disease ○ Methotrexate and infliximab (anti-TNF monoclonal antibody) for refractory disease • Lung transplantation ○ For progressive refractory disease with fibrosis (minority of patients) and pulmonary hypertension

Prognosis • Remission often occurs with medical therapy • Increased mortality with onset of pulmonary hypertension (indication for transplantation) • Asymptomatic recurrence in majority of transplanted lungs but does not affect survival • Median survival is 6 years post transplant

IMAGING Radiographic Findings • Scadding chest radiographic staging system ○ Hilar lymphadenopathy ○ Parenchymal changes including nodular opacities in perilymphatic distribution ○ Pulmonary fibrosis ± traction bronchiectasis

MACROSCOPIC

• Diffuse fibrosis and honeycombing (late stage)

MICROSCOPIC Histologic Features • Multiple submucosal and interstitial granulomas ○ Bronchovascular and lymphatic distribution (replaced diffusely by fibrosis and honeycombing in late stage) • Well-circumscribed granulomas containing epithelioid histiocytes, multinucleated giant cells (often), and rim of lymphocytes ○ Classic nonnecrotizing granulomas – Microscopic foci of central necrosis can be seen • Schaumann or asteroid bodies may be present in giant cells ○ Not sensitive or specific for sarcoidosis • Granulomatous vasculitis involving adventitia and media ○ Without vascular necrosis ○ Progressive pulmonary hypertensive change – Due to vasculitis and interstitial fibrosis • Extensive involvement of hilar lymph nodes • Necrotizing sarcoidosis (distinct subset) ○ Confluent granulomas with large areas of nonsuppurative necrosis; adjacent areas of more classic nonnecrotizing granulomas

ANCILLARY TESTS Histochemistry • Negative acid-fast bacillus (AFB) and silver stains ○ Perform on multiple blocks if necrosis present

DIFFERENTIAL DIAGNOSIS Mycobacterial or Fungal Infection • Mycobacteria: Classic necrotizing granulomas ○ Positive AFB stain or tissue PCR • Histoplasmosis: Positive silver stain or serology

Chronic Hypersensitivity Pneumonitis • Poorly formed bronchiolocentric and interstitial granulomas with interstitial fibrosis • No lymph node involvement

Granulomatosis With Polyangiitis (Wegener) • Geographic necrotizing granulomas associated with positive c-ANCA serology

Chronic Berylliosis • Similar histology, requires clinical correlation

Drug Reaction or Intravenous Drug Abuse • Eosinophils or foreign body granulomas, requires clinical correlation

Common Variable Immunodeficiency • Granulomas often with marked lymphocytic infiltrate, hypogammaglobulinemia, and recurrent infections

SELECTED REFERENCES 1.

General Features • Yellow nodules along interlobular septa and around bronchovascular bundles

Lung Transplantation

TERMINOLOGY

2.

Sawahata M et al: An epidemiological perspective of the pathology and etiology of sarcoidosis. Sarcoidosis Vasc Diffuse Lung Dis. 33(2):112-116, 2016 Rossi G et al: Pathology of sarcoidosis. Clin Rev Allergy Immunol. 49(1):36-44, 2015

377

Lung Transplantation

Pulmonary Arterial Hypertension KEY FACTS

TERMINOLOGY

IMAGING

• Clinical classification of pulmonary arterial hypertension [group 1 of pulmonary hypertension (PH)] includes diverse group of diseases affecting pulmonary arteries • May be idiopathic, heritable, drug or toxin induced, or associated with congenital heart disease or connective tissue disease (CTD)

• Echocardiography to measure pulmonary artery pressures and right ventricular size

MICROSCOPIC

• Accumulation of vascular endothelial and smooth muscle cells due to imbalance of apoptosis and proliferation • Vasoconstriction, vessel wall remodeling, and thrombosis contribute to increased pulmonary vascular resistance

• Pulmonary arteries may have intimal thickening with concentric intimal fibrosis, plexiform lesions, medial hypertrophy, or perivascular (adventitial) collagen deposition • Pulmonary arterioles may become muscularized with marked perivascular lymphocytic infiltrate • Normal alveolar parenchyma (or features of CTD-associated interstitial lung disease)

CLINICAL ISSUES

TOP DIFFERENTIAL DIAGNOSES

• Average life expectancy without transplant is 3 years • Posttransplant survival rates: 5 year = 52-75%, 10 year = 4566%

• Secondary PH • Thromboembolic disease

ETIOLOGY/PATHOGENESIS

Medial Hypertrophy

Plexiform Lesion

Plexiform Lesion

Plexiform Lesion (Elastin)

(Left) This parenchymal arteriole shows marked hypertensive changes due to intimal and medial proliferation. There is neartotal luminal constriction. Note the congested capillaries in the normal parenchyma. (Right) Intraarterial endothelial proliferation creates multiple lumina as seen in this plexiform lesion ﬉. Note the typical location at branch point of the pulmonary artery ſt and adjacent dilated vessels.

(Left) This high-power photomicrograph of a plexiform lesion is from the lung of a 29-year-old female patient who died of severe pulmonary hypertension. There are multiple fibrin thrombi ſt within the endothelium-lined vascular spaces of the lesion. (Right) Elastic stain, especially helpful in highlighting the vasculature, shows a plexiform lesion and adjacent dilated vessels.

378

Pulmonary Arterial Hypertension

Abbreviations • Pulmonary arterial hypertension (PAH) • Pulmonary hypertension (PH)

Definitions • Pulmonary vascular disease originating in pulmonary arterial circulation • Mean pulmonary artery pressure ≥ 25 mm Hg at rest

Classification • Current Dana Point 2008 clinical classification: 5 major types of PH, of which PAH is group 1 ○ Idiopathic – Majority of PAH (group 1) cases ○ Heritable or familial – BMPR2, ALK1, END, SMAD9, CAV1 mutations ○ Drug or toxin induced – Anorexigens, methamphetamines, various chemotherapeutic agents ○ Associated conditions – Congenital heart disease – Connective tissue disease (CTD), most commonly systemic sclerosis – Human immunodeficiency virus (HIV) – Portal hypertension ○ Persistent PH of newborn – Meconium aspiration, infection ○ Pulmonary venoocclusive disease &/or pulmonary capillary hemangiomatosis (PVOD/PCH) – Idiopathic, drug/toxin induced, or associated with CTD, HIV infection, or EIF2AK4 mutation • Pathologic features do not correlate with clinical classification

ETIOLOGY/PATHOGENESIS Pathophysiology • Accumulation of vascular endothelial and smooth muscle cells due to loss of apoptosis • Vasoconstriction, vessel wall remodeling, and thrombosis contribute to increased pulmonary vascular resistance • Right ventricle responds by increasing systolic pressure to maintain cardiac output • Increased demand on right ventricle eventually leads to ischemia and right heart failure

CLINICAL ISSUES

• Patients with severe right ventricular dysfunction or ventricular septal defects may receive heart-lung transplant

Prognosis • Average life expectancy without transplant is 3 years • PVOD/PCH has worst prognosis, followed by CTDassociated PAH, idiopathic PAH, and congenital heart disease-associated PAH (best prognosis) • Posttransplant survival rates: 5 year = 52-75%, 10 year = 4566%

IMAGING Ultrasonographic Findings • Echocardiography to measure pulmonary artery pressures and right ventricular size

MACROSCOPIC General Features • Atherosclerotic plaques in larger pulmonary arteries • Right ventricular hypertrophy in combined heart-lung explant

MICROSCOPIC Histologic Features • Pulmonary arteries may have ○ (1) Medial smooth muscle hypertrophy, (2) intimal ("onion skin") fibrosis ± adventitial thickening ○ Plexiform lesions may be seen in idiopathic, heritable, drug-induced, HIV-associated, and some CTD-associated PAH – Formed by intraarterial endothelial and stromal proliferation creating multiple vascular channels • May have (1) muscularized pulmonary arterioles, (2) marked perivascular lymphocytic infiltrate • Normal alveolar parenchyma (or features of CTD-associated interstitial lung disease) • PVOD/PCH: Fibrous obliteration of pulmonary veins surrounded by congested alveolar capillaries

DIFFERENTIAL DIAGNOSIS Secondary Pulmonary Hypertension • Parenchymal etiology, such as interstitial lung disease

Thromboembolic Disease • Recanalized vessels can mimic plexiform lesions

DIAGNOSTIC CHECKLIST

Presentation

Clinically Relevant Pathologic Features

• Unexplained exertional dyspnea • Symptoms of right heart failure • May be asymptomatic

• Plexiform lesions are irreversible

Treatment • Transplantation offered to patients who failed conservative and medical therapy ○ Most receive bilateral lungs with subsequent gradual improvement in native cardiac function – Hemodynamic instability is frequent postoperative issue

Lung Transplantation

TERMINOLOGY

Pathologic Interpretation Pearls • Pulmonary arteries with concentric intimal fibrosis and medial hypertrophy • Plexiform lesions are pathognomonic for PAH

SELECTED REFERENCES 1. 2.

Nazzareno G et al: 2015 ESC/ERS Guidelines for the diagnosis and treatment of pulmonary hypertension. Eur Respir J. 46(6):903-975, 1855-6, 2015 Price LC et al: Inflammation in pulmonary arterial hypertension. Chest. 141(1):210-21, 2012

379

Lung Transplantation

Other Causes of End-Stage Lung Disease KEY FACTS

TERMINOLOGY • Entities described here include ○ Uncommon indications for lung transplantation ○ Pulmonary manifestations encountered in patients with other solid organ or hematopoietic stem cell transplantation (HSCT)

MICROSCOPIC • Surfactant dysfunction disorders and pulmonary alveolar proteinosis: PAS-positive, mucicarmine-negative eosinophilic granular material within alveolar spaces • Lymphangioleiomyomatosis: Thin-walled cysts with patchy nodular proliferation of bland smooth muscle cells • Langerhans cell histiocytosis: Nodular proliferation of Langerhans cells around cystically dilated small airways; progression to end-stage fibrosis (honeycombing) with few Langerhans cells • Iatrogenic

○ Amiodarone toxicity: Foamy alveolar macrophages, lipid vacuoles in pneumocytes and endothelial cells ○ Bleomycin: Lower lobe predominant diffuse alveolar damage or organizing pneumonia; late stage has interstitial fibrosis with UIP pattern ○ Methotrexate: Interstitial fibrosis with overlapping pattern of nonspecific interstitial pneumonia • HSCT: Chronic graft-vs.-host disease can manifest as bronchiolitis obliterans or, rarely, as unclassifiable interstitial pneumonia • Radiation pneumonitis: Acute: Diffuse alveolar damage and interstitial lymphocytes; chronic: Interstitial/alveolar fibrosis and intimal fibrosis with foamy macrophages

TOP DIFFERENTIAL DIAGNOSES • Emphysema, lipoid pneumonia

DIAGNOSTIC CHECKLIST • Clinical correlation is essential since many histologic features are nonspecific and often overlap

Surfactant Deficiency Disorder

Pulmonary Alveolar Proteinosis

Lymphangiomyomatosis

Bleomycin Toxicity

(Left) Lung wedge biopsy from a 1-month-old term baby shows intraalveolar eosinophilic material ﬈ and reactive type 2 pneumocytes ﬇, suggestive of surfactant dysfunction disorder. (Right) PAS stain of a lung section from a patient with longstanding pulmonary alveolar proteinosis shows characteristic PAS-positive intraalveolar granular material ﬈. Note minimal alveolar wall changes.

(Left) H&E in the lung of a 38year-old woman with lymphangioleiomyomatosis shows abnormal smooth muscle proliferation involving bronchiolar wall and alveolar septa. The muscle cells have plump, spindled nuclei. (Right) Explanted lung from a 25year-old with bleomycininduced pulmonary fibrosis, treated for rhabdomyosarcoma at age 11, shows extensive interstitial fibrosis and mild chronic inflammation.

380

Other Causes of End-Stage Lung Disease

Definitions • Uncommon etiologies of acute or chronic lung injury, which may lead to respiratory failure • Entities described include ○ Uncommon indications for lung transplantation ○ Pulmonary manifestations encountered in other solid organ or hematopoietic stem cell transplantation (HSCT)

ETIOLOGY/PATHOGENESIS Surfactant Dysfunction Disorders • Abnormal lung physiology due to congenital defects in surfactant proteins or processing/homeostasis

Pulmonary Alveolar Proteinosis • Autoimmune: Impaired surfactant protein clearance by macrophages ○ Neutralizing antibodies to granulocyte-macrophage colony stimulating factor (GM-CSF) • Secondary: Decreased number and function of macrophages ○ Associated with certain malignancies (e.g., hematologic) • Hereditary: Impaired macrophage function and decreased surfactant protein clearance ○ Mutation in GM-CSF receptor genes

Lymphangioleiomyomatosis • Loss of heterozygosity of chromosomes 9p and 16p • Tuberous sclerosis complex gene mutation

Langerhans Cell Histiocytosis • Reactive proliferation of Langerhans cells likely due to aberrant immune response to cigarette smoke antigens • BRAF V600E mutations in some may represent clonal proliferation

Bleomycin • Intrinsically low levels of metabolizing enzyme bleomycin hydrolase in lungs lead to high local levels of this cytotoxic drug

Amiodarone • Directly cytotoxic by accumulating in lysosomes and preventing turnover of endogenous phospholipids

Radiation Pneumonitis • Ionizing radiation has direct dose-dependent cytotoxic effect involving altered lipid metabolism • Risk factors: Older age, female gender, nonsmokers, preexisting lung disease, specific individual genetics

Chronic Graft-vs.-Host Disease in HSCT Patients Resulting in Bronchiolitis Obliterans • Engrafted hematopoietic cells attack host lung cells in alloimmune response involving innate immune system and genetic factors

CLINICAL ISSUES Epidemiology

• Lymphangioleiomyomatosis (LAM) accounts for 1.1% of adult lung transplants in International Society for Heart and Lung Transplantation registry ○ Adenocarcinoma accounts for 0.1% • Chemotherapy-related lung fibrosis is rare ○ Affects minority of exposed patients • Bronchiolitis obliterans (BO) occurs in ~ 6% of all HSCT but may be underrecognized ○ Chronic graft-vs.-host disease (GVHD) is predominant risk factor for BO in HSCT with prevalence of 14%

Lung Transplantation

TERMINOLOGY

Presentation • Generally dyspnea, cough, fever • Surfactant dysfunction disorders (SDD): Neonatal respiratory distress; some types present in later childhood/adolescence • LAM: Women of child-bearing age with dyspnea or pneumothorax; can associate with tuberous sclerosis • Langerhans cell histiocytosis (LCH): Adult smokers with chronic cough; may be asymptomatic • Chemotherapy-related lung fibrosis usually has insidious onset of symptoms • Radiation pneumonitis: Acute occurs within 6 months after therapy; chronic occurs after 6 months

Prognosis • Adenocarcinoma ○ Good prognosis for stage I if immunosuppression is reduced ○ Poor prognosis and rapid disease progression if diagnosed/explanted at advanced stage ○ Single lung transplant recipients with chronic obstructive pulmonary disease (COPD) or idiopathic pulmonary fibrosis – 2-4% incidence of developing lung carcinoma in native lung • BO in HSCT ○ 90% survival at 1 year and 75% at 5 years following lung transplantation • Little data on posttransplant prognosis available for other conditions discussed

IMAGING CT Findings • SDDs ○ Ground-glass opacities and peripheral cysts ○ Septal thickening in ABCA3 mutation but not in SFPTC mutation • PAP ○ Diffuse ground-glass opacities with septal thickening results in characteristic crazy-paving pattern • LAM ○ Bilateral, diffuse, thin-walled cystic lesions • LCH ○ Nodules and cysts involve all lobes in pediatric patients ○ Lower lobes spared in adult patients • Chronic GVHD in HSCT patients resulting in BO ○ Mosaic attenuation and bronchial thickening with pleural-based expiratory air-trapping

• Surfactant protein B deficiency is most common indication (16.7%) for lung transplantation in patients < 1 year old 381

Lung Transplantation

Other Causes of End-Stage Lung Disease

382

MICROSCOPIC Histologic Features • SDDs ○ PAS-positive, mucicarmine-negative eosinophilic granular material in alveolar spaces ○ Interstitial fibrosis with alveolar simplification and type II pneumocyte hyperplasia • PAP ○ PAS-positive, mucicarmine-negative eosinophilic granular material in alveolar spaces ○ Type II pneumocyte hyperplasia with preserved lung architecture (no fibrosis) ○ Alveolar spaces may have macrophages, sloughed pneumocytes, and cholesterol clefts • LAM ○ Thin-walled cysts (2-5 mm but sometimes larger) with patchy nodular proliferation of bland smooth muscle cells ○ Nodules occur around airways, lymphatics, and blood vessels ○ Lymphovascular proliferation: Slit-like (within muscular nodules) or dilated • LCH ○ Early cellular phase: Nodular proliferation of Langerhans cells around small airways – Airways can be cystically dilated – Eosinophils are common but not required for diagnosis – Langerhans cells typically have large nuclei with nuclear grooves ○ Late fibrotic phase: End-stage fibrosis (honeycombing) with few Langerhans cells • Adenocarcinoma ○ Ranges from well to poorly differentiated with multiple histologic types ○ Features of COPD or usual interstitial pneumonia (UIP) in sections away from carcinoma • Iatrogenic ○ Chemotherapy-related lung fibrosis – Bleomycin: Lower lobe predominant diffuse alveolar damage (DAD) or organizing pneumonia (OP) □ Late stage has interstitial fibrosis with UIP pattern – Cyclophosphamide: Interstitial fibrosis with UIP pattern or nonspecific interstitial pneumonia (NSIP) pattern; thickened and fibrotic pleura – Methotrexate: Interstitial fibrosis with overlapping pattern of NSIP, hypersensitivity pneumonitis (eosinophils, granulomas), OP, DAD ○ Amiodarone toxicity – Foamy alveolar macrophages, lipid vacuoles in pneumocytes and endothelial cells, OP, organizing DAD ○ Radiation pneumonitis – Acute: DAD, interstitial lymphocytes – Chronic: Interstitial and alveolar fibrosis; intimal fibrosis with foamy macrophages – Atypical pneumocytes and fibroblasts: Enlarged cells with nucleomegaly and hyperchromasia but preserved nuclear:cytoplasmic ratio ○ Immunosuppressants

– Sirolimus: Alveolar hemorrhage, pulmonary alveolar proteinosis (PAP), OP, BO, interstitial fibrosis – Tacrolimus: BO • HSCT: Chronic GVHD can manifest as BO or, rarely, as unclassifiable interstitial pneumonia

Cytologic Features • PAP: Can be diagnosed on Papanicolaou-stained bronchoalveolar lavage fluid (BAL) as orange globules with distinct green borders • Amiodarone toxicity: BAL shows foam cells, increased phospholipid content

ANCILLARY TESTS Immunohistochemistry • SDD: Confirm absence of immunohistochemical staining (i.e., deficiency) for surfactant B or C • PAP: Positive for surfactants A, B, and C by immunohistochemistry • LAM: Proliferative smooth muscle cells are positive for desmin, smooth muscle actin, HMB45 (focal/faint), estrogen receptor, and progesterone receptor • LCH: Langerhans cells are positive for S100, CD1a, and langerin

Electron Microscopy • SDD: Abnormal lamellar bodies associated with surfactant dysfunction

DIFFERENTIAL DIAGNOSIS Emphysema • Cystic airspace disease • May be radiographically similar to LAM and LCH

Lipoid Pneumonia • Alveolar foamy macrophage accumulation • Histologically similar to amiodarone toxicity

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Clinical correlation is essential since many histologic features are nonspecific and often overlap

SELECTED REFERENCES 1.

2.

3. 4.

Benden C et al: The Registry of the International Society for Heart and Lung Transplantation: sixteenth official pediatric lung and heart-lung transplantation report--2013; focus theme: age. J Heart Lung Transplant. 32(10):989-97, 2013 Bergeron A et al: Bronchiolitis obliterans syndrome after allogeneic hematopoietic SCT: phenotypes and prognosis. Bone Marrow Transplant. 48(6):819-24, 2013 Holm AM et al: Lung transplantation for bronchiolitis obliterans syndrome after allo-SCT. Bone Marrow Transplant. 48(5):703-7, 2013 Christie JD et al: The Registry of the International Society for Heart and Lung Transplantation: 29th adult lung and heart-lung transplant report-2012. J Heart Lung Transplant. 31(10):1073-86, 2012

Other Causes of End-Stage Lung Disease

Langerhans Cell Histiocytosis (Left) Langerhans cell histiocytosis in an explanted lung shows a nodular proliferation of Langerhans cells with occasional nuclear irregularities and grooves ſt. Accompanying eosinophils are abundant ﬈. (Right) Langerhans cell histiocytosis in an explanted lung has extensive fibrosis and only a few diagnostic Langerhans cells ﬇. Note that fibrosis is variable and there are reactive pneumocytes ſt.

Langerhans Cell Histiocytosis (Langerin)

Lung Transplantation

Langerhans Cell Histiocytosis

Langerhans Cell Histiocytosis (S100) (Left) Immunohistochemical stain with langerin shows strong cytoplasmic and membranous positivity in the lesional cells. Adjacent clusters of lymphocytes are negative ﬊. (Right) Immunohistochemistry for S100 in this case of Langerhans cell histiocytosis confirms the diagnosis. Note that there are only small areas of positive cells ﬊. Adjacent lymphocytes are negative ﬈. Note the uninvolved lung in the upper left part of image.

Radiation Atypia

Chronic Graft-vs.-Host Disease (Left) Chronic radiation pneumonitis shows extensive interstitial fibrosis and focal atypical pneumocytes with enlarged nuclei ﬈. Note that the fibroblasts are also large and atypical ﬊. (Right) Chronic graft-vs.-host reaction is seen in this explanted lung. The normal pulmonary artery branch ﬇ is accompanied by an airway that is completely obliterated ﬈ by fibrosis and chronic inflammation (bronchiolitis obliterans), recognized by residual muscle layer ſt.

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Lung Transplantation

Surgical Aspects and Complications, Lung KEY FACTS

TERMINOLOGY • Single lung transplantation: Unilateral excision of diseased lung with least pulmonary reserve and replacement with allograft lung • Bilateral sequential lung transplantation: Excision of both diseased lungs and replacement with allograft lungs with separate perihilar airway anastomoses • Heart-lung transplantation: En bloc excision of diseased heart and bilateral lungs and replacement with allograft heart-lung bloc • Lobar lung transplantation: Unilateral excision of diseased lung and replacement with bilateral right and left lower lobes from living donor

ETIOLOGY/PATHOGENESIS • Pleural space complications (hemothorax, pneumothorax, effusion, empyema) • Primary graft dysfunction (PGD) • Vascular insufficiency to anastomotic site

• • • •

Acute graft failure Anastomotic stenosis Thromboembolism Graft infection

CLINICAL ISSUES • Most common type in USA is single lung transplant ○ Followed by bilateral lung transplant • Surgical complications have steadily decreased since 1990s • PGD affects 10-25% of all lung transplant recipients and is leading cause of early death after transplantation

MICROSCOPIC • Diffuse alveolar damage in PGD and acute graft failure • Fibrinopurulent debris ○ ± bacterial colonies ○ ± pleural tissue in empyema

Pulmonary Venous Stenosis

Bronchial Anastomotic Stenosis

Acute Graft Failure

Bronchial Anastomotic Site

(Left) 3D reconstruction of a radiologic image after lung transplant shows stenosis of pulmonary venous anastomosis ſt. (Courtesy W. Vigneswaran, MD.) (Right) Gross photograph shows posterior view of a heart-lung specimen at autopsy, from a patient post left single lung transplant. Note bronchial anastomosis stenosis ﬈, for which a stent had been placed.

(Left) Diffuse alveolar damage is seen in this patient with acute graft failure 3 days after transplant. Note focal hyaline membranes ﬉, reactive pneumocytes ﬊, and interstitial edema ﬈. (Right) Necrotic cartilage ﬇ at the bronchial anastomotic site can be seen in vascular insufficiency or airway dehiscence and can precede stenosis.

384

Surgical Aspects and Complications, Lung

Definitions • Single lung transplantation: Unilateral excision of diseased lung with least pulmonary reserve and replacement with allograft lung • Bilateral sequential lung transplantation: Excision of both diseased lungs and replacement with allograft lungs with separate perihilar airway anastomoses • Heart-lung transplantation: En bloc excision of diseased heart and bilateral lungs and replacement with allograft heart-lung bloc • Lobar lung transplantation: Unilateral excision of diseased lung and replacement with bilateral right and left lower lobes from living donor

ETIOLOGY/PATHOGENESIS 24 Hours to 1 Week • Pleural space complications (hemothorax, pneumothorax, effusion, empyema) ○ In bilateral lung recipients, normal anatomic barrier separating right and left pleural space is lost – Allows extension of hemorrhage or effusion • Primary graft dysfunction (PGD) ○ Typically within 72 hours ○ Associated with ischemia-reperfusion injury, reactive oxygen species, and inflammation

8 Days to 2 Months • Vascular insufficiency to anastomotic site ○ Irregular blood flow due to vascular compromise ○ May be iatrogenic due to surgical procedure ○ May be associated with healing process, such as vascular compression from local edema • Acute graft failure ○ Unknown etiology after exclusion of preservation injury/PGD, hyperacute rejection/antibody-mediated rejection, acute respiratory distress syndrome, heart failure

Within 4 Months • Anastomotic stenosis ○ Excessive scarring at vascular or airway anastomotic site, leading to stricture and stenosis ○ May be preceded by necrosis • Thromboembolism ○ Pulmonary or other vascular embolism related to postoperative hypercoagulable state

At Any Time Post Transplant • Graft infection ○ Multiple posttransplant factors contribute to increased infection risk: Immunosuppression, impaired cough reflex, mucociliary dysfunction due to denervation of donor lung, altered lymphatic flow

CLINICAL ISSUES Epidemiology • Most common type in USA is single lung transplant ○ Followed by bilateral lung transplant • Heart-lung transplants performed infrequently in USA

• Surgical complications have markedly decreased over time • Pulmonary venous stenosis or complete occlusion more common than pulmonary artery stenosis (PAS) • PGD affects 10-25% of all lung transplant recipients

Treatment • Airway stenosis can be treated with endobronchial stent introduced via bronchoscopy • Airway dehiscence can be treated by bronchoscopic stent placement or by surgical intervention • Pulmonary venous stenosis/occlusion often requires lobar resection • PAS requires surgical intervention • Empyema may require surgical decortication

Lung Transplantation

TERMINOLOGY

Prognosis • Lack of allograft bronchial artery revascularization associated with airway hypoxia and chronic rejection • PGD is leading cause of early death ○ Impaired long-term graft function and increased risk of bronchiolitis obliterans syndrome • Without emergent repair, vascular anastomotic complications have high morbidity and mortality

IMAGING CT Findings • Pulmonary venous stenosis/occlusion is seen as marked infiltrates in single lung lobe • Peribronchial collections of air in airway dehiscence • Septal thickening, marked ground-glass opacities, and consolidation associated with PGD

MACROSCOPIC Anastomotic Stenosis • Narrowing at anastomotic site; ± stent; ± necrosis

Pleural Complications (Hemothorax) • Lung and bronchial surfaces with hemorrhage and clot

Thromboembolism • Vascular thrombus adherent to vessel wall; ± multiple

Vascular Insufficiency to Anastomotic Site • Soft yellow necrotic tissue at anastomotic site

Graft Infection • Bacterial: Diffuse or focal consolidation • CMV: Firm parenchyma with hemorrhagic nodules

MICROSCOPIC Empyema • Fibrinopurulent debris, ± bacterial colonies, ± pleural tissue

Primary Graft Dysfunction and Acute Graft Failure • Diffuse alveolar damage

SELECTED REFERENCES 1.

Krishnam MS et al: Postoperative complications of lung transplantation: radiologic findings along a time continuum. Radiographics. 27(4):957-74, 2007

385

Lung Transplantation

Pathologic Classification of Rejection

TERMINOLOGY Definitions • Alloimmune response: Recipient recognition of donor antigens • International Society for Heart and Lung Transplantation (ISHLT) published most recent guidelines for pathologic grading of acute rejection in 2007 ○ Grade should reflect most advanced pattern rather than most predominant ○ Perivascular and small airway inflammation can exist concurrently – Should be reported as such (i.e., grade A2 with B2R)

CLINICAL ISSUES Epidemiology • Incidence ○ Between 2004-2012, 33% of adult lung transplant recipients had ≥ 1 episode of acute rejection within 1 year ○ Acute rejection is less common in pediatric lung recipients, occurring in 4.5% of recipients within 1st year ○ Bronchiolitis obliterans syndrome (BOS) occurs in ~ 50% of lung transplant recipients within 5 years and in 75% within 10 years

ETIOLOGY/PATHOGENESIS Immunologic Basis of Lung Allograft Rejection • Recipient T cells recognize donor major histocompatibility complex (MHC) or human leukocyte antigen (HLA) peptides ○ HLA peptides presented initially by donor antigen presenting cells (APCs) and later by recipient APCs – Direct pathway: Donor APCs present donor MHCpeptide complexes to recipient T cells – Indirect pathway: Recipient APCs present donor peptides to recipient T cells ○ Recipient T cells recognize these antigens as foreign through T-cell receptor, and response is triggered – Donor allograft cells targeted by host immune system for destruction, resulting in graft injury – Both CD4(+) and CD8(+) T cells are found in perivascular cuff, diagnostic of acute cellular rejection (ACR) grade A on biopsy specimens • Local innate immunity is also activated, contributing to allograft injury ○ Mechanisms of innate immune system activation include – Tissue injury associated with transplantation procedure – Autoimmune response to self-antigens, which may become exposed during ischemia-reperfusion injury – Infection • HLA matching not routinely performed for lung transplantation • Lung thought to be primary site of T-cell activation ○ Acute rejection can occur in immediate postoperative period – Lymphatics not yet functioning to drain donor APCs to lymph nodes

386

– Lung lymphoid tissue, or bronchial-associated lymphoid tissue (BALT), functions similarly to lymph node as site of T-cell activation • In antibody-mediated rejection (AMR), alloantibodies deposit within graft vasculature, leading to complement activation, endothelial cell damage, and cell death • In BOS, immune response to airway epithelial antigens leads to Th17/IL-17 activation ○ Increased IL-17(+) CD8(+) T cells also found in inflammatory infiltrates in lymphocytic bronchiolitis (LB)

CLINICAL IMPLICATIONS Prognosis • Frequency and severity of acute rejection is one of most important risk factors for development of BOS • BOS is most common cause of death in all lung transplant recipients after 1 year post transplant

Clinical Presentation • Presenting signs and symptoms of ACR, when present, are nonspecific • Surveillance biopsies routinely performed to identify ACR and AMR ○ Histologic assessment is gold standard for diagnosis of rejection • BOS can occur anytime after transplant and presents with declining pulmonary function tests, dyspnea, and cough

ISHLT Guidelines and Ongoing Challenges • 1st published in 1990; revised in 1996 and 2007 ○ Some centers use 1996 system, although most use 2007 system • Major changes in 2007 from 1996 ○ Grade B – Grade B1R is combination of grade B1 and grade B2 from 1996 – Grade B2R is combination of grade B3 and grade B4 from 1996 classification – 1996 classification allowed for 5 grades of LB (B0-B4) or just stating presence or absence of LB, depending on transplant clinician's preference ○ Grade C – 2007 classification has no requirement to report inflammation associated with BO – Reported as grade C1a (active) or C1b (no inflammation) in 1996 classification • Because grade B1R (ISHLT 2007 classification) is broader category, including grades B1 and B2 (ISHLT 1996 classification), this may contribute to subsequent treatment differences for these patients • Clarification needed regarding utilization of terms grade AX and grade BX, as pathologists and pulmonologists may interpret differently ○ Neutrophilia can be present in higher A and B grades of ACR – Biopsies should not be underestimated as ungradable due to potential infection • Interobserver variability persists in grading of lung allograft biopsies

Pathologic Classification of Rejection

MACROSCOPIC FINDINGS Specimen Adequacy • ≥ 5 fragments of alveolated lung in transbronchial biopsy

Specimen Handling • Gently agitate in formalin to ensure expansion of alveoli • Avoid using forceps to minimize crush artifact

MICROSCOPIC FINDINGS Grading • ACR ○ Grade A: Perivascular inflammation – Grade A0: No acute rejection – Grade A1: Minimal acute rejection □ Infrequent circumferential cuffing of vessels by sparse mononuclear cell infiltrates – Grade A2: Mild acute rejection □ Frequent densely compacted or loose perivascular mononuclear cell infiltrates that may contain eosinophils or macrophages – Grade A3: Moderate acute rejection □ Extension of inflammatory infiltrate into subendothelium (endotheliitis), interstitium, and alveolar spaces □ May contain neutrophils – Grade A4: Severe acute rejection (relatively uncommon) □ Diffuse mononuclear perivascular, interstitial, and alveolar space infiltrates with pneumocyte damage and localized necrosis – Grade AX: Ungradable specimen □ Inadequate sample (< 5 fragments of alveolated lung), artifactual crush distortion, no arteriole/venule for evaluation ○ Grade B: Small airway inflammation (LB) – Grade B0: No airway inflammation – Grade B1R: Low-grade small airway inflammation □ Scattered or circumferential mononuclear cell infiltrate within bronchiolar submucosa with occasional eosinophils – Grade B2R: High-grade small airway inflammation □ Bronchial submucosal mononuclear cell infiltrate with accompanying eosinophils and even neutrophils, respiratory epithelial necrosis/ulceration with fibrinopurulent exudate – Grade BX: Ungradable specimen

□ Concurrent infection (suggested by disproportionate neutrophilic/eosinophilic infiltrate), tangential sectioning, crush artifact • Chronic airway rejection: Grade C (BO) ○ Eccentric or concentric hyaline fibrosis of bronchiolar submucosa, leading to luminal occlusion and fragmentation of airway architecture ○ BOS is clinical correlate ○ Requires wedge (not transbronchial) biopsy • Chronic vascular rejection: Grade D (accelerated graft vascular sclerosis) ○ Fibrointimal thickening and paucicellular hyaline sclerosis of arteries and veins ○ Requires wedge (not transbronchial) biopsy • AMR: Humoral rejection ○ Alloantibody injury to microvasculature and capillaries ○ No consensus on histologic criteria for AMR ○ Associated with poorer graft survival but overall rare in lung transplant recipients

Lung Transplantation

○ Moderate agreement reported for both grade A and grade B rejection, especially after revision of 1996 classification system • Many issues related to lung allograft biopsy interpretation remain unresolved ○ Degree of inflammation in grade B rejection ○ Large airway submucosal inflammation ○ Inflammation associated with BO ○ Specific definitions of grades AX and BX ○ Histologic features of AMR ○ Distinguishing signs of infection ○ Distinguishing features of BALT

Specimen Adequacy • Minimum of 3 levels of H&E-stained sections • Connective tissue stains (trichrome, elastin, etc.) may aid in identification of ○ Submucosal airway fibrosis in BO (grade C) ○ Arteriosclerosis in chronic vascular rejection (grade D) • Silver stains may help identify fungal elements • Grade AX &/or BX may be reported if specimen is not adequate for grading ○ ≤ 100 alveolar spaces total ○ ≤ 1 bronchiole ○ Presence of tangential cutting &/or processing artifact

Reporting • Nonrejection biopsy findings that should be reported ○ Infection – Presence of numerous neutrophils, viral inclusions, fungal hyphae, or yeast ○ Microaspiration – Exogenous lipoid pneumonia and multinucleated giant cells ○ Organizing pneumonia – May be subacute infection, ischemia-reperfusion injury, or resolving acute rejection

SELECTED REFERENCES 1. 2. 3.

4.

Husain AN et al: Lung transplantation: the state of the airways. Arch Pathol Lab Med. 140(3):241-4, 2016 Bhorade SM et al: Interobserver variability in grading transbronchial lung biopsy specimens after lung transplantation. Chest. 143(6):1717-24, 2013 Verleden SE et al: Involvement of interleukin-17 during lymphocytic bronchiolitis in lung transplant patients. J Heart Lung Transplant. 32(4):44753, 2013 Stewart S et al: Revision of the 1996 working formulation for the standardization of nomenclature in the diagnosis of lung rejection. J Heart Lung Transplant. 26(12):1229-42, 2007

387

Lung Transplantation

Antibody-Mediated Rejection, Lung KEY FACTS

TERMINOLOGY • Antibody-mediated rejection (AMR) ○ Synonyms: Humoral rejection or hyperacute rejection ○ Definition: Rejection primarily targeting microvasculature and capillaries due to alloantibodies

CLINICAL ISSUES • Very rare in lung transplant recipients

MICROSCOPIC • Histologic features of AMR in general have not been agreed upon in consensus setting • Pulmonary capillaritis and capillary injury may be seen but are nonspecific • At least 2 back-to-back intracapillary neutrophils • Unexplained diffuse alveolar damage

TOP DIFFERENTIAL DIAGNOSES • Primary graft dysfunction: Complement deposition may occur with ischemia-reperfusion injury

• Infection ○ Presence of neutrophils on biopsy tissue in both ○ C4d stain may be positive in both • Acute cellular rejection (ACR) ○ Perivascular inflammatory cuff (grade A) and airway inflammation (grade B) are key histologic features ○ Neutrophils may be seen in severe ACR ○ Donor-specific antibodies may be found in patients with ACR

DIAGNOSTIC CHECKLIST • Care must be taken to interpret only continuous strong endothelial immunohistochemical (IHC) staining as positive • Interpretation of IHC is difficult due to background staining of elastic fibers both in alveolar septa and blood vessel walls • Hematoxylin and eosin findings are nonspecific • Clinicopathologic correlation is necessary

Antibody-Mediated Rejection

Pneumonia and C4d IHC

C4d IHC Pitfall

C4d IHC Pitfall

(Left) The presence of alveolar capillary neutrophils ﬊, as seen in this lung transplant surveillance biopsy, should raise suspicion for antibodymediated rejection (AMR). Immunofluorescence for C4d was positive in this case. (Right) In this explanted lung allograft, C4d(+) staining of capillaries cannot be attributed to AMR since pneumonia is also present. Note alveolar neutrophils and foamy macrophages ﬈.

(Left) IHC stain for C4d nonspecifically stains the elastic fibers of blood vessels ﬊ and alveolar septa ﬈, as shown here on high power. Note the lack of any endothelial staining. (Right) IHC stain for C4d shows nonspecific interstitial staining of the alveolar septa ﬈ and arteriole ﬊.

388

Antibody-Mediated Rejection, Lung

MICROSCOPIC

Abbreviations

Histologic Features

• Antibody-mediated rejection (AMR)

• Have not been agreed upon in consensus setting ○ Recent International Society for Heart and Lung Transplantation summary statement thoroughly reviews literature • Neutrophilic infiltration score proposed, with ≥ 2 capillary neutrophilia most often being found in AMR ○ Defined as at least 2 back-to-back intracapillary neutrophils ○ Diffuse alveolar damage also found in AMR • Hyperacute rejection associated with preformed alloantibodies ○ Fibrin thrombi in alveolar wall capillaries ○ Fibrinoid necrosis of alveolar walls ○ Alveolar hemorrhage

Synonyms • Humoral rejection • Hyperacute rejection

Definitions • AMR: Rejection primarily targeting microvasculature and capillaries due to alloantibodies • Alloantibodies: Preexisting or newly formed antibodies against target molecules in allograft ○ Alloantibodies also called donor-specific antibodies (DSA)

ETIOLOGY/PATHOGENESIS Proposed Mechanisms • Deposition of alloantibodies within graft vasculature followed by complement activation, endothelial damage, and cell death ○ Can also activate coagulation cascade, resulting in microvascular thrombi • Alloantibodies generally directed against ○ Class I and II major histocompatibility antigens or human leukocyte antigens (HLA) but may also involve non-HLA antigens

CLINICAL ISSUES Epidemiology • Very rare in lung transplant recipients • Risk factors (sensitization) ○ Increased number of postoperative days requiring mechanical ventilation ○ Previous transplantation ○ History of blood product transfusion ○ History of pregnancy

Presentation • Reported to occur within 1st month or months to years after transplantation • Presents with rapid onset of graft dysfunction ○ Clinical presentation is "primary graft dysfunction" – Hypoxia or significant decline in forced expiratory volume in 1 second

Treatment • Drugs ○ Suppression of antibody production – Intravenous immunoglobulin (IVIg) – Rituximab ○ Removal of antibodies from circulation – Plasmapheresis, immunoadsorption

Prognosis • Not well documented ○ Recovery from individual episodes is typical • Associated with development of bronchiolitis obliterans syndrome

Lung Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Primary Graft Dysfunction • Due to poor preservation of donor tissue • Diffuse alveolar damage may be seen in both • Complement deposition may occur with ischemiareperfusion injury

Infection • May occur with AMR • Presence of neutrophils on biopsy tissue in both • Features that favor infection ○ Positive culture from bronchoalveolar lavage fluid ○ Positive viral serology ○ Clinical suspicion • C4d stain may be positive in both

Acute Cellular Rejection • May occur with AMR • Perivascular inflammatory cuff (grade A) and airway inflammation (grade B) are key histologic features • Neutrophils may be seen in severe acute cellular rejection (ACR) • DSA may be found in patients with ACR

SELECTED REFERENCES 1.

2.

3.

4.

5. 6. 7.

8.

Roden AC et al: Diagnosis of acute cellular rejection and antibody-mediated rejection on lung transplant biopsies: a perspective from members of the Pulmonary Pathology Society. Arch Pathol Lab Med. 141(3):437-444, 2017 Basha HI et al: Critical role for IL-17A/F in the immunopathogenesis of obliterative airway disease induced by Anti-MHC I antibodies. Transplantation. 95(2):293-300, 2013 Berry G et al: Pathology of pulmonary antibody-mediated rejection: 2012 update from the Pathology Council of the ISHLT. J Heart Lung Transplant. 32(1):14-21, 2013 Daoud AH et al: Diagnosis and treatment of antibody mediated rejection in lung transplantation: a retrospective case series. Transpl Immunol. 28(1):1-5, 2013 DeNicola MM et al: Pathologic findings in lung allografts with anti-HLA antibodies. J Heart Lung Transplant. 32(3):326-32, 2013 Glanville AR: Antibody-mediated rejection in lung transplantation: Turning myth into reality. J Heart Lung Transplant. 32(1):12-3, 2013 Lobo LJ et al: Donor-specific antibodies are associated with antibodymediated rejection, acute cellular rejection, bronchiolitis obliterans syndrome, and cystic fibrosis after lung transplantation. J Heart Lung Transplant. 32(1):70-7, 2013 Yousem SA et al: The histopathology of lung allograft dysfunction associated with the development of donor-specific HLA alloantibodies. Am J Surg Pathol. 36(7):987-92, 2012

389

Lung Transplantation

Acute Cellular Rejection, Grade A KEY FACTS

TERMINOLOGY • Perivascular rejection • Perivascular inflammatory infiltrate, predominantly lymphocytic

CLINICAL ISSUES • Dyspnea, cough, increased sputum production • Associated with development of bronchiolitis obliterans syndrome (chronic rejection)

MICROSCOPIC • Grade A0: No acute rejection • Grade A1: Minimal acute rejection ○ Alveolated parenchyma with rare scattered perivascular mononuclear cell infiltrates ○ Mononuclear cell infiltrates composed of – Small round or plasmacytoid lymphocytes – Larger transformed lymphocytes ○ Perivascular cuff 2-3 cells thick and circumferential

• Grade A2: Mild acute rejection ○ More frequent perivascular infiltrates – Perivascular cuff 4-8 cell layers thick ○ Alveolated parenchyma with more frequent perivascular infiltrates ○ Easily visible at low-power magnification • Grade A3: Moderate acute rejection ○ Dense, thick perivascular infiltrates with extension into alveolar septa and airspaces ○ Often with endothelialitis • Grade A4: Severe acute rejection ○ Diffuse mononuclear infiltrates in perivascular, interstitial, and alveolar distribution ○ Pneumocyte damage

DIAGNOSTIC CHECKLIST • Grade AX used for inadequate specimens, such as those with no parenchymal vessels

Grade A1 Rejection

Grade A2 Rejection

Mild Acute Perivascular Rejection

Grade A3 Rejection

(Left) Low-power view of posttransplant transbronchial biopsy shows a relatively small perivascular inflammatory cuff (grade A1 rejection). (Right) Low-power view of transbronchial biopsy in this transplant patient shows obvious perivascular cuffing around several vessels. The cuffs are complete and > 3 cell layers thick, making this grade A2.

(Left) High-power view shows activated and mature lymphocytes and one eosinophil ſt in the inflammatory infiltrate around a parenchymal vessel. There is no extension into the lung parenchyma. (Right) Grade A3 acute cellular rejection is characterized by perivascular inflammatory infiltrates, which are much more extensive and involve the adjacent alveolar walls, as seen here.

390

Acute Cellular Rejection, Grade A

Abbreviations • Acute cellular rejection (ACR)

Synonyms • Perivascular rejection

Definitions • Perivascular inflammatory infiltrate, predominantly lymphocytic

CLINICAL ISSUES Presentation • May be asymptomatic • Dyspnea, cough, sputum production, fever, or hypoxia

Treatment • Increase or change immunosuppressant regimen • Tacrolimus often used in recurrent acute rejection

Prognosis • Associated with development of bronchiolitis obliterans syndrome (chronic rejection)

MICROSCOPIC Histologic Features • Grade A0: No acute rejection ○ Normal parenchyma without mononuclear cell infiltrates • Grade A1: Minimal acute rejection ○ Alveolated parenchyma with rare scattered perivascular mononuclear cell infiltrates ○ Mononuclear cell infiltrates composed of – Small round or plasmacytoid lymphocytes – Larger transformed lymphocytes ○ Perivascular cuff – 2- to 3-cell thick infiltrate in adventitia – Incomplete cuff or circumferential ○ Usually no eosinophils and no endothelialitis • Grade A2: Mild acute rejection ○ Alveolated parenchyma with more frequent perivascular infiltrates – Around arterioles, venules, and lymphatics ○ Infiltrates composed of – Small round and larger activated lymphocytes – Plasmacytoid lymphocytes and macrophages – Few eosinophils and rare neutrophils ○ Perivascular cuff – Easily visible at low-power magnification – ≥ 4 cell layers thick ○ Endothelialitis may be present – Subendothelial infiltrates of mononuclear cells – Endothelial hyperplasia or regenerative changes ○ May have concurrent lymphocytic bronchiolitis (i.e., grade B acute rejection) – Small airways with submucosal inflammation ○ No extension of infiltrates into alveolar septa or air spaces • Grade A3: Moderate acute rejection ○ Dense perivascular mononuclear cell infiltrates

– Often with associated endothelialitis (lifting of endothelial layer by inflammatory infiltrate) – Infiltrates extend into alveolar septa and airspaces as single cells or sheets – Continuity between perivascular and interstitial infiltrates – May see alveolar macrophage accumulation and type II pneumocyte hyperplasia ○ Infiltrate composed of – Lymphocytes, eosinophils, and occasional neutrophils ○ Perivascular cuff – Easily recognizable at low power • Grade A4: Severe acute rejection ○ Very rare with current immunosuppressive regimens ○ Diffuse mononuclear infiltrates in perivascular, interstitial, and alveolar distribution ○ Pneumocyte damage present ○ Endothelialitis often seen ○ Alveolar spaces may contain – Sloughed necrotic pneumocytes and macrophages – Hyaline membranes and hemorrhage – Neutrophilic inflammation

Lung Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Primary Graft Dysfunction • Diffuse alveolar damage often present • Distinguish from grade ACR by absence of perivascular and interstitial mononuclear cells

Antibody-Mediated Rejection • Capillaritis or capillary injury with neutrophils and lack of mononuclear infiltrates • May occur simultaneously with ACR

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Features distinguishing grade A2 from grade A1 ○ Thicker perivascular cuff of mononuclear cells in A2 ○ Presence of endothelialitis &/or eosinophils in A2 ○ Either can have coexistence of lymphocytic bronchiolitis • Features distinguishing grade A3 from grade A2 ○ Infiltrates extend continuously from perivascular location into alveolar septa in A3 ○ Presence of occasional neutrophils more common in A3 • Features distinguishing grade A4 from grade A3 ○ Pneumocyte damage and necrosis with sloughing into alveolar spaces in A4 ○ Hyaline membranes and alveolar hemorrhage in A4 • Grade AX used for inadequate specimens, such as those with no parenchymal vessels

SELECTED REFERENCES 1.

2.

3.

Roden AC et al: Diagnosis of acute cellular rejection and antibody-mediated rejection on lung transplant biopsies: a perspective from members of the Pulmonary Pathology Society. Arch Pathol Lab Med. 141(3):437-444, 2017 Stewart S et al: Revision of the 1996 working formulation for the standardization of nomenclature in the diagnosis of lung rejection. J Heart Lung Transplant. 26(12):1229-42, 2007 Yousem SA et al: Revision of the 1990 working formulation for the classification of pulmonary allograft rejection: Lung Rejection Study Group. J Heart Lung Transplant. 15(1 Pt 1):1-15, 1996

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Lung Transplantation

Acute Cellular Rejection, Grade B KEY FACTS

MICROSCOPIC

TOP DIFFERENTIAL DIAGNOSES

• Grade B0: No airway inflammation • Grade B1R: Low-grade small airway inflammation ○ Bronchiolar submucosal inflammation with no fibrosis ○ Inflammation does not involve airway epithelium ○ Grade B1R: Combination of grades B1 and B2 from 1996 ISHLT classification • Grade B2R: High-grade small airway inflammation ○ Bronchiolar submucosal inflammation with epithelial damage ○ Submucosa expanded by inflammatory infiltrate ○ Eosinophils and plasmacytoid cells present ○ Epithelial cell metaplasia, apoptosis, and necrosis ○ Grade B2R: Combination of grades B3 and B4 from 1996 ISHLT classification • Grade BX: Ungradeable small airway inflammation ○ Sampling issue, infection, tangential cut, artifact

• Infection ○ Airway submucosa and epithelium with more neutrophils than mononuclear cells • Bronchial-associated lymphoid tissue ○ Circumscribed nodular submucosal collection of lymphocytes and rare eosinophils without epithelial damage ○ CD21 immunohistochemical stain highlights characteristic follicular dendritic cell network

DIAGNOSTIC CHECKLIST • Grade B1R: ISHLT 2007 terminology combines grades B1 and B2 from 1996 ISHLT classification • Grade B2R: ISHLT 2007 terminology combines grades B3 and B4 from 1996 ISHLT classification

Mild Acute Rejection (Grade B2/B1R)

Moderate Acute Rejection (Grade B3/B2R)

Bronchial-Associated Lymphoid Tissue

Bronchial-Associated Lymphoid Tissue

(Left) There is a band-like mixed inflammatory infiltrate in the submucosa, which does not damage the overlying respiratory epithelium. (Right) There is much more inflammatory infiltrate than that seen in mild rejection, which extends into and partly destroys the overlying epithelium ﬉.

(Left) A small collection of mature lymphocytes, as seen here, should not be interpreted as acute rejection (it is neither airway nor perivascular). Also, there are not activated lymphocytes or eosinophils. (Right) CD21 stain highlights the dendritic cell network in the center, which confirms that this is indeed BALT and not acute rejection.

392

Acute Cellular Rejection, Grade B

Abbreviations • Acute cellular rejection (ACR)

Synonyms • Airway inflammation • Lymphocytic bronchiolitis (LB)

Definitions • Inflammation around bronchioles (small airways) representing acute rejection of airways, which can be diagnosed on transbronchial biopsy

ETIOLOGY/PATHOGENESIS Environmental Exposure • Changes in particulate air pollution associated with increased risk for LB in one study

○ Grade B2R is combination of grade B3 and grade B4 from 1996 ISHLT classification – Grade B3 (moderate airway inflammation): Similar to grade B2 but more intense and with epithelial cell damage – Grade B4 (severe airway inflammation): Grade B3 plus epithelial ulceration and sloughing into lumen with fibrinopurulent debris and neutrophils • Grade BX: Ungradeable small airway inflammation ○ No airway in biopsy, or tangential cutting or processing artifact ○ Evidence of infection

Lung Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Chronic Rejection (Bronchiolitis Obliterans) • Airway fibrosis always present, which may contain inflammatory infiltrate

Infection

CLINICAL ISSUES Prognosis • Associated with increased risk of chronic rejection (bronchiolitis obliterans syndrome)

MICROSCOPIC

• Airway submucosa and epithelium with more neutrophils than mononuclear cells • Increased eosinophils as compared to mononuclear cells should also raise suspicion • Bronchoalveolar lavage with culture can aid in identifying etiology

Histologic Features

Bronchial-Associated Lymphoid Tissue

• Grade B0: No airway inflammation ○ No bronchiolar inflammation • Grade B1R: Low-grade small airway inflammation ○ Bronchiolar submucosal inflammation – Infrequent, scattered, mononuclear inflammatory cells and occasional eosinophils – May form incomplete or circumferential band around bronchiole ○ Inflammation does not involve airway epithelium – No damage to airway epithelium – Airway epithelium may show mild hyperplasia ○ Grade B1R is combination of grade B1 and grade B2 from 1996 International Society for Heart & Lung Transplantation (ISHLT) classification – Grade B1 (minimal airway inflammation): Rare scattered mononuclear cells within airway submucosa – Grade B2 (mild airway inflammation): Circumferential airway submucosal band of mononuclear cells, some eosinophils, intraepithelial lymphocytes, without epithelial cell damage • Grade B2R: High-grade small airway inflammation ○ Bronchiolar submucosal inflammation with epithelial damage – Submucosa expanded by inflammatory infiltrate – Mononuclear inflammatory cells larger (activated lymphocytes) and often extend into epithelium – Eosinophils and plasmacytoid cells present – Epithelial cell metaplasia, apoptosis, and necrosis – Very severe forms can have epithelial ulceration with neutrophils, fibrinopurulent exudate, and sloughed cellular debris – More mononuclear cells than neutrophils present; if more neutrophils, likely infection

• Circumscribed nodular submucosal collection of lymphocytes and rare eosinophils without epithelial damage • Located adjacent to airways and often contains anthracotic pigment • CD21 immunohistochemical stain helps highlight characteristic follicular dendritic cell network

Acute Cellular Rejection, Grade A • May coexist with grade B ACR

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Grade B1R: ISHLT 2007 terminology combines grades B1 and B2 from 1996 ISHLT classification • Grade B2R: ISHLT 2007 terminology combines grades B3 and B4 from 1996 ISHLT classification

SELECTED REFERENCES 1. 2. 3. 4.

5.

6.

Husain AN et al: Lung transplantation: the state of the airways. Arch Pathol Lab Med. 140(3):241-4, 2016 Bhorade SM et al: Interobserver variability in grading transbronchial lung biopsies after lung transplantation. Chest. 143(6): 1717-24, 2013 Gordon IO et al: SaLUTaRy: survey of lung transplant rejection. J Heart Lung Transplant. 31(9):972-9, 2012 Verleden SE et al: Lymphocytic bronchiolitis after lung transplantation is associated with daily changes in air pollution. Am J Transplant. 12(7):1831-8, 2012 Hodge G et al: Lymphocytic bronchiolitis is associated with inadequate suppression of blood T-cell granzyme B, IFN-gamma, and TNF-alpha. Transplantation. 89(10):1283-9, 2010 Stewart S et al: Revision of the 1996 working formulation for the standardization of nomenclature in the diagnosis of lung rejection. J Heart Lung Transplant. 26(12):1229-42, 2007

393

Lung Transplantation

Chronic Allograft Dysfunction, Lung KEY FACTS

TERMINOLOGY • Clinical: Bronchiolitis obliterans syndrome (BOS) • Chronic lung allograft dysfunction (CLAD) recently defined to cover all chronic graft dysfunction, including obstructive and restrictive forms • BOS ○ Progressive obstructive physiology ○ Permanent loss of forced expiratory volume at end of 1st second of expiration ○ Bronchiolitis obliterans (BO) is pathologic hallmark • Restrictive allograft syndrome manifests as ○ Progressive, restrictive physiology with increasing fibrosis on radiology ○ Often coexists with BOS • Pleuropulmonary fibrosis is main pathologic finding

CLINICAL ISSUES

• CLAD is most common cause of death in adult and pediatric lung transplant recipients after 1st year

MICROSCOPIC • In BO, asymmetric or concentric submucosal fibrosis of small airways ○ Results in partial or complete luminal occlusion ○ Associated airway submucosal and peribronchiolar inflammation may be present • Chronic vascular rejection characterized by ○ Venous and arterial fibrointimal thickening by paucicellular hyaline sclerosis • Chronic rejection is difficult to diagnose on biopsy due to ○ Patchy nature of changes ○ Lack of sufficient airway sampling • RAS: Fibrosis of large areas of parenchyma and pleura • Almost all patients also have BO

• Can occur weeks to months or years after transplantation

Normal Lung

Chronic Lung Allograft Dysfunction

Bronchiolitis Obliterans

Chronic Vascular Rejection

(Left) This section of the lung is from a 13-year-old child who died of a gunshot wound. The bronchiole and pulmonary artery are about the same diameter, and there is very little submucosal fibrous tissue ﬈. (Right) Chronic rejection in this lung transplant recipient is manifested by bronchiolitis obliterans. Note submucosal fibrosis partially occluding the lumen ﬉ without inflammation (grade Cb).

(Left) There is complete occlusion of the airway lumen by fibrosis. The airway wall is recognizable by a discontinuous muscle layer ﬉ and normal accompanying pulmonary artery ﬈. (Right) Elastic stain demonstrates subtotal occlusion of the pulmonary artery lumen due to intimal fibrosis. The arterial wall elastic fibres are easily seen here.

394

Chronic Allograft Dysfunction, Lung

Definitions

Prognosis

• Chronic lung allograft dysfunction (CLAD) recently defined to cover all chronic graft dysfunction, including obstructive and restrictive forms • Bronchiolitis obliterans syndrome (BOS) ○ Progressive obstructive physiology ○ Permanent loss of forced expiratory volume at end of 1st second of expiration ○ Bronchiolitis obliterans (BO) is pathologic hallmark • Restrictive allograft syndrome (RAS) manifests as ○ Progressive, restrictive physiology with increasing fibrosis on radiology ○ Pleuropulmonary fibrosis is main pathologic finding ○ Often coexists with BOS

• CLAD is most common cause of death in adult and pediatric lung transplant recipients after 1st year

Synonyms

ETIOLOGY/PATHOGENESIS Multifactorial • Increased risk of BO with ○ Acute perivascular rejection ○ Lymphocytic bronchitis and bronchiolitis (acute airway rejection) ○ Infection, especially viral or fungal ○ Gastroesophageal reflux disease/microaspiration and gastroparesis ○ Reperfusion injury and extended graft ischemic time • Important molecules in fibroproliferative process include ○ Platelet-derived growth factor B ○ Transforming growth factor β ○ Chemokine (CXC motif) receptor 2

Lung Transplantation

• Chronic rejection

• Progression of established BOS may be slowed by lymphocyte-depletion therapies • Retransplantation has some success with 5-year survival rate of ~ 50%

TERMINOLOGY

MICROSCOPIC Histologic Features • BO: Asymmetric or concentric submucosal fibrosis of small airways ○ Results in partial or complete luminal occlusion ○ Can have associated fragmentation and destruction of airway smooth muscle and elastic tissue ○ Associated obstructive changes include mucostasis &/or foamy macrophages in distal airspaces ○ Early lesions show polypoid protrusion of submucosal fibrosis into lumen ○ Associated airway submucosal and peribronchiolar inflammation may be present in BO ○ Presence or absence of inflammation not categorized in 2007 International Society for Heart and Lung Transplantation classification • Chronic vascular rejection is characterized by ○ Venous and arterial fibrointimal thickening by paucicellular hyaline sclerosis, ± inflammation • Chronic rejection is difficult to diagnose on biopsy due to patchy nature of changes and lack of sufficient airway sampling ○ May see associated changes of mucostasis &/or foamy macrophages, which are suggestive of BO • RAS: Fibrosis of large areas of parenchyma and pleura ○ Almost all patients also have BO

Role of Immune System in Bronchiolitis Obliterans • Autoimmune response to airway epithelial self-antigens leads to Th17/IL-17 pathway activation ○ Induced by alloimmune response to donor human leukocyte antigen • Genetic polymorphisms have been described ○ In innate immune system receptors ○ In other immune pathway molecules • Association with presence of donor-specific antibodies in cystic fibrosis patients

CLINICAL ISSUES Epidemiology • Incidence ○ BOS occurs in 50% by 5 years and in 76% by 10 years ○ RAS occurs in significant minority of patients

Presentation • Can occur weeks to months or years after transplantation • Dyspnea, cough • Worsening pulmonary function tests

Treatment • Focuses on prevention by treatment of known risk factors • Immunosuppressive therapies have not been effective

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Submucosal airway fibrosis of BO results in narrowing of lumen and leads to obstructive signs and symptoms • Lung parenchyma fibrosis usually leads to rapidly progressive restrictive lung disease • CLAD is clinical diagnosis; biopsy often not needed

SELECTED REFERENCES 1.

2. 3.

4. 5.

6.

Ruttens D et al: An association of particulate air pollution and traffic exposure with mortality after lung transplantation in Europe. Eur Respir J. 49(1), 2017 Husain AN et al: Lung transplantation: the state of the airways. Arch Pathol Lab Med. 140(3):241-4, 2016 Goldfarb SB et al: The Registry of the International Society for Heart and Lung Transplantation: eighteenth official pediatric lung and heart-lung transplantation report--2015; focus theme: early graft failure. J Heart Lung Transplant. 34(10):1255-63, 2015 Verleden SE et al: Linking clinical phenotypes of chronic lung allograft dysfunction to changes in lung structure. Eur Respir J. 46(5):1430-9, 2015 Yusen RD et al: The Registry of the International Society for Heart and Lung Transplantation: thirty-second official adult lung and heart-lung transplantation report--2015; focus theme: early graft failure. J Heart Lung Transplant. 34(10):1264-77, 2015 Verleden GM et al: A new classification system for chronic lung allograft dysfunction. J Heart Lung Transplant. 33(2):127-33, 2014

395

Lung Transplantation

Organizing Pneumonia KEY FACTS

TERMINOLOGY • Reversible distal airways and intraalveolar proliferation of fibroblasts and myofibroblasts • Bronchiolitis obliterans with organizing pneumonia ○ Older term, not recommended now

ETIOLOGY/PATHOGENESIS • 4 stages of pathogenesis: Injury, proliferation, maturation, and resolution • In transplant setting, etiology related to infection, acute rejection, &/or drug toxicity

MICROSCOPIC • Nodules of fibroblasts and myofibroblasts arranged in whorls with pale gray matrix (Masson bodies) within alveoli and distal airspaces ○ Capillaries, macrophages, and few inflammatory cells often present within Masson bodies

• Type 2 pneumocyte hyperplasia, mild chronic interstitial inflammation, and intraalveolar macrophages often present • Patchy distribution with preserved lung architecture and no interstitial fibrosis • Preserved lung architecture without interstitial fibrosis • If areas away from organizing pneumonia (OP) pattern are abnormal, consider OP superimposed on or coexistent with another process

TOP DIFFERENTIAL DIAGNOSES • Usual interstitial pneumonia ○ Fibroblastic foci, located within airway wall (interstitial) – May be confused with Masson bodies of OP, especially if epithelialization over nodules occurs • Organizing diffuse alveolar damage ○ Interstitial organizing fibroblastic proliferation often with extensive type 2 pneumocyte hyperplasia

Masson Bodies

Masson Body

Cryptogenic Organizing Pneumonia

Intrabronchial Masson Body

(Left) Low-power view of organizing pneumonia shows several intraalveolar rounded ﬈ and elongated st balls of fibrous tissue (Masson bodies), which compress adjacent, relatively normal lung. (Right) At high power, the intraalveolar Masson body is seen to be composed of fibroblasts ﬈ with abundant intercellular matrix ﬉. Note chronic inflammatory cells present ﬊ within the Masson body.

(Left) This organizing pneumonia presented as a mass that was resected. The mass consists of nodules of fibrosis. Normal lung tissue is present on the right. (Right) A Masson body is filling the bronchiolar lumen. Note the residual bronchial mucosa ſt.

396

Organizing Pneumonia

Abbreviations • Organizing pneumonia (OP)

Synonyms • OP pattern • Bronchiolitis obliterans with OP ○ Older term, not recommended now

Definitions • Reversible distal airway and alveolar proliferation of fibroblasts and myofibroblasts

ETIOLOGY/PATHOGENESIS

Treatment • Corticosteroids • Antibiotics or antivirals if infectious etiology • Stop administering or lower dose of offending agent if associated with drug toxicity

IMAGING CT Findings • Typical pattern: Patchy airspace consolidation in peribronchial distribution • Diffuse pattern: Diffuse infiltrative opacities or reticulonodular pattern

MICROSCOPIC

Infectious Agents

Histologic Features

• Resolution of infectious bronchopneumonia can progress through phase of OP pattern

• Nodules of fibroblasts and myofibroblasts arranged in whorling pattern with pale gray matrix (Masson bodies) ○ Located within alveolar ducts and alveoli – Can also involve distal airway spaces (bronchioles) ○ Patchy and apparent bronchiolocentric distribution • Preserved lung architecture without interstitial fibrosis • Mild chronic interstitial inflammation may be present • Type 2 pneumocyte hyperplasia often seen • May see increased airspace macrophages • Uninvolved areas histologically normal ○ If areas away from OP pattern are abnormal, consider OP superimposed on or coexistent with another process • Epithelialization can occur over individual nodules, making recognition of intraalveolar location difficult

Drug Toxicity • Response to direct cytotoxic effect on pneumocytes

Acute Rejection • Pneumocyte damage can occur in moderate to severe acute rejection (AR) or in repeated episodes of lower grade AR

Cryptogenic • Unknown insult to alveolar pneumocytes • Not thought to occur in setting of lung transplantation ○ Usually identifiable causative factor

Lung Transplantation

TERMINOLOGY

Stages of Pathogenesis • Injury phase ○ Insult to alveolar epithelium resulting in type I pneumocyte necrosis and epithelial denudation ○ Plasma proteins enter alveolar spaces ○ Activation of coagulation cascade, resulting in fibrin deposition ○ Intraluminal migration of acute and chronic inflammatory cells in response to fibrin • Proliferating phase ○ Inflammatory cells, including macrophages fragment fibrin ○ Activated fibroblasts enter lumen, proliferate, and differentiate into myofibroblasts ○ Inflammatory cells and fibrin are replaced by fibroblasts and myofibroblasts with collagen-rich loose connective tissue matrix ○ Alveolar basement membrane reepithelializes • Mature phase ○ Fibroblasts and myofibroblasts arrange in concentric rings alternating with layers of collagen matrix • Resolution phase ○ Dissipation of alveolar nodules and return to normal cellularity and architecture ○ Preserved alveolar basement membrane critical for complete resolution

CLINICAL ISSUES Presentation • Cough, dyspnea, flu-like symptoms

DIFFERENTIAL DIAGNOSIS Usual Interstitial Pneumonia • Fibroblastic foci, located within airway wall (interstitial) ○ May be confused with Masson bodies of OP, especially if epithelialization over nodules occurs • Areas of OP often present in usual interstitial pneumonia

Organizing Diffuse Alveolar Damage • Interstitial organizing fibroblastic proliferation often with extensive type 2 pneumocyte hyperplasia • May see residual features of acute phase (e.g., hyaline membranes &/or vascular fibrin thrombi) if in transition to organizing phase

Hypersensitivity Pneumonitis • OP pattern may be component of hypersensitivity pneumonitis • Would also see granulomas, interstitial fibrosis, and chronic inflammation

Eosinophilic Pneumonia • OP pattern may be component of eosinophilic pneumonia • Would also see eosinophils, alveolar macrophages with eosinophilic granular material, and alveolar fibrin

SELECTED REFERENCES 1. 2.

Zare Mehrjardi M et al: Radio-pathological correlation of organizing pneumonia (OP): a pictorial review. Br J Radiol. 90(1071):20160723, 2017 Tabaj GC et al: Histopathology of the idiopathic interstitial pneumonias (IIP): a review. Respirology. 20(6):873-83, 2015

397

Lung Transplantation

Microaspiration KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Microscopic aspiration of gastric contents ○ Usually associated with symptoms of gastroesophageal reflux disease (GERD)

• Exogenous lipoid pneumonia ○ Single or multiple large fat vacuoles within macrophages (intracytoplasmic) • Multinucleated giant cells within alveoli or interstitium ○ ± foreign material (e.g., degenerating food particles, cellulose from tablets, barium crystals)

CLINICAL ISSUES • GERD is common in pediatric and adult lung transplant patients • May be asymptomatic or present with ○ Repeated episodes of acute cellular rejection (ACR) ○ Symptoms of GERD – Heartburn – Regurgitation – Nausea – Dysphagia • Precipitates ACR • Correlation reported with BOS ○ Both in retrospective analysis and several prospective studies

TOP DIFFERENTIAL DIAGNOSES • Rejection ○ ACR or chronic airway rejection • Artifact ○ Air bubbles in tissue, which are round vacuoles but are not intracellular • Macroaspiration (aspiration pneumonia/pneumonitis) ○ Bronchopneumonia with foreign material ○ Clinically obtunded patient

Microaspiration With Giant Cells

Air Bubble Artifact

Microaspiration With Fibrosis

Massive Aspiration Pneumonia

(Left) Histologic changes of microaspiration can be very subtle and easily missed, as shown here. Within the alveolar spaces, there are 2 multinucleated giant cells with large vacuoles ﬈. (Right) Not all vacuoles are due to aspiration of lipid material. The holes shown here ﬈ are likely to be air artifacts. They are not intracellular and can be easily mistaken for exogenous lipoid pneumonia.

(Left) Small multinucleated giant cells ﬈ are present in the interstitium, which is widened by fibrosis and inflammation including eosinophils. (Right) Aspiration pneumonia, as shown here, is distinct from microaspiration. Note numerous neutrophils and oral squamous cells.

398

Microaspiration

Synonyms

CT Findings • Diffuse basilar centrilobular nodules • Tree-in-bud pattern with airway and interstitial thickening

• Exogenous lipoid pneumonia

Definitions • Microscopic aspiration of gastric contents ○ Usually associated with symptoms of gastroesophageal reflux disease (GERD)

ETIOLOGY/PATHOGENESIS Multifactorial Predisposition to GERD • Diminished cough reflex ○ Transection of bronchial nerves during transplant operation • Operative trauma to proximal aspects of vagus nerve • Postoperative gastroparesis • Immunosuppressants may lead to decreased lower esophageal sphincter pressure • Esophageal dysfunction in patients with connective tissue disease • Obesity and obstructive sleep apnea

CLINICAL ISSUES Epidemiology • Age ○ GERD common in pediatric and adult lung transplant patients

Presentation • • • •

May be asymptomatic Repeated episodes of acute cellular rejection (ACR) Cough, shortness of breath Symptoms of GERD ○ Heartburn ○ Regurgitation ○ Nausea ○ Dysphagia

Treatment • Surgical approaches ○ Nissen fundoplication – Proposed standard of care operation in both adult and pediatric patients after lung transplantation • Drugs ○ Proton pump inhibitor for GERD • Clinical ○ Raise head of bed ○ Go to bed on empty stomach

Prognosis • Precipitates ACR • Correlation reported with BOS ○ Both in retrospective analysis and several prospective studies • Overall survival may not be affected

IMAGING Radiographic Findings • Lung opacities

MICROSCOPIC Histologic Features • Exogenous lipoid pneumonia ○ Single or multiple large fat vacuoles within macrophages that may push nucleus to one side ○ Macrophages are intraalveolar or interstitial ○ Can be seen on transbronchial biopsy • Multinucleated giant cells within alveoli or interstitium ± foreign material (e.g., degenerating food particles, cellulose from tablets, barium crystals) • Minimal inflammatory infiltrates of lymphocytes and neutrophils • Fibrinous exudate and organizing pneumonia • May see features of ACR ○ Airway submucosal ± intraepithelial inflammation ○ Perivascular inflammatory cuff

Lung Transplantation

TERMINOLOGY

ANCILLARY TESTS Cytology • Quantitative assessment of oil red O-stained alveolar macrophages in bronchoalveolar lavage specimens proposed to detect aspirated lipid

DIFFERENTIAL DIAGNOSIS Acute Cellular Rejection • May be concurrent

Infection • Viral or fungal • Tracheobronchitis

Chronic Airway Rejection • Submucosal small airways fibrosis ± inflammation

Macroaspiration (Aspiration Pneumonia/Pneumonitis) • Bronchopneumonia with foreign material • Clinically obtunded patient • Not often biopsied; seen at autopsy

Endogenous Lipoid Pneumonia/Postobstructive • Small lipid vacuoles • No foreign body type multinucleated giant cells

Artifact • Air bubbles are round vacuoles but are not intracellular

SELECTED REFERENCES 1.

2. 3. 4.

Lo WK et al: Both pre-transplant and early post-transplant antireflux surgery prevent development of early allograft injury after lung transplantation. J Gastrointest Surg. 20(1):111-8; discussion 118, 2016 Patti MG et al: The intersection of GERD, aspiration, and lung transplantation. J Laparoendosc Adv Surg Tech A. 26(7):501-5, 2016 Cardasis JJ et al: The spectrum of lung disease due to chronic occult aspiration. Ann Am Thorac Soc. 11(6):865-73, 2014 Fisichella PM et al: Aspiration, localized pulmonary inflammation, and predictors of early-onset bronchiolitis obliterans syndrome after lung transplantation. J Am Coll Surg. 217(1):90-100; discussion 100-1, 2013

399

Lung Transplantation

Bacterial Infections KEY FACTS

ETIOLOGY/PATHOGENESIS • • • • • •

Vancomycin-resistant Enterococcus (VRE) Methicillin-resistant Staphylococcus aureus (MRSA) Streptococcus pneumoniae Haemophilus influenzae Pseudomonas aeruginosa Mycobacteria ○ Tuberculous and nontuberculous • Carbapenem-resistant gram-negative bacteria • Clostridium difficile

CLINICAL ISSUES

○ Opportunistic infections are more common • After 6 months ○ Community-acquired pneumonia and urinary tract infections occur

MICROSCOPIC • Neutrophils within airway submucosa and involving airway epithelium suggest bacterial infection • Intraalveolar fibrin may also be present • Inflammatory response to bacterial growth may be lacking

TOP DIFFERENTIAL DIAGNOSES • Airway inflammation with mostly lymphocytes

• > 50% of lung transplant recipients die from infectious complications ○ Mostly bacterial • ~ 70% of infections occur within 1st year after transplant ○ Most bacterial infections occur within 2 weeks to 1 month after lung transplantation • Between 1-6 months

DIAGNOSTIC CHECKLIST • Diagnosis depends on clinical-pathologic correlation with emphasis on microbiological cultures

Airway Infection

Bacterial Pneumonia

Necrotizing Pneumonia

Bacterial Pneumonia

(Left) Predominance of intraepithelial and submucosal neutrophils as seen on this transbronchial biopsy is highly suggestive of infection that may be bacterial; correlation with culture is needed for diagnosis. (Right) Often, lung transplant recipients are quite immunocompromised, allowing bacteria ﬈ to grow around blood vessels and in lung tissue without any inflammatory response, as seen here at autopsy.

(Left) Microscopic examination shows areas of degenerating neutrophils as well as necrotic parenchyma. Note small bluish rounded structures ﬊, calcium in this case, which should prompt GMS stain to look for fungus. (Right) A small amount of inflammation with neutrophils and fibrin is shown on lung biopsy, suggestive of bacterial infection.

400

Bacterial Infections

Definitions • Bacterial infection of lung, pleura, mediastinum, serum, urinary tract, skin, or gastrointestinal tract

ETIOLOGY/PATHOGENESIS Infectious Agents • • • • • • •

Vancomycin-resistant Enterococcus (VRE) Methicillin-resistant Staphylococcus aureus (MRSA) Streptococcus pneumoniae Haemophilus influenzae Pseudomonas aeruginosa Mycobacteria, tuberculous (TB), and nontuberculous (NTM) Carbapenem-resistant Acinetobacter baumannii and Klebsiella pneumoniae • Clostridium difficile

Sources of Infection • Allograft lung, remaining native lung • Hospital or community acquired

Microaspiration • Impaired function of gastroesophageal junction with reflux and impaired cough reflex • Introduces oral flora into lung environment • Oral flora may become pathogenic in these immunocompromised patients

CLINICAL ISSUES Epidemiology • ~ 70% of infections occur within 1st year after transplant ○ Most bacterial infections occur within 2 weeks to 1 month after lung transplantation ○ VRE and MRSA are most common in 1st month ○ Opportunistic infections are more common between 1-6 months ○ Community-acquired pneumonia and urinary tract infections occur after 6 months – Reported S. pneumoniae infection rate of 6.4% at median of 1.3 years post transplantation despite antibiotic prophylaxis • P. aeruginosa colonization associated with increased risk of bronchiolitis obliterans (BO) syndrome within 2 years of transplant • TB incidence ranges from 2.5-10.0% • NTM incidence is 1.1/100 person-years, mostly from Mycobacterium avium complex • S. aureus bacteremia reported in 10-30% of lung transplant recipients • Urinary tract infections reported in 3% of lung transplant recipients • C. difficile colitis due to antibiotic exposure reported in 7.4% of lung transplant recipients • Staphylococcus species skin infections affect 5.5% of lung transplant recipients

• Complications from infection are most common cause of death in BO • NTM infection associated with increased risk of death in primary lung transplant patients • Fatal infections reported from donor transmission of carbapenem-resistant organisms

Prevention • Pneumococcal vaccination recommended pretransplant and at regular intervals post transplant • H. influenzae type B vaccination recommended in pediatric recipients • Yearly influenza vaccination also recommended • Vaccination not shown to increase rejection risk in other solid organ transplant recipients

Lung Transplantation

TERMINOLOGY

MACROSCOPIC Autopsy Findings • Greenish-yellow exudate and areas of consolidation

MICROSCOPIC Histologic Features • On biopsy, neutrophils within airway submucosa and involving airway epithelium suggest bacterial infection • Fibrin may be present • Significant lung inflammation may be lacking in gramnegative infections • Severely immunosuppressed patients may lack inflammatory response, especially at autopsy • Look for multiple sources or types of infection due to immunosuppressed state

ANCILLARY TESTS Microbiology Culture • Obtain additional tissue and send for culture and sensitivity at time of transbronchial biopsy for histology • Necessary for definitive diagnosis

DIFFERENTIAL DIAGNOSIS Acute Cellular Rejection, Grade B • Airway inflammation with mostly lymphocytes • Few neutrophils may be present within epithelium ○ Nonspecific response to bronchoscopy procedure ○ Coexisting bacterial or other infection

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Histology can often only suggest bacterial infection • Diagnosis depends on clinical-pathologic correlation with emphasis on microbiological cultures

SELECTED REFERENCES 1.

Prognosis

2.

• > 50% of lung transplant recipients die from infectious complications ○ Bacterial infections are most common

3.

Troxell ML et al: Practical applications in immunohistochemistry: evaluation of rejection and infection in organ transplantation. Arch Pathol Lab Med. 140(9):910-25, 2016 Martin-Gandul C et al: The impact of infection on chronic allograft dysfunction and allograft survival after solid organ transplantation. Am J Transplant. 15(12):3024-40, 2015 Witt CA et al: Pulmonary infections following lung transplantation. Thorac Surg Clin. 22(3):403-12, 2012

401

Lung Transplantation

Viral Infections KEY FACTS

TERMINOLOGY • Viral infection in lung transplant recipient affecting lungs, liver, or other organs

ETIOLOGY/PATHOGENESIS • Cytomegalovirus (CMV) ○ Seropositive donor transmits virus via lymphocytes containing latent virus within allograft ○ Seropositive recipient may have reactivation of infection associated with immunosuppression

CLINICAL ISSUES • Symptomatic CMV pneumonia has significantly decreased from 50% incidence of 1980s • Overall risk of EBV-associated PTLD (not necessarily pulmonary involvement) after lung transplant: 1% at 1 year and 1.5 % at 5 years

○ Enlarged infected cells ○ Single nuclear inclusion with halo ("owl's eye") ○ Granular amphophilic cytoplasmic inclusions • Pulmonary EBV-associated PTLD ○ Lymphocytic vasculitis with fibrinoid necrosis ○ Sheets of cells with individual and focal coagulative necrosis ○ Range cytologically from immunoblastic cells, large lymphoid cells, and small round lymphocytes ○ Brisk mitoses • Due to immunosuppression, coinfection with multiple agents is not uncommon and must be considered in differential, as histologic (and radiologic and clinical) features often overlap

ANCILLARY TESTS • For CMV, HSV, adenovirus, EBV

MICROSCOPIC • CMV pneumonia

Pulmonary Posttransplant Lymphoproliferative Disorder

Posttransplant Lymphoproliferative Disorder (EBER)

HSV Pneumonia

HSV1 and 2 (IHC, Red Chromogen)

(Left) This high-power view of a posttransplant lymphoproliferative lesion shows characteristic sheets of dyscohesive large lymphoid cells with prominent nucleoli at varying stages of maturity. (Right) This stain for active Epstein-Barr virus (EBER) demonstrates nuclear activity in a significant proportion of the lymphoid proliferation.

(Left) This patient died of rapidly progressive pneumonia. Within a focus of lung necrosis, multiple smudged intranuclear inclusions are identified ſt, which are most consistent with herpes simplex virus (HSV). (Right) IHC stain (red chromogen) for HSV1 and HSV2 confirms the virus seen on H&E.

402

Viral Infections

Definitions • Viral infection in lung transplant recipient affecting lungs, liver, or other organs

ETIOLOGY/PATHOGENESIS Infectious Agents • Cytomegalovirus (CMV) ○ Seropositive donor transmits virus via donor lymphocytes containing latent virus within allograft ○ Seropositive recipient may have – De novo infection – Reactivation of prior infection associated with immunosuppression • Herpes simplex virus (HSV) and varicella zoster virus (VZV) ○ Most cases due to reactivation • Human herpesviruses 6 and 7 (HHV-6 and HHV-7) ○ Frequently detected due to reactivation • Epstein-Barr virus (EBV) infection leading to posttransplant lymphoproliferative disorder (PTLD) ○ Seropositive donor transmission of latent virus or reactivation of recipient latent virus – Concurrent immunosuppression releases B cells from counter-regulatory effects of T cells, eventually resulting in clonal B-cell proliferation • Respiratory syncytial virus (RSV) ○ Common community-acquired childhood infection • Adenovirus ○ Due to progressive and rapid early posttransplantation course, theorized to either be acquired from upper respiratory tract of recipient, or originating and reactivating from donor lung • Other community-acquired respiratory viruses ○ Influenza, parainfluenza, rhinovirus, coronavirus, metapneumovirus • Parvovirus B19 ○ Acquired in childhood • Hepatitis B virus (HBV) ○ Hepatitis B surface antigen-positive [HBsAg(+)] recipient ○ HBsAg(-) and hepatitis B core antibody-positive [HBcAb(+)] donor • Hepatitis C virus (HCV) ○ Underlying recipient disease

CLINICAL ISSUES Epidemiology • Incidence ○ Symptomatic CMV pneumonia has significantly decreased from 50% incidence in 1980s due to – Matching of CMV status of donor and recipient – Appropriate prophylaxis and viral load monitoring ○ HSV: Before era of prophylaxis, associated with significant morbidity and mortality – Currently rare in transplant setting ○ VZV reactivation in lung transplant: 12-15% ○ HCV seropositivity in potential lung transplant recipients: ~ 2%

○ Overall risk of EBV-associated PTLD (not necessarily pulmonary involvement) after lung transplant: 1% at 1 year and 1.5 % at 5 years • Factors rendering lung transplant patients more susceptible to infection ○ Immunosuppression ○ Altered phagocytosis ○ Graft denervation leads to absence of cough reflex and impaired mucociliary function ○ Compromised lymphatic flow

Lung Transplantation

TERMINOLOGY

Presentation • CMV and HSV: Fever, cough, dyspnea • RSV: Upper respiratory infection (cough, pharyngitis) or lower respiratory infection (tachypnea, wheezing) • Parvovirus B19: Unexplained and refractory anemia, pure red cell aplasia, low reticulocyte count • HBV and HCV: Hepatitis and cirrhosis • Adenovirus: Sudden-onset respiratory failure with upper respiratory manifestations (tonsillar/pharyngeal erythema)

Treatment • Drugs ○ CMV and HSV: Ganciclovir or valganciclovir – CMV may become resistant ○ HBV: Lamivudine, entecavir, adefovir ○ HCV: Ribavirin and Peginterferon-alfa ○ RSV: Aerosolized ribavirin ○ Adenovirus: Primarily supportive; cidofovir shown to improve survival in group of pediatric transplant patients ○ EBV-associated PTLD: Responds well to decrease in immunosuppression • Supportive care: Influenza, parainfluenza, rhinovirus, coronavirus, metapneumovirus • Monitoring ○ CMV viral load monitoring after drug therapy is important to detect early recurrence

Prognosis • Unclear whether CMV or RSV infection imparts increased risk for bronchiolitis obliterans syndrome

Risk Factors for Early Postoperative Infection • Suboptimal donor organ conditions (prolonged ischemia, arterial oxygen tension < 350 mm Hg) prior to harvest • Recipient age > 40 years • Prolonged ventilatory support • Copious tracheobronchial secretions

MACROSCOPIC General Features • CMV pneumonia ○ Heavy, consolidated lungs with nodular foci of necrosis and hemorrhage • HSV pneumonia ○ Airway-centered necrosis, airway ulceration with exudate and pseudomembrane • Adenovirus pneumonia ○ Bilateral necrotizing bronchocentric pneumonia ± acute hemorrhage • EBV-associated PTLD 403

Lung Transplantation

Viral Infections

404

○ Bilateral well-circumscribed, firm to rubbery, gray-tan to yellow necrotic nodules within lung parenchyma ○ Diffuse involvement of reticuloendothelial system ± liver, GI tract, kidneys, CNS

MICROSCOPIC Histologic Features • Pulmonary features characteristic of infections affecting lung transplant recipients described here • CMV pneumonia ○ Miliary necroinflammatory lesions – Necrotic inflamed alveolar walls with central small nodules of hemorrhage and fibrin – Zonal necrosis ○ Diffuse alveolar damage ○ Minimal nonspecific inflammation ○ Diffuse interstitial pneumonitis with alveolar wall inflammatory infiltrates ○ Characteristic cytopathic effects – Enlarged infected cells: Epithelial, endothelial, fibroblasts, macrophages – Single nuclear inclusion with peripheral halo ("owl's eye") – Small granular amphophilic cytoplasmic inclusions • HSV pneumonia ○ Miliary pattern of inflammation with necrosis ○ Diffuse alveolar damage ○ Airway ulceration ○ Characteristic cytopathic effects – Intranuclear amorphous inclusion with chromatin margination – Multinucleation with nuclear molding • RSV pneumonia ○ Bronchiolitis: Airways dilated with mucus, inflammatory cells, and necrotic epithelial debris – Chronic inflammation around bronchioles – Bronchiolar epithelial ulceration and necrosis – Characteristic cytopathic effects (multinucleation, cytoplasmic inclusions) are difficult to find • Adenovirus pneumonia ○ Diffuse alveolar damage ○ Bronchocentric necrosis with hemorrhage and neutrophilic/karyorrhectic debris ○ Characteristic cytopathic effects – Affects pneumocytes, bronchial epithelial cells, and histiocytes – "Smudgy" basophilic intranuclear inclusions • Pulmonary EBV-associated PTLD ○ Erosion/ulceration of cartilaginous bronchi ○ Lymphocytic vasculitis with fibrinoid necrosis ○ Sheets of cells with individual and focal coagulative necrosis ○ Range cytologically from immunoblastic cells, large lymphoid cells, and small round lymphocytes ○ Brisk mitoses • Due to immunosuppression, coinfection with multiple agents is not uncommon and must be considered in differential, as histologic (and radiologic and clinical) features often overlap

ANCILLARY TESTS Immunohistochemistry • For CMV, HSV, adenovirus, EBV

PCR • Quantitative detection of viral DNA in blood • Nucleic acid amplification test (NAAT) for communityacquired viruses using swab or aspirate

Genetic Testing • Hybrid capture CMV DNA assay: Quantitative • Nucleic acid sequence-based amplification for CMV RNA: Quantitative • CMV antigenemia assay for pp65: Semiquantitative

Culture • Conventional or shell vial: Low sensitivity and labor intensive

Serology • For HBV and HCV detection and monitoring

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Clinical correlation is essential as many histologic features of individual pulmonary viral infections overlap

SELECTED REFERENCES 1.

Almaghrabi RS et al: Cytomegalovirus infection in lung transplant recipients. Expert Rev Respir Med. 11(5):377-383, 2017 2. Green M et al: Epstein-Barr virus infection and posttransplant lymphoproliferative disorder. Am J Transplant. 13 Suppl 3:41-54; quiz 54, 2013 3. Sayah DM et al: Rhinovirus and other respiratory viruses exert different effects on lung allograft function that are not mediated through acute rejection. Clin Transplant. 27(1):E64-71, 2013 4. Uhlin M et al: Update on viral infections in lung transplantation. Curr Opin Pulm Med. 18(3):264-70, 2012 5. Weigt SS et al: CXCR3 chemokine ligands during respiratory viral infections predict lung allograft dysfunction. Am J Transplant. 12(2):477-84, 2012 6. Kwakkel-van Erp JM et al: Mannose-binding lectin deficiency linked to cytomegalovirus (CMV) reactivation and survival in lung transplantation. Clin Exp Immunol. 165(3):410-6, 2011 7. Paraskeva M et al: Cytomegalovirus replication within the lung allograft is associated with bronchiolitis obliterans syndrome. Am J Transplant. 11(10):2190-6, 2011 8. Vu DL et al: Respiratory viruses in lung transplant recipients: a critical review and pooled analysis of clinical studies. Am J Transplant. 11(5):1071-8, 2011 9. Kumar D et al: A prospective molecular surveillance study evaluating the clinical impact of community-acquired respiratory viruses in lung transplant recipients. Transplantation. 89(8):1028-33, 2010 10. Gottlieb J et al: Community-acquired respiratory viral infections in lung transplant recipients: a single season cohort study. Transplantation. 87(10):1530-7, 2009 11. Hakim FA et al: Severe adenovirus pneumonia in immunocompetent adults: a case report and review of the literature. Eur J Clin Microbiol Infect Dis. 27(2):153-8, 2008

Viral Infections

CMV Pneumonia (Left) Multiple inclusions diagnostic of cytomegalovirus (CMV) are present in this lung biopsy from a transplant recipient. The infected nuclei are enlarged ﬈. Note multiple granular cytoplasmic inclusions ﬊. (Right) This transbronchial biopsy from a lung transplant recipient shows perivascular inflammation extending into alveolar septa, which would be moderate acute rejection if not for the CMV inclusion ſt.

CMV (IHC)

Lung Transplantation

CMV Inclusions

Adenovirus Cytopathic Effect (Left) IHC stain for CMV in this lung biopsy is positive in nuclei of 2 only slightly enlarged cells, which on corresponding H&E did not show obvious inclusions. IHC stain is very helpful in such cases. (Right) Amidst an inflammatory fibrinous and hemorrhagic exudate are the "smudgy" basophilic intranuclear inclusions ﬈ characteristic of adenovirus.

Adenovirus on Frozen Section

Adenovirus (IHC) (Left) This high-power image, from an intraoperative consultation, shows cells with enlarged nuclei and poorly delineated "smudgy" nuclear inclusions ﬊ that should raise concern for adenovirus infection (frozen was reported as favor infection). Note the surrounding inflammatory exudate with karyorrhectic debris ﬈. (Right) Immunohistochemical stain for adenovirus highlights the irregular nuclear inclusions in sloughed epithelial cells and macrophages.

405

Lung Transplantation

Fungal Infections, Lung KEY FACTS

• Aspergillus may manifest as colonization or infection • High overall mortality rate (40-80%)

○ Cannot reliably distinguish from Aspergillus • Cryptococcus ○ Variable-sized yeast, 2-10 μm, with thick capsule

IMAGING

ANCILLARY TESTS

• Consolidation &/or pulmonary nodules

• Microbiologic culture does not conclusively indicate infection, because colonization can produce positive culture • In cases with positive histology, culture necessary for speciation • Gomori methenamine silver (GMS): Most sensitive • Periodic acid-Schiff (PAS): Does not stain pneumocystis

CLINICAL ISSUES

MICROSCOPIC • Aspergillus spp. ○ Septate hyphae with thick parallel walls and acute angle branching • Granulomas, necrotizing pneumonia, and angioinvasion can be seen in pulmonary aspergillosis ○ May be no tissue reaction in overwhelming infection • Candida ○ Budding yeasts with pseudohyphae ○ Granulomas with multinucleated giant cells and acute bronchopneumonia • Histologic features can suggest NAMF

TOP DIFFERENTIAL DIAGNOSES • Exogenous lipoid pneumonia • Pneumocystis pneumonia • Bacterial or other nonfungal pneumonia

Fungal Pneumonia

Anastomotic Aspergillus Infection

Anastomotic Aspergillus Infection

Aspergillus niger

(Left) Cut surface of the lung at autopsy shows multiple areas of consolidation st, some with cavitation ſt, and abscess formation in this transplant patient who died of fungal pneumonia. (Right) This lung transplant patient developed anastomotic infection and underwent endobronchial debridement. Both cartilage ﬈ and soft tissue ﬊ are necrotic. GMS and or PAS stain should be done.

(Left) GMS stain highlights invasive fungal hyphae in both the necrotic cartilage and soft tissue from the anastomotic site that was shown in the prior image. (Right) Fruiting head of Aspergillus niger with diagnostic pigmented conidiophores is shown in a transplant patient with an intrabronchial mass. Note that this is a rare finding on tissue sections.

406

Fungal Infections, Lung

Infectious Agents • Aspergillus: Most often Aspergillus fumigatus • Candida: Most often Candida albicans • Non-Aspergillus mycelial fungi (NAMF) ○ Zygomycetes: Rhizopus, Mucorales, Rhizomucor, Absidia ○ Fusarium spp., especially Fusarium solani ○ Scedosporium apiospermum or Scedosporium prolificans • Cryptococcus: Mostly Cryptococcus neoformansvar grubii (serotype A) or Crytpococcus neoformans var neoformans • Histoplasmosis: Histoplasma capsulatum

Immunosuppression • Necessary to prevent rejection ○ Allows growth of opportunistic and low virulence organisms

Routes of Infection • Preexisting colonization • Postsurgical hospital acquired • Community acquired

CLINICAL ISSUES Epidemiology • Incidence ○ Fungal infection occurs in 15-35% of lung recipients – 2.4% within 1st year post transplant – Majority occur after 1st year of transplant ○ 80% are Aspergillus spp. and Candida spp. – Invasive candidiasis rates have fallen recently • Risk factors for infection with Aspergillus ○ Cytomegalovirus infection ○ Chronic rejection ○ Cystic fibrosis ○ Additional risk factors in pediatric patients include – Grade A2 rejection – Multiple episodes of acute rejection – CMV-positive donor – Tacrolimus use • Risk factors for infection with Candida ○ Chronic glucocorticoids, broad-spectrum antibiotics, malnutrition • Risk factors for infection with NAMF ○ Scedosporium: Single lung transplant and cystic fibrosis ○ Zygomycetes: Broad-spectrum antibiotics, diabetic ketoacidosis, neutropenia

Presentation • Aspergillus may manifest in several different ways ○ Colonization: < 20% progress to invasive infection ○ Anastomotic site infection – Cough, dyspnea, hemoptysis – Necrosis and dehiscence of anastomosis can occur – Excessive granulation tissue can be obstructive ○ Invasive pulmonary infection ○ Disseminated infection, rare • Candida can present as colonization, mucocutaneous disease, or disseminated disease • NAMF can be localized or disseminated

• Cryptococcus can manifest as colonization, pneumonia, meningitis, or disseminated disease

Prognosis • • • •

High overall mortality rate (40-80%) Bronchial anastomosis infections have 23% mortality Disseminated infections have 90-100% mortality NAMF more likely to disseminate and have high mortality

Lung Transplantation

ETIOLOGY/PATHOGENESIS

MICROSCOPIC Histologic Features • Aspergillus spp. have septate hyphae with thick parallel walls and acute angle branching ○ Birefringent oxalate crystals may be present ○ Conidia (fruiting heads) rare within cavitary lesions where organism exposed to air ○ Granulomas, necrotizing pneumonia, and angioinvasion can be seen in lung • Candida spp. are budding yeasts with pseudohyphae ○ Granulomas with multinucleated giant cells and acute bronchopneumonia • Histologic features can suggest NAMF but cannot reliably distinguish from Aspergillus ○ Ribbon-like hyphae with nonparallel walls • Cryptococcus, variable-sized yeasts, 2-10 μm, thick capsule ○ Histiocytic pneumonia with numerous macrophages packed with yeast forms ○ Mucoid pneumonia with alveolar yeasts and little inflammatory reaction • Disseminated histoplasmosis has macrophages packed with small, oval H. capsulatum yeast

ANCILLARY TESTS Microbiological Culture • Necessary for speciation • Does not conclusively indicate infection, because colonization can produce positive culture

Histologic Stains for Fungi • Gomori methenamine silver (GMS): Most sensitive • Periodic acid-Schiff (PAS): Does not stain pneumocystis

DIFFERENTIAL DIAGNOSIS Exogenous Lipoid Pneumonia • Resembles cryptococcal pneumonia or disseminated histoplasmosis

Pneumocystis Pneumonia • Fluffy pink acellular alveolar exudates with sparse inflammation

Bacterial or Other Nonfungal Pneumonia • Fungal elements not identified

SELECTED REFERENCES 1. 2.

Pasupneti S et al: Aspergillus-related pulmonary diseases in lung transplantation. Med Mycol. 55(1):96-102, 2017 Danion F et al: Mucormycosis: New developments into a persistently devastating infection. Semin Respir Crit Care Med. 36(5):692-705, 2015

407

Lung Transplantation

Fungal Infections, Lung

Fungal Pneumonia

Aspergillus Infection

Oxalate Crystals in Aspergillosis

Oxalate Crystals in Aspergillosis

Aspergillus Pneumonia

Aspergillus Pneumonia

(Left) Intraoperative evaluation (frozen section) is very helpful in identifying fungal hyphae, which can be cut longitudinally ﬈ or seen on cross section ﬊ as rounded structures. Cutting the section at 6 μm and overstaining with hematoxylin makes the fungus easier to see. Giemsa stain can also highlight the hyphae. (Right) Presence of pale yellow oxalate crystals ﬈, as seen here within necrotic cartilage, is very helpful in suggesting a diagnosis of Aspergillus infection.

(Left) Polarized light microscopy shows this dramatic picture of strongly birefringent oxalate crystals in a transplant patient with invasive aspergillosis involving bronchial cartilage ﬇ and adjacent airway soft tissue. The size of the crystals is variable. (Right) High-power photomicrograph shows oxalate crystals on polarized light. Note the characteristic bright appearance, rounded pinwheel shape, and mostly yellow color (but with tinge of others).

(Left) Gomori methenamine silver (GMS) stain shows the characteristic radiating growth pattern of invasive Aspergillus. It is centered on a pulmonary artery branch and is growing out into the adjacent lung parenchyma. Note the dichotomous branching, which gives it the radial appearance. (Right) High-power view of GMS stain demonstrates acute angle branching and parallel walls of septate hyphae ﬇, which are straight. These are suggestive but not diagnostic of aspergillosis.

408

Fungal Infections, Lung

Candidiasis (Left) GMS stain shows broad, irregular hyphae of Rhizopus species. The walls are not parallel, and the hyphae appear twisted and ribbon-like compared with those of Aspergillus. (Right) Occasionally, granulomatous reaction can be seen (with or without necrosis), as in this patient with invasive candidiasis. Highly immunosuppressed patients would not form granulomas. Thus, special stains in such a case should always include silver and acid-fast stains.

Candidiasis

Lung Transplantation

Mucormycosis

Disseminated Histoplasmosis (Left) GMS stain highlights the pseudohyphae of Candida species. Note pinching in of fungal walls ſt formed by successive budding. (Right) Numerous small, oval yeasts (intracellular) of Histoplasma capsulatum are shown in a transplant patient with disseminated disease. The yeasts vary somewhat in size and are pointed at one end and rounded at the other (pear-shaped) ﬈. Occasional budding ﬉ is present.

Cryptococcal Pneumonia

Cryptococcal Pneumonia (Left) Disseminated cryptococcosis appears as vacuolated histiocytes, which could easily be mistaken for lipoid pneumonia. Each macrophage is packed with yeasts pushing the nucleus to one side (signet ring appearance). Note lack of neutrophilic inflammation. (Right) PAS stain highlights yeast walls but does not stain mucinous capsule. The yeasts are somewhat larger than Histoplasma but can be difficult to differentiate on morphology alone. Mucin stain is not always positive.

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SECTION 8

Intestinal Transplantation

Pathologic Classification of Intestinal Allograft Diseases Indications and Evaluation of Explant Reperfusion Injury History of Intestinal and Multivisceral Transplantation

412 414 418 422

Allograft Rejection/Immunological Injury Acute Antibody-Mediated Rejection, Intestine Acute Cellular Rejection, Intestine Chronic Rejection, Intestine Stomach Rejection Colon Rejection Graft-vs.-Host Disease, Intestine

424 428 436 440 444 448

Infections Bacterial and Fungal Infections Adenovirus, Intestine Rotavirus, Cytomegalovirus, and Herpes Simplex Virus Epstein-Barr Virus, Intestine

452 456 460 464

Intestinal Transplantation

Pathologic Classification of Intestinal Allograft Diseases

TERMINOLOGY Abbreviations • • • • •

Acute cellular rejection (ACR) Acute antibody-mediated rejection (AMR) Cytomegalovirus (CMV) Epstein-Barr virus (EBV) Posttransplant lymphoproliferative disorder (PTLD)

DEFINITIONS Reperfusion Injury • Ischemic injury to allograft bowel ○ Results from reperfusion of organs post transplantation • Donor factors responsible • Increases risk of subsequent rejection episodes

Acute Cellular Rejection • T-cell-mediated injury to bowel ○ Occurs due to large antigenic load of donor bowel • Alloimmune reaction of T cells to donor antigens ○ Predominantly those of major histocompatibility complex (HLA) • Other cells may also play role, such as ○ Macrophages ○ Plasma cells

Acute Antibody-Mediated Rejection • Alloantibody-mediated injury to cells expressing donor antigens ○ Predominantly HLA classes I and II antigens ○ Frequently IgG lymphocytotoxic antibodies ○ Preformed donor-specific antibodies predispose to AMR

Enteritis • Any inflammatory process involving allograft or native bowel due to infection • Usually induces influx of inflammatory cells ○ Mainly neutrophils and lymphocytes ○ Causative agents may or may not be seen on biopsy

Chronic Rejection • End result of prolonged episodes of rejection ○ Either cell mediated or due to antibody-mediated injury • Usually end result of vascular injury resulting in graft vascular insufficiency

Types of Intestinal Transplants • Isolated small bowel transplant ○ Usually with anastomosis with native jejunum and colon • Combined small bowel and colon transplant ± liver ○ Usually anastomosis with jejunum and rectum/rectosigmoid colon • Multivisceral transplant ○ Combinations of small bowel, colon, stomach, duodenum, pancreas, and liver ○ Modified multivisceral transplant with exclusion of liver ○ Combined liver and intestine with pancreas

ALLOIMMUNE RESPONSE Cell-Mediated Rejection • ACR 412

○ Usually with increased lamina propria cellularity with lymphocytes and some plasma cells ○ Crypt injury manifested by crypt epithelial apoptosis required ○ Ulceration frequent in severe injury ○ Categorize into following grades – Indeterminate – Mild – Moderate – Severe ACR ○ If persistent, results in exfoliative rejection ○ Rejection may be patchy in biopsies and between sites • Chronic rejection ○ Can manifest as chronic nonhealing ulcerated mucosa ○ Difficult to diagnose on mucosal biopsies – May need to examine serial biopsies ○ Clues include persistent architectural changes and lamina propria fibrosis ○ Fibrosing peritonitis may be manifestation (on resected grafts only) ○ Characteristic mesenteric and penetrating vascular intimal injury with inflammation and marked narrowing of lumen ○ Vascular changes seen on resection specimens only – Need to carefully examine any segmental resection specimens for vascular changes • Antibody-mediated rejection ○ Acute injury – Within days to weeks ○ Capillary dilatation, endothelial activation, and intravascular neutrophils ○ Mucosal ulceration in severe cases ○ C4d shows diffuse deposition within capillaries in lamina propria ○ No consistent correlation between C4d and AMR episodes ○ Circulating donor-specific antibodies demonstrated • Graft-vs.-host disease ○ Large population of donor lymphoid tissue predisposes to graft-vs.-host disease ○ Variable presentation with multiorgan involvement; fatal ○ Native organs affected ○ Allograft uninvolved

NONALLOIMMUNE DISEASES Reperfusion Injury • Immediate injury within 1st few hours of transplant • Characterized by ○ Mucosal hemorrhages ○ Crypt injury ○ Ischemic necrosis with neutrophils

Bacterial Infections • Usually enteric bacteria most common • Clostridium difficile in children • Neutrophils present

Fungal Infections • Usually result of severe immune suppression • Early postsurgical wound infections due to Candida

Pathologic Classification of Intestinal Allograft Diseases 5.

• Adenovirus ○ Early ○ More often pediatric • CMV • Herpesvirus • Rotavirus • Calicivirus • Inclusions visualized in allograft biopsies for adenovirus and CMV ○ Rarely herpes • EBV ○ Early infection in pediatric patients ○ Reactivation in adults ○ EBER stain helpful in assessing degree of viral colonization

6.

Posttransplant Tumors

13.

• • • •

14.

Typically lymphoproliferative disorders Frequently affects intestinal graft Polymorphous or monomorphic Most EBV related ○ Some not associated with EBV ○ Some T-cell PTLD • Can be incidental finding in allograft surgical specimens

7.

8.

9. 10.

11. 12.

15.

Mangus RS et al: Multivisceral Transplantation: expanding Indications and improving outcomes. J Gastrointest Surg. 17(1):179-86, 2012 Remotti H et al: Small-bowel allograft biopsies in the management of smallintestinal and multivisceral transplant recipients: histopathologic review and clinical correlations. Arch Pathol Lab Med. 136(7):761-71, 2012 Tsai HL et al: Association between donor-specific antibodies and acute rejection and resolution in small bowel and multivisceral transplantation. Transplantation. 92(6):709-15, 2011 Gupta A et al: Elevated myeloid: plasmacytoid dendritic cell ratio associates with early acute cellular rejection in pediatric small bowel transplantation. Transplantation. 89(1):55-60, 2010 Vianna RM et al: Current status of small bowel and multivisceral transplantation. Adv Surg. 42:129-50, 2008 Ruiz P et al: Histological criteria for the identification of acute cellular rejection in human small bowel allografts: results of the pathology workshop at the VIII International Small Bowel Transplant Symposium. Transplant Proc. 36(2):335-7, 2004 Klaus A et al: Diffuse mesenterial sclerosis: a characteristic feature of chronic small-bowel allograft rejection. Virchows Arch. 442(1):48-55, 2003 Parizhskaya M et al: Chronic rejection of small bowel grafts: pediatric and adult study of risk factors and morphologic progression. Pediatr Dev Pathol. 6(3):240-50, 2003 Adams DH: Immunologic aspects of small bowel transplantation. Transplant Proc. 30(6):2557-9, 1998 Mueller AR et al: Differentiation between preservation reperfusion injury and acute rejection after small bowel transplantation. Transplant Proc. 30(6):2657-9, 1998 Banner B et al: Transplantation of the small intestine: the pathologist's perspective. Am J Surg Pathol. 14 Suppl 1:109-16, 1990

Intestinal Transplantation

Viral Infections

MULTIVISCERAL TRANSPLANT ISSUES Gastric and Colonic Rejection • May or may not occur in conjunction with small bowel rejection • Gland or crypt apoptosis characteristic • Lesser degree of apoptoses than in small bowel allograft biopsies • Liver/small bowel combined transplants have better outcome than isolated small bowel transplants

RECURRENT INTESTINAL DISEASES Causes • Crohn disease • Rare abdominal tumors (e.g., desmoids)

RETRANSPLANTATION Causes of Graft Loss • • • •

Severe ACR, including exfoliative rejection Chronic rejection or sclerosing mesenteritis PTLD Adhesions and graft dysmotility

SELECTED REFERENCES 1. 2.

3.

4.

Bharadwaj S et al: Current status of intestinal and multivisceral transplantation. Gastroenterol Rep (Oxf). 5(1):20-28, 2017 Liu L et al: Sequential histologic changes in the healing process in small bowel allografts treated for acute cellular rejection. Transplant Proc. 45(2):643-8, 2013 Swanson BJ et al: Histologic analysis of chronic rejection in small bowel transplantation: mucosal and vascular alterations. Transplantation. 95(2):378-82, 2013 Tsuruyama T et al: Histology of intestinal allografts: lymphocyte apoptosis and phagocytosis of lymphocytic apoptotic bodies are diagnostic findings of acute rejection in addition to crypt apoptosis. Am J Surg Pathol. 37(2):17884, 2013

413

Intestinal Transplantation

Indications and Evaluation of Explant

TERMINOLOGY Definitions • Intestinal transplantation (ITx) ○ Includes isolated small intestine or both small and large intestines • Liver and intestinal transplantation (L-ITx) • Multivisceral transplantation (MVT) ○ L-ITx and stomach ○ Duodenum ○ Pancreas ○ Spleen • Modified multivisceral transplantation (MMVT) ○ MVT with some exclusions

○ ○

EPIDEMIOLOGY Age Range • < 1 year to > 65 years

Incidence • Organ Procurement and Transplantation Network (OPTN) database • Source ○ 1990-2012 – Total intestinal transplants = 2,232 ○ Peak incidence in 1-5 year age group – 60% of transplants in pediatric age group – 40% in adults ○ Slight progressive decrease in number of intestinal transplants in last 3 years across all age groups

○

ETIOLOGY/PATHOGENESIS Indications • Pediatric: Isolated ITx or L-ITx ○ Gastroschisis – Congenital defect in abdominal wall □ Results in persistence of intestinal organs within large extraabdominal sac – May include

Tufting Enteropathy (Left) This small bowel biopsy shows subtle changes of the surface epithelium with occasional epithelial cells showing "piling up" (tufting enteropathy) ﬈. (Right) A case of tufting enteropathy stained for MOC-31 shows complete loss of membranous staining in the surface epithelium ﬈ as a result of a genetic defect involving EPCAM.

414

○

□ Liver □ Stomach □ Intestines □ Pancreas □ Spleen – Narrow opening of abdominal wall results in □ Frequent kinks □ Ischemic bowel injury □ Requires surgical resection of intestines – Usually very little residual bowel for removal at time of transplantation □ Show serosal adhesions □ May be normal Volvulus Necrotizing enterocolitis – Frequent cause of short gut syndrome (SGS) □ In children who survive acute phase of this condition – Common in premature infants □ Especially severely premature babies in 1st few weeks after birth – Much less common in full-term babies – May be related to inflammatory mediators □ Following bacterial colonization with possible ischemic component – Neonates often require multiple resections to remove large portions of bowel, resulting in SGS – Explants show serosal adhesions and some fibrosis of wall – Active disease never seen at time of transplantation Intestinal atresia – Typically when multiple resections for multifocal atresias result in SGS – Underlying cause may be cystic fibrosis □ Testing for cystic fibrosis recommended – Can involve small intestine or both small and large intestine Hirschsprung disease (HSCR) – Only long-segment variant associated with SGS

Tufting Enteropathy: MOC31 Stain

Indications and Evaluation of Explant

○

○

○

○

○

– Chronic rejection • Pediatric: MVT or MMVT ○ HSCR involving stomach ○ Pseudoobstruction involving stomach ○ Familial adenomatous polyposis ○ Crohn disease – Unusual in pediatric age group – Usually in adulthood • Adults: Isolated ITx or L-ITx ○ Crohn disease – 2nd most common cause of transplantation in adults □ 14% of total – Multiple bowel resections result in SGS and dependence on TPN □ Necessitates ITx – Can recur in allograft ○ Tumors – Epithelial tumors involving root of mesentery – Pancreatic tumors – Desmoids – Metastatic neuroendocrine tumors ○ Pseudoobstruction/motility disorders ○ Ischemic bowel disease most common cause ○ Familial adenomatous polyposis/Gardner syndrome and desmoids ○ Trauma ○ Retransplantation due to similar causes as pediatric retransplants

Intestinal Transplantation

○

– Aganglionosis extends from rectum to small intestine to varying extent □ May involve jejunum – Most have stoma placement as treatment for aganglionosis – At time of transplant, portions of aganglionic bowel may still be present in explant □ If resection was not performed earlier □ Rare frozen section evaluation needed to determine ganglionic bowel to use for anastomosis/stoma Pseudoobstruction – Includes diverse group of lesions, such as □ Mitochondrial diseases □ Neuromuscular diseases □ Some hollow visceral myopathies – Can present from early in infancy and mimic HSCR □ Requires rectal biopsies to exclude HSCR – Invariably results in SGS with dependence on total parenteral nutrition (TPN) Microvillus inclusion disease – Cause of intractable diarrhea of infancy – Usually presents in early infancy with failure to thrive and secretory diarrhea – Histological diagnostic hallmark is absence of brush border in small intestine □ Demonstrated by PAS, CD10, or alkaline phosphatase – MYO5B gene mutation recently identified – Some children show liver insufficiency due to canalicular brush border defects – ITx or MVT only current options for treatment Tufting enteropathy – Manifests as intractable diarrhea of infancy □ Secretory diarrhea □ Failure to thrive – Histological diagnosis by demonstration of epithelial tufts on surface of small and large intestinal mucosa – EPCAM gene mutations □ Identified as cause of disease – Requires ITx including colon □ Often MVT Familial adenomatous polyposis – Rare cause of MVT or ITx – Usually following prophylactic or therapeutic ileocolectomy Tumors: Desmoid fibromatosis with adhesions – Rare cause of ITx – Typically in intraabdominal tumors that involve mesentery □ Cause marked adhesions of bowel and intestinal obstruction – Usually in setting of familial adenomatous polyposis ± intestinal polyps Retransplantation due to – Hyperacute or severe antibody-mediated rejection – Exfoliative rejection – Ischemic/vascular thrombosis – Posttransplant lymphoproliferative tumors □ Involving graft and mesentery

Contraindications to Intestinal Transplantation • Pediatric and adult ○ Profound neurologic abnormalities and multisystem diseases ○ Life-threatening or noncorrectable illness of gastrointestinal tract ○ Severe congenital or acquired immunological disorders ○ Nonresectable malignancies

CLINICAL IMPLICATIONS Graft Survival Following Intestinal Transplantation • Short-term graft and patient survival have significantly improved (80-90%) ○ Survival for L-ITx and MVT are lower (~ 70%) • Improved immunomodulation resulted in significantly improved survival • Long-term survival still low ○ From 1997-2006, 5- and 10-year patient survival only 54% and 43%, respectively, with lower graft survival rates

Causes of Graft Failure • Chronic allograft rejection • Vascular compromise (either acute or chronic) leading to graft failure • Severe exfoliative rejection • Posttransplant tumors necessitate withdrawal of immunosuppression over prolonged periods with graft survival affected by rejection • Severe systemic infections preclude use of immunosuppressive agents for graft survival 415

Intestinal Transplantation

Indications and Evaluation of Explant • Some causes may be indications for retransplantation ○ Others may prove fatal

MACROSCOPIC Evaluation of Explant • May be routine procedure of stoma closure after intestinal transplant or resection of entire allograft • Bowel segments resected at time of stoma closure are easy to evaluate ○ Grossly assess for nodules or ulcers ○ Adequately section bowel mucosa and serosal vessels as well as mesentery ○ Incidental findings – Ulcers – Nodules of small posttransplant tumors – Arterial intimal changes of chronic rejection • Evaluation of resected allograft ○ Can be complex due to multiple adhesions and lack of proper mesenteric attachments ○ Important to identify anastomotic points ○ Look for any areas of discoloration – Sample both wall and mesentery at these sites ○ Look for nodules (incidental or expected) regardless of resection indication ○ Triage tumor specimens for any further studies or cytogenetics if indicated ○ Look for evidence of thickened mesenteric vessels or thrombosis ○ Submit sections longitudinally to assess vessels penetrating muscle layers for changes of chronic rejection ○ Adequately sample bowel mucosa (both ulcers and adjacent mucosa) to evaluate for rejection ○ Identify and evaluate any separate bowel fragments included for changes and submit sections accordingly ○ Judiciously sample and submit sections for evaluation

MICROSCOPIC Stomal Changes • Evaluate for ○ Evidence of rejection ○ Vascular changes of chronic rejection in subserosa and serosa ○ Subtle signs of early chronic rejection – Neuronal hyperplasia – Pyloric metaplasia ○ Evidence of viral infection/inclusions ○ Evidence of posttransplant tumors • Use EBER staining where indicated to exclude any EBVdriven process • Compare histology with prior biopsies to better understand changes

Explant Changes • Evaluate for ○ Acute cellular rejection ○ Evidence of exfoliative rejection in mucosa ○ Penetrating and serosal/mesenteric vessels – Intimal proliferation – Changes of chronic rejection 416

○ Serosa – Sclerosing mesenteritis (form of chronic rejection) – Fibrosis ○ Tumors and nodules for any PTLD ○ Infectious agents ○ Severe rejection of resection margins – If segmental resection of allograft performed • Immunostain for C4d when suspicious of antibodymediated rejection as cause of graft failure

SELECTED REFERENCES 1. 2. 3.

4.

5.

6. 7. 8. 9. 10.

11. 12. 13. 14. 15.

Mangus RS et al: Multivisceral transplantation: expanding indications and improving outcomes. J Gastrointest Surg. 17(1):179-87, 2013 Tzakis AG et al: Intestinal and multivisceral autotransplantation for tumors of the root of the mesentery: long-term follow-up. Surgery. 152(1):82-9, 2012 Cruz RJ Jr et al: Modified multivisceral transplantation with spleenpreserving pancreaticoduodenectomy for patients with familial adenomatous polyposis "Gardner's Syndrome". Transplantation. 91(12):1417-23, 2011 Mazariegos GV et al: Current status of pediatric intestinal failure, rehabilitation, and transplantation: summary of a colloquium. Transplantation. 92(11):1173-80, 2011 Cruz RJ Jr et al: Modified "liver-sparing" multivisceral transplant with preserved native spleen, pancreas, and duodenum: technique and long-term outcome. J Gastrointest Surg. 14(11):1709-21, 2010 Millar AJ et al: Intestinal transplantation for motility disorders. Semin Pediatr Surg. 18(4):258-62, 2009 Kato T et al: Intestinal and multivisceral transplantation in children. Ann Surg. 243(6):756-64; discussion 764-6, 2006 Grant D et al: 2003 report of the intestine transplant registry: a new era has dawned. Ann Surg. 241(4):607-13, 2005 Bond GJ et al: Intestinal transplantation for total/near-total aganglionosis and intestinal pseudo-obstruction. Semin Pediatr Surg. 13(4):286-92, 2004 Parizhskaya M et al: Chronic rejection of small bowel grafts: pediatric and adult study of risk factors and morphologic progression. Pediatr Dev Pathol. 6(3):240-50, 2003 Tzakis AG et al: Intestinal transplantation: advances in immunosuppression and surgical techniques. Transplant Proc. 35(5):1925-6, 2003 Wu T et al: A schema for histologic grading of small intestine allograft acute rejection. Transplantation. 75(8):1241-8, 2003 Nishida S et al: Ninety-five cases of intestinal transplantation at the University of Miami. J Gastrointest Surg. 6(2):233-9, 2002 Noguchi Si S et al: Pediatric intestinal transplantation: the resected allograft. Pediatr Dev Pathol. 5(1):3-21, 2002 Starzl TE et al: Multivisceral and intestinal transplantation. Transplant Proc. 24(3):1217-23, 1992

Indications and Evaluation of Explant

Microvillus Inclusion Disease (Left) The same allograft bowel resection cut surface shows extensive congestion and hemorrhages as well as wall thickening and adhesions between bowel loops. Some thickened arteries ﬈ are noted in the mesentery. (Right) PAS stain shows an absence of brush border in this case of microvillus inclusion disease ﬈. Note instead the pink zone of staining in the enterocyte cytoplasm at the apex of the cells suggesting lysosomal accumulation.

Microvillus Inclusion Disease

Intestinal Transplantation

Resected Allograft: Chronic Rejection

Microvillus Inclusion Disease (Left) High-magnification image of an microvillus inclusion disease (MID) case shows absence of the brush border on the surface ﬈ with reactive-appearing epithelial cells and no significant inflammation. (Right) CD10 immunostain of an MID case highlights the absence of the brush border on the surface of small intestinal epithelial cells ﬈ with a corresponding superficial enterocyte cytoplasmic staining similar to the PAS stain.

Hirschsprung Disease

Hirschsprung Disease (Left) Low-magnification image of the small bowel shows close apposition of the 2 muscle layers ﬈ without development of a myenteric plexus in an example of longsegment Hirschsprung disease involving the entire small intestine. A hypertrophic nerve trunk may not always be evident. (Right) A section of colon shows an absence of development of myenteric plexus and no separation between the muscle layers ﬈ in an example of total colonic aganglionosis.

417

Intestinal Transplantation

Reperfusion Injury KEY FACTS

• Preservation injury

• Associated with increased risk of acute cellular rejection in graft

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• May be related to increased inflammatory mediators in cadaveric donor organ ○ Presently unknown • Possible association with length of preservation of donor organ in cold preservatives • Preliminary data suggest upregulation of inflammatory genes in blood of recipients in immediate posttransplant period

• • • • • • •

TERMINOLOGY

CLINICAL ISSUES • Occurs immediately or up to 1 week post transplant • Intestinal dysmotility • Inapparent clinically ○ Diagnosis made on histology

Epithelial lifting Edema of villi Villous blunting Surface denudation Crypt epithelial regeneration and mitoses Few crypt apoptoses in severe ischemic injury Severe cases with neutrophilic and monocytic infiltration within graft • More severe cases with frank ulceration and hemorrhage

TOP DIFFERENTIAL DIAGNOSES • Antibody-mediated rejection • Acute cellular rejection • Bacterial/viral infections

Mild Ischemia/Reperfusion Injury

Severe Ischemia/Reperfusion Injury

Ischemia/Reperfusion Injury

Ischemia/Reperfusion Injury: Day 3

(Left) An example of an immediate postperfusion biopsy shows mild reactive changes of the surface epithelium and crypts with villous blunting and mild mucosal congestion. (Right) Severe reperfusion injury results in epithelial denudation, mucosal hemorrhage and loss of epithelium, and crypts with reactive stromal changes in lamina propria.

(Left) A scanning view of a small bowel biopsy taken at day 3 shows marked regenerative mucosal changes with hemorrhages, some loss of crypts, and stromal changes within lamina propria. (Right) Higher magnification image shows crypt injury, hemorrhages, regenerative epithelium, stromal changes, and some architectural distortion with basal separation of crypts

418

Reperfusion Injury

Abbreviations • Ischemia/reperfusion injury (IRI)

Synonyms • Preservation injury

Definitions • Ischemic graft injury in immediate posttransfusion period from vascular perfusion following cold preservation that results in inflammatory response

ETIOLOGY/PATHOGENESIS Donor Organ Status and Preservation • May be related to increased inflammatory mediators in deceased donor organ • Prior inflammatory conditions in bowel before donor death • Possible association with length of preservation of donor organ in cold preservatives

At Time of Surgery • Surgical manipulation releases inflammatory mediators • Preliminary data suggest upregulation of inflammatory genes in recipients in immediate posttransplant period ○ Inflammatory mediators, such as T-bet and TLR9 • Oxygen free radicals also implicated in injury • Post cold preservation, revascularization of graft causes release of inflammatory mediators • Increase intragraft CD14(+) monocytes in some reperfused bowel transplants

CLINICAL ISSUES Presentation • Occurs immediately or up to 1 week post transplant • Intestinal dysmotility • Infections and fevers related to bacterial translocation and sepsis • Rare instances of immediate graft congestion and hemorrhages with discoloration at end of surgery • Inapparent clinically, but diagnosis made by biopsy

Treatment • Preconditioning regimens implemented to prevent injury • Possible role of glycine treatment in recipients to inhibit inflammatory mediators • Recent oxygen insufflation of University of Wisconsin solution reduces reperfusion injury

Prognosis • Usually good outcome with resolution within 1 to 2 weeks • Associated with increased risk of acute cellular rejection

IMAGING

• • • • • •

Villous blunting and surface denudation Congestion of capillaries in lamina propria Crypt epithelial regeneration and mitoses Smooth muscle contraction and some eosinophilia No significant inflammatory cell increase in lamina propria Severe cases ○ Frank ulceration and hemorrhage ○ Ischemic injury extends transmurally ○ Few crypt apoptoses in severe ischemic injury ○ Neutrophilic and monocytic infiltration within graft ○ Attenuated mucosa with loss of crypts • Features of recovery ○ Regeneration usually evident at days 5-7 ○ Usually subsides by day 14 ○ May be associated with acute cellular rejection on day 7 or 14 biopsy ○ Villous height may take longer to return to normal ○ Reactive stromal increase may be only evidence of undetected reperfusion injury

Intestinal Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Antibody-Mediated Rejection • Occurs often in 1st week but can occur later • Endoscopically, mucosa congested or denuded • Dilated mucosal capillaries with plump endothelial cells and neutrophilic margination (not usually seen in IRI) • Strongly positive C4d in capillary endothelial cells for antibody-mediated rejection

Acute Cellular Rejection • Can occur in 1st week • Crypt apoptosis prominent • Ulceration of severe acute cellular rejection associated with crypt apoptosis and lymphocytic infiltrates • Needs distinction from IRI due to different treatments • Villous alterations may be common to both

Bacterial/Viral Infections • Rarely occurs, most likely bacterial in 1st week • Usually associated with extensive neutrophilic infiltrates and exudates with ulcers • No hemorrhage or crypt regeneration • No apoptosis in crypts • Pseudomembrane in cases of Clostridium difficile enteritis

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Mucosal congestion and stromal prominence in biopsies within 7 days of transplantation • Mucosal sloughing and crypt regeneration in more severe cases without crypt apoptosis • Frank hemorrhage and sloughing in severe cases; need to exclude AMR

Radiographic Findings • Evidence of ileus with dilatation of bowel loops

SELECTED REFERENCES 1.

MICROSCOPIC Histologic Features • Epithelial lifting and edema of villi

2.

Glowka TR et al: Oxygen Insufflation in University of Wisconsin Solution Ameliorates Reperfusion Injury in Small Bowel after Cold Storage and Reperfusion. Ann Transplant. 20:469-77, 2015 Lee RG et al: Pathology of human intestinal transplantation. Gastroenterology. 110(6):1820-34, 1996

419

Intestinal Transplantation

Reperfusion Injury

Mild Ischemia/Reperfusion Injury

Mild Ischemia/Reperfusion Injury

Ischemia/Reperfusion Injury

Severe Ischemia/Reperfusion Injury

Severe Ischemia/Reperfusion Injury

Severe Ischemia/Reperfusion Injury

(Left) Low-power image shows villous alteration with superficial hemorrhage and marked reactive changes in epithelium and crypts in a day 0 postperfusion bowel specimen. The crypts appear normal in distribution but are elongated. (Right) Highmagnification image of a postperfusion specimen shows flattening of villi and hyperchromasia of epithelial cell nuclei with some loss of goblet cells in the surface epithelium, suggestive of regenerative changes ﬈.

(Left) High-magnification image of a postperfusion specimen shows crypt regenerative changes, a rare apoptotic cell ſt within crypts, and minimal lamina propria inflammation. (Right) This example of severe ischemic injury in a postperfusion specimen resulted in poor graft function in the immediate posttransplant period. Note the mucosal hemorrhage, loss of epithelium and crypts, and congestion.

(Left) Close-up view of a postperfusion specimen shows epithelial sloughing and loss of crypts with a single residual crypt ﬈ showing epithelial injury and inflammatory infiltrate in the lumen. Congestion and hemorrhage are also noted. (Right) Highmagnification image highlights epithelial denudation, loss of crypts, and residual crypts with necrotic debris ﬈ and sloughed cells within the lumina. The lamina propria is congested. No apoptoses or viral inclusions are noted.

420

Reperfusion Injury

Differential Diagnosis: AMR (Left) Biopsy shows denudation of epithelium with dilated capillaries and a granulation tissue appearance of the lamina propria with loss of crypts. This raises the possibility of acute cellular rejection (ACR) or antibodymediated rejection (AMR) rather than ischemic/ reperfusion injury. (Right) Another section shows dilated capillaries in the lamina propria that are filled with inflammatory cells, including neutrophils ﬈, and lined by prominent endothelial cells. This should warrant a C4d immunostain to exclude AMR.

Differential Diagnosis: ACR

Intestinal Transplantation

Differential Diagnosis: ACR/AMR

Differential Diagnosis: ACR (Left) This example shows epithelial denudation, but the crypts reveal multiple apoptoses ﬈ that would favor the diagnosis of ACR in this biopsy. ACR may coexist with ischemia/reperfusion injury. (Right) Photomicrograph demonstrates marked crypt injury, and multiple crypts reveal a blown-out appearance with ring apoptosis of crypts ﬈ in this example of severe ACR. Note also the increased lymphocytic infiltration in the lamina propria.

Differential Diagnosis: AMR

Differential Diagnosis: AMR (Left) C4d immunohistochemistry in an ulcerated small bowel allograft biopsy reveals, even at this low magnification, diffuse positive capillary endothelial staining ﬈, raising a strong suspicion for AMR. This needs correlation with circulating donor-specific antibodies. (Right) Highmagnification image shows strong endothelial staining of C4d with a granular cytoplasmic staining pattern. The surface epithelium of this plump villous structure is denuded.

421

Intestinal Transplantation

History of Intestinal and Multivisceral Transplantation

TERMINOLOGY Abbreviations

•

• Intestinal transplantation (ITx) • Multivisceral transplantation (MVT)

Types of Transplants • Isolated small bowel or intestinal • Intestinal and liver (most common type) • ITx as part of MVT that includes colon, stomach, and duodenum, or liver and pancreas in various combinations

CHRONOLOGY AND EVOLUTION

•

•

•

Timeline • 1902 ○ Carrel – 1st attempt at canine ITx • 1959 ○ Lillehei et al – Technique for isolated ITx • 1960 ○ Starzl and Kaupp – 1st successful MVT in dogs • 1962 ○ Recognition that ITx as part of MVT has better outcome than ITx alone • 1962 ○ Lillehei – Autotransplantation of cold-preserved small bowel • 1968 ○ Dudrick et al – Introduction of total parenteral nutrition (TAN) • 1972 ○ Fortner et al – Longest survival: 76 days for ITx in precyclosporin era ○ Azathioprine, steroids, and antilymphocyte globulin (ALG) used as immunosuppressive agents • 1986 ○ Cohen et al – 1st transplant at University of Toronto using cyclosporine • 1987 ○ Starzl et al – 1st MVT in child at Pittsburgh, survived > 6 months • 1987 ○ Grant et al – 1st combined liver-intestine transplantation in London • 1986-1990 ○ 13 intestinal transplants worldwide with 15% success rate • 1988 ○ Deltz et al – Isolated intestinal transplant from live donor – Nutritional function for 56 months • 1989 ○ FK506 (tacrolimus or Prograf) introduced to clinical transplantation • 1990 ○ Starzl 422

•

•

•

•

•

– 1st clinical trial in Pittsburgh using FK506 – Dramatic improvement in success rate 1995 ○ White et al – Pathology of pediatric small ITx 1996 ○ Lee et al – Grading system for small intestine allograft rejection 1996 ○ Fujisaki et al – Description of pathology of antibody-mediated rejection in small bowel transplant in rats 2004 ○ Garcia et al – Grading acute cellular rejection in gastric allografts 2006 ○ Wu et al – Pathology in patients pretreated with Thymoglobulin or alemtuzumab 2009 ○ Abu-Elmagd et al – Reported experience with 500 ITx and MVT at single center 2009 ○ Sindhi et al – Demonstration of CD15(+) T-cytotoxic cells as markers of ACR in children with small bowel transplantation 2015 ○ Grant et al – Human and mouse tissue-engineered small intestine both demonstrate digestive and absorptive function □ Attempt to generate organoids for short bowel syndrome and substitute for transplantation 2016 ○ Sindhi et al – Profile of Pleximmune blood test for transplant rejection risk prediction

Other Historically Significant Transplant Landmarks • 1943 ○ Medawar et al – Tissue rejection is immune reaction • 1953 ○ Billingham et al – Tolerance to skin allografts induced by inoculating mice with immunocompetent spleen from adult donors • 1958 ○ Dausset – 1980 Nobel Prize for discovery of 1st HLA • 1959 ○ Merrill, Murray – 1st kidney transplantation in twins with long survival • 1959 ○ Billingham – Definition of 3 prerequisites for development of graftvs.-host disease • 1960 ○ Goodwin

History of Intestinal and Multivisceral Transplantation

Year

Landmark Event

Pioneer

1902

1st attempt at canine intestinal transplantation

Carrel

1959

Developed technique for isolated intestinal transplantation

Lillehei

1987

1st multivisceral transplant in child

Starzl et al

1990

1st clinical trial using tacrolimus (FK506) in intestinal transplants; improved survival

Starzl

1995

Description of pathology of pediatric small intestine transplantation

White et al

1996

Grading of small intestinal transplant rejection

Lee et al

1996

Description of pathology of antibody-mediated rejection in small bowel transplant in rats

Fujisaki et al

2004

ACR grading scheme for gastric allografts

Garcia et al

2004

Pathology of intestinal transplant with preformed antibodies

Wu et al

2006

Description of histology in patients pretreated with Thymoglobulin or alemtuzumab

Wu et al

2010

Identification of allospecific CD154(+) B cells in intestinal allograft rejection

Ashokkumar et al

•

•

•

•

•

• •

•

•

• •

•

•

– Methotrexate and cyclophosphamide for immune suppression in renal transplant 1960 ○ Calne et al – Preclinical studies of 6-mercaptopurine and azathioprine 1963 ○ Mathe et al – Success with bone marrow transplantation 1963 ○ Starzl et al – Reversal of rejection using prednisone in addition to azathioprine 1963 ○ Gowan – 1st description of function of lymphocytes 1964 ○ Starzl et al – Liver described as most tolerogenic of organs 1966 ○ ALG trials begin 1967 ○ Starzl – 1st liver transplantation 1967 ○ Starzl – Seminal paper "Death after transplantation" 1968 ○ Barnard – 1st cardiac transplantation 1968 ○ "Brain death" defined by Harvard Commission 1976 ○ Dreyfuss et al – Discovery of cyclosporin A and cyclosporin C 1977 ○ Kostakis et al and Calne et al – Use of cyclosporin in animal transplants 1978-79 ○ Calne et al – Use of cyclosporin in human transplants

Intestinal Transplantation

Timeline of Significant Events in ITx and MVT

• 1986 ○ United Network for Organ Sharing starts keeping records • 1988 ○ Belzer et al – University of Wisconsin preservation solution • 1992-93 ○ Starzl et al – Chimerism in organ recipients • 2014 ○ FDA approves Pleximmune test for prediction of acute cellular rejection for liver and small bowel transplants in children

SELECTED REFERENCES 1. 2. 3.

4. 5.

6. 7.

8.

9. 10.

11. 12. 13.

14.

Brenner DA: Thomas E. Starzl: Transplantation pioneer. Proc Natl Acad Sci U S A. 114(41):10808-10809, 2017 Sindhi R et al: Profile of the Pleximmune blood test for transplant rejection risk prediction. Expert Rev Mol Diagn. 16(4):387-93, 2016 Grant CN et al: Human and mouse tissue-engineered small intestine both demonstrate digestive and absorptive function. Am J Physiol Gastrointest Liver Physiol. 308(8):G664-77, 2015 Ruiz P: Updates on acute and chronic rejection in small bowel and multivisceral allografts. Curr Opin Organ Transplant. 19(3):293-302, 2014 Ashokkumar C et al: Increased expression of peripheral blood leukocyte genes implicate CD14+ tissue macrophages in cellular intestine allograft rejection. Am J Pathol. 179(4):1929-38, 2011 Nayyar N et al: Pediatric small bowel transplantation. Semin Pediatr Surg. 19(1):68-77, 2010 Abu-Elmagd KM et al: Evolution of the immunosuppressive strategies for the intestinal and multivisceral recipients with special reference to allograft immunity and achievement of partial tolerance. Transpl Int. 22(1):96-109, 2009 Abu-Elmagd KM et al: Five hundred intestinal and multivisceral transplantations at a single center: major advances with new challenges. Ann Surg. 250(4):567-81, 2009 Linden PK: History of solid organ transplantation and organ donation. Crit Care Clin. 25(1):165-84, ix, 2009 Wu T et al: Histopathologic characteristics of human intestine allograft acute rejection in patients pretreated with thymoglobulin or alemtuzumab. Am J Gastroenterol. 101(7):1617-24, 2006 Starzl TE et al: Tolerogenic immunosuppression for organ transplantation. Lancet. 361(9368):1502-10, 2003 Starzl TE et al: Transplantation tolerance from a historical perspective. Nat Rev Immunol. 1(3):233-9, 2001 Groth CG et al: Historic landmarks in clinical transplantation: conclusions from the consensus conference at the University of California, Los Angeles. World J Surg. 24(7):834-43, 2000 Starzl TE et al: Antigen localization and migration in immunity and tolerance. N Engl J Med. 339(26):1905-13, 1998

423

Intestinal Transplantation

Acute Antibody-Mediated Rejection, Intestine KEY FACTS

ETIOLOGY/PATHOGENESIS • Usually due to preformed lymphocytotoxic antibodies ○ IgG type due to prior allosensitization (panel reactive antibodies) • Frequently coexists with acute cellular rejection

CLINICAL ISSUES • Hyperacute rejection: Within minutes of transplant organ reperfusion • Antibody-mediated rejection: Usually in first 2 weeks but can present later • Leads invariably to graft failure

MICROSCOPIC • Dilated capillaries with neutrophilic aggregates in lumen and plump, prominent endothelial cells • Ulceration in severe cases • Capillary fibrin thrombi, arterial if deeper submucosal tissue included

• No significant lamina propria lymphocytosis or apoptoses in crypts

ANCILLARY TESTS • C4d staining ○ Usually diffuse staining of all capillaries in mucosa and submucosa ○ Needs correlation with DSA titers and should not be interpreted in its absence

TOP DIFFERENTIAL DIAGNOSES • • • •

Acute cellular rejection Severe reperfusion injury Infectious enteritis Ischemic injury

DIAGNOSTIC CHECKLIST • Extent of mucosal injury • Whether frank ulceration is present or not • Evidence of arteritis may suggest more severe injury

Acute Antibody-Mediated Rejection With Ulceration

Mucosa in Antibody-Mediated Rejection

C4d in Mucosal Capillaries

Arteritis

(Left) Resected bowel specimen shows large areas of ulceration and congestion with edema in the submucosa. There is loss of crypts due to dropout in many areas. The absence of significant fibrosis suggests that this is an acute, rather than a chronic, process. (Right) Mucosal congestion, rare crypt apoptoses, and dilated capillaries with prominent endothelial cells are all features that suggest a diagnosis of antibodymediated rejection (AMR).

(Left) A C4d immunohistochemical study shows strong and diffuse endothelial cytoplasmic staining, typical of a positive reaction, suggesting the diagnosis of AMR. (Right) Arteritis can be associated with AMR or mediated by T cells. This image shows severe, transmural arteritis with mononuclear cells infiltrating through the media ſt.

424

Acute Antibody-Mediated Rejection, Intestine

Abbreviations • Antibody-mediated rejection (AMR)

Synonyms • Acute humoral rejection • Acute vascular rejection

Definitions • Rejection of donor organ due to circulating preformed lymphocytotoxic antibodies that results in vascular injury and graft loss • Few cases in literature; remains controversial

ETIOLOGY/PATHOGENESIS Hyperacute Rejection • Usually due to preformed lymphocytotoxic antibodies ○ IgG type due to prior allosensitization [panel reactive antibodies (PRA)] • Antibodies present in high titers before transplant ○ Not detected due to late crossmatch results • Immediate posttransplant period injury to donor organ • Antibodies to class I or II human lymphocyte antigens or ABO blood group antigens • Activation of complement cascade with fibrin and platelet aggregation, leading to thrombosis

Acute Antibody-Mediated Rejection • Usually due to low levels of preformed antibodies that increase over time • Can be due to de novo allosensitization resulting from acute cellular rejection (ACR) • Usually associated with rising donor-specific antibodies (DSA) • Less often seen in combined liver-small bowel transplant due to protection by liver • Common in individuals receiving 2nd transplants due to failed 1st allografts (high PRA) • Frequently coexists with ACR

CLINICAL ISSUES Presentation • Hyperacute rejection: Usually within minutes of transplant organ reperfusion ○ Intraoperative swelling of graft with discoloration ○ Can present in 24-48 hours as surgical emergency ○ Less common now due to availability of crossmatch results prior to transplant and immunosuppression in tacrolimus era • AMR: Usually in first 2 weeks but can present later • Increased stomal output as evidence of graft dysfunction with bleeding in severe cases • Rare cases of mild AMR associated with ACR (incidental biopsy finding)

Endoscopic Findings • Mucosal congestion and discoloration • Mucosal ulcers and hemorrhage

Treatment • Plasmapheresis ○ Best method to reduce circulating antibodies • Steroids • Anti-CD20 antibody (rituximab) • Antirejection drugs to treat accompanying ACR, including anti-CD3 antibody • Rare use of anti-CD138 antibody (bortezomib)

Prognosis • • • •

Intestinal Transplantation

TERMINOLOGY

Hyperacute rejection can result in immediate graft loss Difficult to predict due to paucity of data/literature Leads invariably to graft failure Prolonged graft survival (> 1 year) reported in some instances but final outcome unclear

MACROSCOPIC Hyperacute Rejection • Marked purplish discoloration of bowel with hemorrhage • Transmural ischemic damage • Uncommon to receive resections for AMR other than hyperacute rejection

MICROSCOPIC Histologic Features • Mucosal changes on biopsies ○ Marked congestion ○ Mucosal edema ○ Dilated capillaries with neutrophilic aggregates in lumina and plump, prominent endothelial cells ○ Capillary fibrin thrombi, arterial if deeper submucosal tissue included ○ Large arteries usually not sampled; may show vasculitis and necrosis ○ Neutrophils in lamina propria ○ Ulceration in severe cases ○ No significant lamina propria lymphocytosis or apoptoses in crypts – If crypt apoptosis present, suspect ACR in addition • Resected bowel for hyperacute rejection ○ Mucosal congestion and hemorrhages ○ Transmural ischemia and necrosis ○ Arterial thrombosis and vasculitis, especially mesenteric vessels

ANCILLARY TESTS Immunohistochemistry • C4d ○ Usually diffuse staining of endothelial cells of all mucosal and submucosal capillaries – Normal small bowel □ Shows no capillary C4d staining □ C4d may stain arterial walls (elastic lamina) and lymphoid aggregates ○ Requires correlation with DSA titers to establish diagnosis of AMR ○ Data still limited on utility in diagnosis, unlike kidney transplant ○ Positive staining helps 425

Intestinal Transplantation

Acute Antibody-Mediated Rejection, Intestine – Negative staining does not exclude diagnosis of AMR ○ C4d should be tested, whenever AMR is suspected ○ C4d staining may persist on serial biopsies – May not correlate with DSA

Serologic Testing and Histocompatibility Assays • Positive crossmatch for T- or B-lymphocytotoxic antibodies • PRA

DIFFERENTIAL DIAGNOSIS Acute Cellular Rejection • Distinguished by presence of crypt apoptosis • Increased lymphoplasmacytic infiltrate • No significant neutrophilic component unless overlying ulcer is present • No endothelial activation noted • No fibrin thrombi in capillaries ○ If thrombi present, consider combined ACR and AMR • C4d may be positive without associated PRA or DSA

Severe Reperfusion Injury • Usually seen in 1st week post transplant • Mucosal congestion and hemorrhage with some crypt apoptosis and injury • Mucosal regeneration in 1st week with crypt regenerative changes • C4d(-) • No DSA or PRA • No capillary neutrophil margination or endothelial activation evident

Infectious Enteritis • Bacterial ○ Usually associated with neutrophilic exudate ○ Extensive mucosal ulceration with pseudomembrane in cases of Clostridium difficile enteritis • Viral ○ Usually associated with lymphoplasmacytic infiltrate and ulceration ○ Viral inclusions evident, usually early in course – Cytomegalovirus – Adenovirus – Epstein-Barr virus • Absence of ○ Endothelial activation ○ Fibrin thrombi ○ C4d capillary staining

Ischemic Injury • Usually result of mechanical factors • Can occur at any time post transplant, though common in immediate postsurgical period • Usually related to vascular kinking due to adhesions • Usually segmental • Acute emergency with discoloration of bowel • Segmental resection may show bowel ischemia that may be transmural with necrosis • No vascular thrombosis or vasculitis noted

426

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • • • •

Extent of mucosal injury Whether frank ulceration is present or not Evidence of arteritis, as this may suggest more severe injury Degree of ischemic injury may predict graft loss due to hyperacute rejection

Pathologic Interpretation Pearls • Dilated capillaries with neutrophilic margination • Edema with some neutrophils without lymphoplasmacytic infiltrate • Absence of crypt apoptosis • Presence of fibrin thrombi in vessels • Can coexist with ACR, so presence of apoptosis does not exclude AMR • C4d staining ○ Test for C4d capillary deposition if persistent neutrophilic infiltrates on serial biopsies ○ C4d immunohistochemistry reliable but studies still evolving ○ Only diffuse C4d capillary deposition may warrant suggestion of AMR ○ Focal staining may be seen in ACR cases • Presence of circulating donor antibodies helpful to diagnose AMR • Neutrophilic exudate or pseudomembrane should warrant exclusion of bacterial enteritis

SELECTED REFERENCES 1.

Fujiwara S et al: Effectiveness of bortezomib in a patient with acute rejection associated with an elevation of donor-specific HLA antibodies after smallbowel transplantation: case report. Transplant Proc. 48(2):525-7, 2016 2. Koo J et al: Allograft biopsy findings in patients with small bowel transplantation. Clin Transplant. 30(11):1433-1439, 2016 3. Dick AA et al: Antibody-mediated rejection after intestinal transplantation. Curr Opin Organ Transplant. 17(3):250-7, 2012 4. Tsai HL et al: Association between donor-specific antibodies and acute rejection and resolution in small bowel and multivisceral transplantation. Transplantation. 92(6):709-15, 2011 5. Ashokkumar C et al: Allospecific CD154+ B cells associate with intestine allograft rejection in children. Transplantation. 90(11):1226-31, 2010 6. Ruiz P et al: Immediate antibody-mediated (hyperacute) rejection in smallbowel transplantation and relationship to cross-match status and donorspecific C4d-binding antibodies: case report. Transplant Proc. 42(1):95-9, 2010 7. de Serre NP et al: Evaluation of c4d deposition and circulating antibody in small bowel transplantation. Am J Transplant. 8(6):1290-6, 2008 8. Kato T et al: Association of emergence of HLA antibody and acute rejection in intestinal transplant recipients: a possible evidence of acute humoral sensitization. Transplant Proc. 38(6):1735-7, 2006 9. Troxell ML et al: Evaluation of C4d staining in liver and small intestine allografts. Arch Pathol Lab Med. 130(10):1489-96, 2006 10. Rifle G et al: Donor-specific antibodies in allograft rejection: clinical and experimental data. Transplantation. 79(3 Suppl):S14-8, 2005 11. Wu T et al: A clinicopathologic study of isolated intestinal allografts with preformed IgG lymphocytotoxic antibodies. Hum Pathol. 35(11):1332-9, 2004 12. Ruiz P et al: Mucosal vascular alterations in the early posttransplant period of small bowel allograft recipients may reflect humoral-based allograft rejection. Transplant Proc. 34(3):869-71, 2002

Acute Antibody-Mediated Rejection, Intestine Suspicious for Antibody-Mediated Rejection (Left) A low-magnification image of a biopsy shows superficial epithelial injury with denudation, loss of crypts, congestion, and stromal prominence. There are gaps in between crypts suggesting the loss of crypts. (Right) Mucosal biopsy shows congestion with dilated capillaries and plump, prominent endothelial cells ﬈. Adjacent crypts do not exhibit apoptosis. This should provide a clue to perform C4d staining.

Fibrinous Exudate in Antibody-Mediated Rejection

Intestinal Transplantation

Acute Antibody-Mediated Rejection

Positive C4d Deposition in Capillaries (Left) Biopsy from a patient with AMR shows mucosal ulceration and fibrinous exudate with fibrin in the superficial lamina propria ﬈ and some loss of crypts. There is an increase in lamina propria inflammation. (Right) A C4d immunostain shows many capillaries with strong endothelial staining ﬈. This finding requires correlation with the presence of circulating donor-specific antibodies to establish a diagnosis of AMR.

Organized Thrombus

Severe Arterial Inflammation (Left) Serosal vessel from a resection specimen shows an organized thrombus with recanalization. The entire lumen is obliterated. The thrombus shows scattered inflammatory cells. (Right) Another vessel showing eccentric inflammatory changes in the wall with fibrinoid necrosis ﬈ and luminal narrowing suggests acute vascular injury as a mechanism for graft loss. This change may be seen in AMR manifesting as hyperacute rejection.

427

Intestinal Transplantation

Acute Cellular Rejection, Intestine KEY FACTS

• Most common and early complication • Usually between 3 days up to years post transplant • Majority in 1st month ○ 1st episode median: 2.5 weeks • Fevers, abdominal pain, vomiting, nausea, and watery diarrhea • Biopsy to include at least 2, but average 3, pieces of mucosa • Most grafts with mild acute cellular rejection (ACR) can be treated and recover normal function • Higher grades of rejection can improve or progress to chronic rejection • Graft loss once chronic rejection evident

○ Monocytes and macrophages ○ Mast cells ○ Eosinophils ○ Few neutrophils • Minimum of 10 crypts needed for evaluation of rejection • Grading system proposed that includes ○ Indeterminate ○ Mild – ≥ 6 apoptoses per 10 crypts ○ Moderate ○ Severe – Mucosal ulceration with exudate and granulation tissue

MICROSCOPIC

TOP DIFFERENTIAL DIAGNOSES

• Apoptosis is hallmark of rejection • Usually associated with inflammatory infiltrate ○ Lymphocytes ○ Plasma cells

• • • •

CLINICAL ISSUES

Reperfusion injury Bacterial infections Viral infections Antibody-mediated rejection

Typical Epithelial Crypt Apoptosis

Severe ACR

Crypt Apoptosis

Multiple Crypt Apoptoses

(Left) H&E shows crypt demonstrating apoptosis with fragmentation of the nucleus and hyperchromasia and vacuolation of the cytoplasm ﬈. Background infiltrate of lymphocytes, plasma cells, and eosinophils is seen. (Right) Scanning image of severe acute cellular rejection (ACR) shows mucosal ulceration with loss of crypts and villous outlines and presence of a few residual crypts. The lamina propria resembles granulation tissue.

(Left) Another example of a small bowel biopsy shows parallel arranged crypts with scattered large epithelial apoptoses ſt at their bases. Note nuclear condensation, rounding with eosinophilia and fragmentation. (Right) H&E shows small bowel crypts with multiple apoptotic bodies ſt and occasional crypt with confluent adjacent apoptoses ﬇. The crypts are visualized in cross section.

428

Acute Cellular Rejection, Intestine Laboratory Tests

• T-cell-mediated rejection

• Fecal calprotectin measurement ○ Increased during ACR but not specific • Granzyme B and perforin increased in ACR • CD154(+) Tc memory cells by flow cytometry ○ Help classify patients as rejectors or nonrejectors

Definitions

Treatment

• Immunologic host response resulting in cellular infiltration and injury of allograft in absence of preformed antibodies

• Depends on severity of changes, both endoscopic and histologic ○ Mild ACR: Steroid bolus and FK506 modulations ○ Moderate: Steroid and FK506 ± Campath or OKT6 ○ Severe: Multiple drugs and alemtuzumab treatment

Abbreviations • Acute cellular rejection (ACR)

Synonyms

ETIOLOGY/PATHOGENESIS Mechanisms of Acute Cellular Rejection • T-cell-mediated disease ○ More common than other organ transplants due to large amount of donor lymphoid tissue • Other cells implicated include ○ Dendritic cells ○ Monocytes/macrophages ○ Possibly eosinophils and mast cells • Isolated small bowel (SB) transplant has higher incidence than SB-liver transplant ○ Protective influence of liver proposed • Toll-like receptors also implicated to have role • Improved survival of graft after antithymoglobulin ○ Due to reduction of donor-specific, inflammatory CD154(+) T cytotoxic (Tc) cells • Aberrant innate immunity via NOD2 gene suggested as possible mechanism

CLINICAL ISSUES Epidemiology • Incidence ○ Most common and early complication of SB transplant ○ Most recipients have ≥ 1 episode of ACR ○ Majority in 1st month – 1st episode median: 2.5 weeks

Presentation • • • • • • •

Fevers Abdominal pain Vomiting, nausea Watery diarrhea Increased stomal output Abdominal distension and ileus Metabolic acidosis

Endoscopic Findings • • • • • • •

Mucosal edema Loss of mucosal pattern Hyperemia Ischemic dusky mucosa with ulcerations Extensive mucosal denudation in severe cases Bleeding and loss of peristalsis Protocol biopsies early on in course, later endoscopies dictated by symptomatology • Biopsy to include at least 2, but average 3, pieces of mucosa • Avoid anastomotic sites and blind loops unless pathology suspected

Intestinal Transplantation

TERMINOLOGY

Prognosis • Most grafts with mild ACR treated and recover normal function • Nonrejectors on low-dose immunosuppressive therapy maintenance do well • Higher grades of rejection can improve or progress to chronic rejection • Worse prognosis for repeated episodes of rejection or severe exfoliative rejection • Graft loss once chronic rejection evident

MACROSCOPIC General Features • Unusual to receive surgical resections in mild or moderate rejection • Severe exfoliative rejection can result in graft enterectomy • Mucosa shows hemorrhagic appearance with extensive mucosal sloughing • Thickening of bowel wall due to fibrosis

MICROSCOPIC Histologic Features • Apoptosis is hallmark of rejection • Usually associated with inflammatory infiltrate ○ Lymphocytes ○ Plasma cells ○ Monocytes and macrophages ○ Mast cells ○ Eosinophils ○ Few neutrophils • No infectious etiology for inflammation or viral inclusions evident • ≥ 10 crypts needed for evaluation of rejection • Proposed grading system includes ○ Indeterminate: 4-5 scattered apoptoses per 10 crypts with minimal inflammation – With thymoglobulin or alemtuzumab pretreatment, mucosal changes precede ACR □ Neutrophils in lamina propria, lesser lymphocytes and eosinophils, and edema ○ Mild ACR – ≥ 6 apoptoses per 10 crypts – Expanded lamina propria with mixed inflammatory infiltrate – No crypt loss or ulceration 429

Intestinal Transplantation

Acute Cellular Rejection, Intestine – Identify epithelial and exclude inflammatory cell apoptoses – Architectural distortion, villous blunting, edema, and congestion ○ Moderate ACR – Increased apoptoses (more than mild ACR) – Many crypts with multiple and confluent apoptoses causing holes in crypt epithelium – Crypt injury with loss of Paneth cells and sloughing of cells into lumen – Areas with crypt dropout with ghost outlines of necrotic crypts – No mucosal ulceration or significant mucosal fibrosis – Increased lamina propria cellularity with many activated blastoid lymphocytes, small lymphocytes, plasma cells, eosinophils, macrophages, and some neutrophils – Crypt regeneration evident with many mitoses and hyperchromasia – On healing, residual crypt distortion ○ Severe ACR – Extensive crypt damage with dropout, can be patchy and variable among fragments – Mucosal ulceration with exudate and granulation tissue – Occasional biopsies with only granulation tissue and no crypts – Residual crypts with apoptoses – Can persist on several mucosal biopsies and can be segmental with regional variations – In prolonged episodes, mucosal regeneration at edges of ulcers begins from surface, subsequent crypt formation – More frequent in lower endoscopic biopsies than in upper portions of allograft – Better assessment of overall graft with both upper and lower biopsies in such cases ○ Exfoliative rejection – Form of severe rejection involving entire bowel – Diffuse mucosal ulceration with granulation tissue – Complete absence of crypts and no evidence of regeneration of epithelium – Can result in fibrosis and distortion of architecture over time, if healing occurs – Progress often to chronic rejection – Frequently necessitate graft enterectomy for severe symptoms

DIFFERENTIAL DIAGNOSIS Reperfusion Injury • Mucosal congestion and hemorrhage rather than inflammatory infiltrate • Crypt regeneration and fewer apoptoses as opposed to ACR (more apoptoses)

Bacterial Infections • Clostridium difficile with ulcers and exudate ○ Usually superficial changes and injury with abundant neutrophils ○ No crypt apoptoses or lymphocytes in lamina propria ○ Stool positive for C. difficile toxin 430

Viral Infections • Rotavirus ○ Superficial epithelial regenerative changes with reactive appearance ○ Lamina propria with few neutrophils and lymphocytes ○ Villous alterations with no viral inclusions ○ No significant crypt apoptoses • Adenovirus ○ Superficial epithelial reactive changes with piling up of cells ○ No significant crypt injury or apoptoses ○ "Smudge" cells: Intranuclear inclusions in surface epithelial cells ○ Variation in size of epithelial nuclei with eosinophilic to amphophilic inclusions with peripheral chromatin ring ○ Adenovirus immunostain diagnostic • Cytomegalovirus (CMV) infection ○ Classic inclusion of CMV: Intranuclear and eosinophilic granular cytoplasmic inclusions ○ Neutrophils in lamina propria ○ No activated lymphocytes or crypt apoptoses ○ Surface ulceration in severe cases ○ Can coexist with rejection in same biopsy • Epstein-Barr virus (EBV) enteritis and posttransplant lymphoproliferative disorder (PTLD) ○ Predominantly lymphoplasmacytic infiltrate ○ No crypt apoptoses ○ Lymphoid nodules with overlying ulcer: Suspect PTLD ○ EBER stain diagnostic in most; enteritis or PTLD

Antibody-Mediated Rejection • Early in transplant with mucosal congestion, hemorrhage, and ulceration • Prominent dilated capillaries with fibrin-platelet thrombi &/or neutrophils in capillaries • No significant lamina propria inflammatory infiltrate • No significant crypt apoptoses ○ If present, consider combination of both ACR and antibody-mediated rejection (AMR) • C4d diffusely positive in capillary endothelium ○ Focal C4d stain interpreted as normal • Circulating donor-specific antibody demonstration required

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Biopsy to include at least 2, but average 3, pieces of mucosa • Apoptosis is hallmark of rejection • Grading system proposed that includes ○ Indeterminate ○ Mild ○ Moderate ○ Severe grades of rejection – Can take weeks to recover to normalcy • Rejection is patchy process ○ Changes may differ in upper and lower GI tract • Exfoliative rejection warrants graft enterectomy if severe • Severe rejection can progress to or accelerate chronic rejection • Check for donor-specific antibodies if persistent rejection

Acute Cellular Rejection, Intestine

Grade

Histologic Features

Indeterminate for acute cellular rejection (ACR)

Relatively preserved architecture Mild increase in lamina propria cellularity lymphocytes, activated lymphocytes, plasma cells, and eosinophils Scattered crypt apoptosis < 6/10 crypts

Mild ACR

Villous architectural changes with blunting Increased cellularity with activated lymphocytes and few plasma cells with monocytes and eosinophils

Intestinal Transplantation

Histologic Grading of Acute Cellular Rejection

Crypt apoptosis ≥ 6/10 crypts or confluent apoptosis in occasional crypt No surface ulceration or viral inclusions Moderate ACR

Increased architectural distortion with areas of crypt dropout and villous alterations Increased lamina propria cellularity: Mononuclear, mainly with eosinophils Readily evident crypt apoptoses with multifocal confluent apoptosis and crypt lumina with cellular debris No ulceration

Severe ACR

Ulceration of mucosal fragments with exudate and granulation tissue Extensive areas of crypt loss Residual crypts with marked apoptoses or regenerative changes

Modified from Wu T et al: A schema for histologic grading of small intestine allograft acute rejection. Transplantation. 75(8):1241-8, 2003 and Ruiz P et al: International grading scheme for acute cellular rejection in small-bowel transplantation: single-center experience. Transplant Proc. 42(1):47-53, 2010.

Differential Diagnosis of Acute Cellular Rejection: When to Consider What Timing of Event

Grade of Rejection

Differential to Consider

1st week

Mild rejection

Reperfusion injury Bacterial infection, sepsis

Severe rejection

Severe reperfusion injury Antibody-mediated rejection Exclude anastomotic ulcer

2nd week to months

Mild rejection

Clostridium difficile, rotavirus, or adenovirus CMV, EBV

Severe rejection/ulceration

Antibody-mediated rejection EBV or PTLD Exfoliative rejection Chronic rejection

Pathologic Interpretation Pearls • Crypt apoptosis: Think rejection • Neutrophils: Think infection or AMR if within capillaries • Lymphoplasmacytic infiltrate without apoptosis: Think viral infection (EBV, others) • Eosinophilia: No clear significance: Frequent accompaniment of rejection; possible drugs as cause • Ulcer: Severe rejection, AMR, PTLD

SELECTED REFERENCES 1. 2.

3.

Ashokkumar C et al: Predicting cellular rejection with a cell-based assay: Preclinical evaluation in children. Transplantation. 101(1):131-140, 2017 Remotti H et al: Small-bowel allograft biopsies in the management of smallintestinal and multivisceral transplant recipients: histopathologic review and clinical correlations. Arch Pathol Lab Med. 136(7):761-71, 2012 Sindhi R et al: Allospecific CD154 + T-cytotoxic memory cells as potential surrogate for rejection risk in pediatric intestine transplantation. Pediatr Transplant. 16(1):83-91, 2012

4.

Tsai HL et al: Association between donor-specific antibodies and acute rejection and resolution in small bowel and multivisceral transplantation. Transplantation. 92(6):709-15, 2011 5. Ruiz P et al: International grading scheme for acute cellular rejection in smallbowel transplantation: single-center experience. Transplant Proc. 42(1):4753, 2010 6. Sindhi R et al: Immune monitoring in small bowel transplantation. Curr Opin Organ Transplant. 15(3):349-56, 2010 7. Ruiz P et al: Histological criteria for the identification of acute cellular rejection in human small bowel allografts: results of the pathology workshop at the VIII International Small Bowel Transplant Symposium. Transplant Proc. 36(2):335-7, 2004 8. Ishii T et al: Exfoliative rejection after intestinal transplantation in children. Pediatr Transplant. 7(3):185-91, 2003 9. Wu T et al: A schema for histologic grading of small intestine allograft acute rejection. Transplantation. 75(8):1241-8, 2003 10. Lee RG et al: Pathology of human intestinal transplantation. Gastroenterology. 110(6):1820-34, 1996 11. White FV et al: Pathology of intestinal transplantation in children. Am J Surg Pathol. 19(6):687-98, 1995 12. Jaffe R et al: Multivisceral intestinal transplantation: surgical pathology. Pediatr Pathol. 9(6):633-54, 1989

431

Intestinal Transplantation

Acute Cellular Rejection, Intestine

Adequate Biopsy

ACR

Mild ACR

Moderate ACR

Moderate ACR

Moderate ACR

(Left) Scanning view shows adequate crypts to evaluate for rejection. Note some blunting of villi and some crypt architectural distortion, suggesting pathology of ACR. (Right) High-power view of ACR shows a crypt with 2 large epithelial apoptoses ſt. Note inflammatory infiltrate of lymphocytes, activated lymphocytes, plasma cells, and eosinophils. The surface epithelium is intact.

(Left) High-power view of mild ACR shows a single crypt with confluent apoptosis, i.e., 3-4 adjacent epithelial cells undergoing apoptosis ſt. This was the only crypt with this finding. Other crypts showed increased apoptoses, supporting the diagnosis of ACR. (Right) H&E shows moderate rejection with some architectural distortion and a crypt with abundant necrotic debris ﬈. Lamina propria shows increased inflammatory infiltrate. No ulceration is noted, and villous architecture is preserved.

(Left) Higher magnification of moderate ACR shows a crypt with a lumen containing apoptotic debris ſt. There is increased inflammation, including lymphocytes, eosinophils, and plasma cells. (Right) Biopsy shows features of moderate ACR. Here, multiple crypts show evidence of confluent apoptosis with debris in the lumina of many crypts. Note the attenuated epithelial lining of the crypts. No ulceration is seen, though the surface epithelium appears injured.

432

Acute Cellular Rejection, Intestine

Day 5 Biopsy: Mild ACR (Left) Day 0 perfused bowel biopsy shows mucosal sloughing with congestion, dilated capillaries, reactive changes in epithelium, and crypt and lamina propria inflammatory cells, all features of reperfusion injury. (Right) Biopsy at day 5 shows a crypt with apoptotic cell debris ſt, consistent with a mild ACR. Reperfusion injury can increase risk of ACR.

Week 2 Biopsy: Severe ACR

Intestinal Transplantation

Day 0 Biopsy: Reperfusion Injury

Week 3 Biopsy: Severe ACR (Left) Biopsy at 2 weeks shows sloughing of epithelium with loss of crypts, ulcer, granulation tissue, and exudate. These features portend the beginning of a severe rejection episode that can persist for several biopsies. (Right) Biopsy at 3 weeks shows persistence of ulceration with granulation tissue with complete loss of crypts and surface epithelial lifting. These are features of an ongoing episode of severe ACR.

Week 5 Biopsy: Regeneration

Week 6 Biopsy: Appearance of Crypts (Left) Biopsy at 5 weeks shows early reepithelialization starting from the sides. The crypts develop subsequently. The lamina propria still shows granulation tissue, suggesting continuation of an episode of severe ACR. (Right) Biopsy at 6 weeks shows presence of epithelium with many goblet cells along with appearance of crypts. Still evident are stromal changes in lamina propria. This biopsy suggests an end to the episode of severe ACR with regeneration of mucosa.

433

Intestinal Transplantation

Acute Cellular Rejection, Intestine

Reperfusion Injury

Reperfusion Injury

Antibody-Mediated Rejection

Rotavirus Enteritis

EBV Enteritis

EBV Enteritis

(Left) Day 0 biopsy shows early injury of reperfusion in the bowel with surface epithelial denudation, stromal changes with vascularity, and rare apoptosis in crypts (ischemic changes). (Right) Higher magnification of reperfusion injury shows some crypt injury as well as increased cellularity in the lamina propria and rare apoptoses.

(Left) Day 4 biopsy of an episode of antibody-mediated rejection shows surface epithelial denudation with stromal vascularity and appearance of granulation tissue without an exudate, mimicking reperfusion injury. This warrants a C4d stain. (Right) Rotavirus enteritis shows villous blunting with reactive appearance of epithelium without viral inclusions. Note the associated increased cellularity in the lamina propria, which includes a few neutrophils. There are no crypt apoptoses.

(Left) Biopsy shows increased lamina propria cellularity with blastic lymphocytes, few plasma cells, and rare eosinophils with no significant apoptosis. This should warrant exclusion of viral etiology, especially Epstein-Barr virus (EBV) infection. (Right) In situ hybridization for EBER probe highlights scattered lamina propria lymphocytes with nuclear staining for EBER ﬈, confirming EBV enteritis. This finding is invariably associated with high blood EBV levels by PCR.

434

Acute Cellular Rejection, Intestine

Severe Rejection (Left) Biopsy shows erosions with marked lamina propria inflammation and distortion of architecture with loss of crypts. Residual crypts show apoptoses with debris in the lumina. There is prominent vascularity. The surface epithelium shows marked regenerative changes with loss of goblet cells. (Right) Biopsy shows partial retention of villous architecture and an area with ulceration ﬈, highlighting the segmental nature of a severe rejection process.

Explant Allograft

Intestinal Transplantation

Moderate to Severe Rejection

Severe Exfoliative Rejection (Left) Gross photo of bowel resected for severe exfoliative rejection shows an erythematous and edematous mucosa with ulcerations and hemorrhage. Areas of cobblestoning are evident ſt. (Right) Scanning view of exfoliative rejection shows edematous submucosa and a thinned-out lamina propria with complete loss of epithelium and crypts with inflammatory infiltrate restricted to the mucosa. Some serosal inflammation is noted in this case.

Severe Exfoliative Rejection

Denuded Mucosa: Exfoliative Rejection (Left) H&E of exfoliative rejection shows mucosal ulceration with surface exudate consisting of fibrin ﬈ with inflammatory cells and complete absence of crypt or surface epithelium. There is some inflammation extending into the submucosa in this region. (Right) High magnification highlights the cellular infiltrate of lymphocytes and plasma cells with prominent vascularity and surface exudate.

435

Intestinal Transplantation

Chronic Rejection, Intestine KEY FACTS

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• Early onset of acute cellular rejection or multiple episodes increase risk of chronic rejection (CR) • Severe exfoliative rejection may progress to CR in many cases • Progressive vascular injury resulting in graft vasculopathy and ischemic injury

• • • • •

CLINICAL ISSUES • Diarrhea or increased stomal output • Hematochezia • Invariably leads to graft loss

MACROSCOPIC • Extensive ulceration • Thickened bowel loops with adhesions • Marked fibrosis of bowel wall and serosa

Ulceration with granulation tissue Distortion of crypt architecture with pyloric metaplasia Crypt loss Fibrosis in wall with muscular hypertrophy/atrophy Fibrosing/sclerosing peritonitis ○ Subserosal and serosal fibrosis ± inflammation • Perforating and mesenteric vessels with intimal thickening and luminal narrowing/obliteration

TOP DIFFERENTIAL DIAGNOSES • Severe acute cellular rejection • Vascular compromise ○ Arterial thrombosis or adhesions • Viral infections

Chronic Rejection, Resection

Graft Vasculopathy

Chronic Allograft Rejection: Gross

Chronic Graft Vasculopathy

(Left) Scanning magnification shows an ulcerated allograft with marked architectural distortion and extensive fibrosis. Note mucosa with indistinct separation from submucosa and complete loss of crypts. (Right) A classic feature of chronic rejection is mesenteric arterial intimal proliferation with luminal narrowing. These changes are present to varying degrees and can only be seen on bowel resection.

(Left) An example of a resected allograft shows extensive adhesions of bowel loops with thickening of the bowel wall and ectatic vessels in the mesentery ﬈. Note an occasional mesenteric artery with thickened wall ſt. (Right) A mesenteric arterial branch shows accumulation of foam cells in the intima with associated narrowing of the lumina (foam cell arteriopathy ﬈). Note intermixed lymphocytes.

436

Chronic Rejection, Intestine

Abbreviations • Chronic rejection (CR)

Definitions

○

• Progressive injury by immunologic factors and ischemia resulting in irreversible vascular damage and graft loss

○

ETIOLOGY/PATHOGENESIS ○ ○

General Aspects • Increased risk of CR with ○ Prolonged cold ischemia time and advanced donor age ○ Early onset or multiple acute cellular rejection (ACR) episodes • Severe exfoliative rejection may progress to CR

CLINICAL ISSUES Presentation • Diarrhea or increased stomal output; hematochezia

Treatment • Aggressive treatment of ACR with steroids and tacrolimus (FK506) • Antibody treatment (e.g., alemtuzumab) for severe ACR • Graft enterectomy in irreversible cases

Prognosis • Poor once CR begins; results in graft loss

MACROSCOPIC General Features • • • •

Marked congestion of mucosa Extensive ulceration Thickened bowel loops with adhesions Large prominent mesenteric vessels with thickened walls

Sections to Be Submitted • Multiple sections of bowel with focus on ulcerated areas and adjoining preserved mucosa • Multiple sections of mesentery to study vessels ○ Always evaluate in stoma revision specimen for unsuspected CR • Full-thickness sections • Longitudinal sections of intestine wall to study perforating arteries • Any nodules incidentally noted on sectioning/dissecting

DIFFERENTIAL DIAGNOSIS Severe Acute Cellular Rejection • Severe ACR with ulceration (especially exfoliative rejection) mimics CR • Large number of apoptoses in residual glands with active ulcerations favor ACR • Lack of architectural distortion in regenerating bowel • Can coexist or evolve into CR

Arterial Thrombosis or Adhesions • Difficult to differentiate on biopsies • Imaging may demonstrate vascular occlusion and explanation for biopsy findings • Usually acute but can result in persistent mucosal ulceration • Usually segmental and diagnosed on resected specimen

Viral Infections • Cytomegalovirus can cause ulcers with crypt distortion and regenerative changes

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Serial biopsies with architectural distortion, especially following episodes of ACR ○ Essential to diagnose changes of CR • Crypt loss and persistent ulceration in biopsies • Fibrosis in lamina propria or transmural fibrosis as seen with trichrome stain • Mesenteric vessels with intimal proliferation and luminal narrowing but no thrombosis • Sclerosing peritonitis/mesenteritis is specifically associated with CR

SELECTED REFERENCES

MICROSCOPIC

1.

Histologic Features

2.

• Biopsy ○ Ulceration with granulation tissue ○ Crypt loss ○ Marked stromal changes with repair and regeneration of surface epithelium ○ Distortion of crypt architecture with pyloric metaplasia • Resection specimen ○ Mucosal changes – Extensive ulceration and denudation of mucosa

– Loss of crypts in large segments – Marked architectural and crypt distortion – Granulation tissue or stromal changes in lamina propria – Pyloric &/or neuronal metaplasia Subserosal and serosal fibrosis ± inflammation: Fibrosing/sclerosing peritonitis Perforating and mesenteric vessels with intimal thickening, foam cells, muscular hyperplasia, and luminal narrowing/obliteration Masson trichrome highlights intestinal wall fibrosis If nodules noted, may represent posttransplant lymphoproliferative disease or smooth muscle tumor

Intestinal Transplantation

TERMINOLOGY

3. 4.

5. 6.

7.

Nayyar N et al: Pediatric small bowel transplantation. Semin Pediatr Surg. 19(1):68-77, 2010 Ramos E et al: Chronic rejection with sclerosing peritonitis following pediatric intestinal transplantation. Pediatr Transplant. 11(8):937-41, 2007 Klaus A et al: Diffuse mesenterial sclerosis: a characteristic feature of chronic small-bowel allograft rejection. Virchows Arch. 442(1):48-55, 2003 Parizhskaya M et al: Chronic rejection of small bowel grafts: pediatric and adult study of risk factors and morphologic progression. Pediatr Dev Pathol. 6(3):240-50, 2003 Noguchi Si S et al: Pediatric intestinal transplantation: the resected allograft. Pediatr Dev Pathol. 5(1):3-21, 2002 Demetris AJ et al: Chronic rejection. A general overview of histopathology and pathophysiology with emphasis on liver, heart and intestinal allografts. Ann Transplant. 2(2):27-44, 1997 Langrehr JM et al: Clinical course, morphology, and treatment of chronically rejecting small bowel allografts. Transplantation. 55(2):242-50, 1993

437

Intestinal Transplantation

Chronic Rejection, Intestine

Chronic Rejection

Chronic Rejection

Severe Acute Cellular Rejection

Chronic Rejection

Chronic Rejection

Chronic Graft Vasculopathy

(Left) This biopsy shows marked architectural distortion with loss of crypts, fibrosis, and scant inflammatory infiltrate in the lamina propria. There is also some splaying of the muscularis mucosa. (Right) Biopsy of chronic rejection shows marked architectural distortion, stromal changes, and loss of crypts. The surface epithelium here demonstrates regeneration. The muscularis mucosa is thickened, and submucosal inflammation is present.

(Left) This example shows progression to chronic rejection with surface epithelial loss, increased lamina propria cellularity with lymphocytes and plasma cells, and crypt loss. Note also crypts ﬊ with apoptoses, suggesting accompanying acute cellular rejection. (Right) A mucosal section illustrates crypt distortion with pyloric metaplasia ﬈, which may provide clues to a diagnosis of chronic rejection.

(Left) Low-power view of bowel resected for chronic rejection shows mucosal architectural distortion with uniform separation of crypts ﬈ from the muscularis mucosa, submucosa with vascular proliferation, thickened muscularis propria ﬊, and serosal edema. (Right) Trichome stain highlights a mesenteric artery with features of intimal hyperplasia ﬊ with inflammatory cells, resulting in luminal narrowing. This represents the pathognomonic sign of chronic rejection.

438

Chronic Rejection, Intestine

Chronic Rejection (Left) Resection specimen of a small bowel allograft that was removed for chronic rejection shows irregular thickening of the wall with submucosal fibrosis and muscular hypertrophy ﬉ and atrophy ﬊. Note a single serosal obliterated artery ﬈. (Right) This section demonstrates the mucosal changes in chronic rejection. There is villous architectural distortion ﬈ with variation in height of villi along with basal separation from muscularis mucosa ﬊. No apoptosis is seen.

Chronic Rejection: Graft Vasculopathy

Intestinal Transplantation

Chronic Rejection

Chronic Rejection (Left) Vascular changes seen in chronic rejection show almost concentric intimal proliferation with near obliteration of the lumen with associated inflammatory infiltrate in the wall composed mainly of lymphocytes (endarteritis) ﬈. This change may be seen in an occasional artery in the serosa. (Right) At low magnification, an elastic trichrome stain of specimen shows a resected allograft with submucosal fibrosis, preserved mucosa, and thickened serosal artery.

Chronic Rejection

Chronic Graft Vasculopathy (Left) Low-magnification image of an elastic trichromestained allograft section reveals a submucosal thickened artery ﬈ as well as a zone of serosal mesenteric fibrosis (sclerosing peritonitis) ﬊. (Right) Trichome stain shows a lesion that is the hallmark of chronic rejection. A large artery in the mesentery shows eccentric intimal proliferation leading to progressive narrowing of the lumen and secondary ischemic effect.

439

Intestinal Transplantation

Stomach Rejection KEY FACTS

CLINICAL ISSUES • • • • • •

Abdominal pain Stomal output increase Fevers Usually in children with multivisceral transplant Often simultaneous rejection of stomach and intestine Endoscopy with redness and erosions possible ○ Involvement of other parts of allograft often visualized

MICROSCOPIC • Similar histologic features to small bowel allograft rejection • Degree of apoptotic activity often differs from intestinal biopsies and should be correlated

• Graft-vs.-host disease ○ Native involvement

DIAGNOSTIC CHECKLIST • Degree of apoptoses may be much less than small bowel mucosa • Degree of inflammation much less than small intestine • Other causes of gastritis can mimic and interfere with rejection diagnosis • Important to confirm that gastric mucosa represents allograft and not native stomach, as interpretations of changes may differ

TOP DIFFERENTIAL DIAGNOSES • Diseases of native stomach ○ Helicobacter pylori gastritis or reactive gastropathy • Epstein-Barr virus gastritis ○ Native or allograft involvement

Long Segment HSCR

Long Segment HSCR

Indeterminate for ACR

Indeterminate for ACR

(Left) Low-magnification image of gastric wall in an example of Hirschsprung disease (HSCR) shows absence of ganglion cells in lamina propria and no separation of muscle layers ﬈ to indicate myenteric plexus development. (Right) Highpower image of gastric muscularis propria of a child with HSCR shows a large hypertrophic nerve trunk ﬈ with ganglion cells ﬊, confirming a transition zone in the stomach.

(Left) Biopsy section of stomach shows minimal inflammation and some glandular damage ﬈ with no significant apoptosis, which is indeterminate for acute cellular rejection (ACR). (Right) Gastric biopsy section shows isolated apoptoses ſt in a gland with no significant inflammation, supporting a diagnosis of indeterminate for ACR.

440

Stomach Rejection

Epidemiology • Stomach transplant usually component of multivisceral transplant • Most common indications for stomach transplantation ○ Long segment Hirschsprung disease (HSCR) ○ Pseudoobstruction • Uncommon site for biopsy ○ Degree of rejection is less than small bowel

Presentation • Abdominal pain, stomal output increase, fevers • Isolated stomach rejection unusual in absence of duodenal or intestinal rejection

• May involve native or allograft stomach • Increased lamina propria inflammatory infiltrate with activated lymphocytes present • Perform EBER (Epstein-Barr virus-encoded RNA) testing when suspected • Apoptoses less likely unless coexisting ACR present

Graft-vs.-Host Disease • Only native stomach involved • Increased apoptosis within glands • Increased lamina propria inflammation including lymphocytes and neutrophils

Intestinal Transplantation

Epstein-Barr Virus Gastritis

CLINICAL ISSUES

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls

MICROSCOPIC Histologic Features • Similar to small bowel allograft rejection ○ Lesser degree of involvement • Grade rejection based on ○ Degree of apoptosis ○ Inflammatory infiltrate ○ Surface ulceration ○ Lamina propria and glandular epithelial changes ○ Severe cases show marked architectural changes and more obvious apoptoses and glandular loss • Inflammatory infiltrate ○ Mainly lymphocytes and plasma cells – Can include eosinophils and neutrophils • Degree of apoptotic activity often differs from intestinal biopsies and should be correlated • Glandular apoptoses are less prominent in stomach ○ Compared to intestine and can be easily missed • Stomal fibrosis, if present, indicates persistent or chronic rejection

• • • •

Gastric mucosal changes can be subtle Degree of apoptoses often much less than small bowel Degree of inflammation much less than in small intestine Reactive mucosal changes with scattered apoptoses may be main finding • Severe cases show ○ Crypt dropout ○ Multiple apoptoses ○ Mucosal ulceration • Other causes of gastritis frequently mimic and interfere with rejection diagnosis • Important to confirm that gastric mucosa represents allograft and not native stomach, as interpretations of changes may differ

SELECTED REFERENCES 1.

2. 3.

DIFFERENTIAL DIAGNOSIS Diseases of Native Stomach

4.

• Chronic active gastritis ± Helicobacter pylori • Chemical or reactive gastropathy ○ Reactive epithelial changes and corkscrew appearance of glands with superficial inflammation • Acute erosive gastritis, drug induced, with superficial erosions and neutrophils

5. 6. 7.

Lee E et al: Multivisceral transplantation for abdominal tumors in children: a single center experience and review of the literature. Pediatr Transplant. 21(5), 2017 Kubal CA et al: Intestine and multivisceral transplantation: current status and future directions. Curr Gastroenterol Rep. 17(1):427, 2015 Abu-Elmagd KM et al: Five hundred intestinal and multivisceral transplantations at a single center: major advances with new challenges. Ann Surg. 250(4):567-81, 2009 Takahashi H et al: Analysis of acute and chronic rejection in multiple organ allografts from retransplantation and autopsy cases of multivisceral transplantation. Transplantation. 85(11):1610-6, 2008 Kato T et al: Intestinal and multivisceral transplantation in children. Ann Surg. 243(6):756-64; discussion 764-6, 2006 Tzakis AG et al: 100 multivisceral transplants at a single center. Ann Surg. 242(4):480-90; discussion 491-3, 2005 Garcia M et al: Acute cellular rejection grading scheme for human gastric allografts. Hum Pathol. 35(3):343-9, 2004

Grading of Stomach Allograft Rejection Grade

Histologic Features

0 (no rejection)

Scant inflammation with no significant apoptosis or surface injury; no lamina propria changes

1 (indeterminate for ACR)

Mild inflammatory infiltrate in LP with epithelial damage without significant apoptoses

2 (mild ACR)

Mixed inflammatory infiltrate with single cell apoptoses in glands; mild distortion of architecture

3 (moderate ACR)

Increased LP inflammatory infiltrate with glandular damage, confluent apoptoses, architectural disarray

4 (severe ACR)

Numerous apoptoses with glandular dropout, architectural distortion, mixed inflammation, and ulceration

LP = lamina propria; ACR = acute cellular rejection. Modified from Garcia M et al: Hum Pathol. 35(3):343-9, 2004.

441

Intestinal Transplantation

Stomach Rejection

Severe ACR

Severe ACR

Moderate ACR

Moderate ACR

Gastric Rejection

Candida Gastritis

(Left) Gastric biopsy section shows areas of mucosal erosion with fibrinous exudate, underlying patchy glandular loss and regeneration. Architectural distortion and increased inflammation in the lamina propria are also present, suggesting severe ACR. (Right) Section of gastric mucosa shows glandular destruction as evidenced by apoptoses with apoptotic debris ﬈ in the lumen of the gland. Note an attenuated and partial lining epithelium in this example of severe ACR.

(Left) Section of gastric mucosa shows inflammatory infiltrate in the lamina propria and glandular apoptosis ſt in an example of moderate ACR. (Right) Another section of allograft stomach shows markedly reactive glandular epithelial changes with rare apoptoses ﬈ and inflammation in the lamina propria. This is an example of moderate ACR.

(Left) An image of a gastric body mucosal biopsy shows glandular injury with apoptotic cellular debris in lumen of glands ﬈ and loss of some epithelial cells. This would suggest a mild ACR, but note the overlap with reactive gastropathy. (Right) A gastric biopsy showing an acute inflammatory exudate with abundant budding yeasts ﬈ and occasional pseudohyphae ſt of Candida species.

442

Stomach Rejection

Arterial Thrombosis Effect (Left) Low-magnification image shows mucosal erosions and superficial fibrin with glandular dropout and hemorrhage compatible with changes of acute ischemia due to arterial thrombosis. (Right) Higher magnification view of acute ischemic injury in the stomach due to arterial thrombosis shows glandular disruption, fibrin, and epithelial necrosis with lamina propria congestion and hemorrhage.

Reactive Gastropathy

Intestinal Transplantation

Acute Ischemic Ulcer

Reactive Gastropathy (Left) Low-magnification image shows acute reactive gastropathy changes in the gastric mucosa of the native stomach with erosions, regenerative epithelial cells, and inflammation that mimics ACR in the allograft. Note the superficial exudate with fibrin. (Right) Higher magnification view of acute reactive gastropathy shows superficial erosions with neutrophils and reactive epithelial changes in the glands ſt. Note also the presence of eosinophils in the lamina propria.

EBV Gastritis

EBV Gastritis (Left) Allograft stomach biopsy shows an intense lamina propria infiltrate of lymphocytes, including some activated forms, and hyperplastic glandular changes without associated apoptoses. These features should prompt testing by EBER in situ hybridization to exclude Epstein-Barr virus (EBV) gastritis. (Right) An EBER probe highlights many positive lymphocytes ﬈, indicating EBV RNA within nuclei, in this example of EBV gastritis involving the allograft stomach.

443

Intestinal Transplantation

Colon Rejection KEY FACTS ○ Similar to small intestine ACR

ETIOLOGY/PATHOGENESIS • Usually part of multivisceral transplant (MVT) or modified MVT • Colon inclusion does not increase transplant complications

CLINICAL ISSUES • Similar to small bowel rejection ○ Coexists with small intestine acute cellular rejection (ACR) • Stomal output increase • Prognosis usually determined by small intestine ACR severity

MACROSCOPIC

TOP DIFFERENTIAL DIAGNOSES • • • •

EBV colitis and posttransplant lymphoproliferative disorder Viral colitis, non-EBV Clostridium difficile colitis (pseudomembranous colitis) Mycophenolate mofetil colitis ○ Morphologic overlap with ACR

DIAGNOSTIC CHECKLIST • Mucosal ulceration ○ Hallmark of severe rejection • Intimal proliferation of serosal vessels ○ Hallmark of chronic rejection

• Severe exfoliative rejection or chronic rejection causes adhesions and wall thickening

MICROSCOPIC • Crypt apoptosis is hallmark • Rejection graded as mild, moderate, and severe

Colon Pseudoobstruction

Colon Pseudoobstruction: Trichrome

Adenomatous Polyposis Coli

Colonic Apoptoses

(Left) H&E of a colon resection specimen with intestinal pseudoobstruction shows expansion of the submucosa with intact mucosa and irregular thickening ſt and thinning ﬇ of muscularis propria. (Right) Trichrome stain highlights submucosal fibrosis and irregular muscularis propria layers, especially the outer layer, in a patient with intestinal pseudoobstruction.

(Left) A total colectomy specimen is shown here performed in a patient with diffuse colonic polyposis ﬈ with evidence of involvement of small bowel and stomach, one of the indications for multivisceral transplant. (Right) Allograft colon biopsy shows large epithelial apoptoses ſt in 2 crypts without any acute inflammation or cryptitis. Epithelial apoptosis is the hallmark for acute cellular rejection in the colon, similar to the small bowel.

444

Colon Rejection

ANCILLARY TESTS

Indications for Colon Transplantation

Immunohistochemistry

• Long-segment Hirschsprung disease (HSCR) and pseudoobstruction, including megacystis microcolon intestinal hypoperistalsis syndrome • Ischemic disease and short gut syndrome • Inflammatory bowel disease, especially Crohn disease • Familial polyposis and malignancy involving colon or mesocolon • Tufting enteropathy (due to colonic involvement) • Often part of multivisceral transplant ○ Initial reluctance to include colon with small intestine during transplant, as complication rates were very high ○ Colon inclusion does not increase complication rates

• Perform C4d immunostain in suspected AMR

CLINICAL ISSUES Presentation • Similar to small bowel rejection and coexists with small intestine acute cellular rejection (ACR) • Stomal output increase • Watery stools, if stoma closed • Fevers

Prognosis • Prognosis usually determined by small bowel ACR severity ○ Colonic rejection changes do not predict graft outcome • Chronic rejection and severe exfoliative rejection can involve colon • Rare recurrence of primary disease in graft ○ Especially Crohn disease

MACROSCOPIC General Features • Severe exfoliative rejection or chronic rejection causes adhesions and wall thickening • Extensive mucosal sloughing with erythema may be seen

MICROSCOPIC Histologic Features • Crypt apoptosis: Hallmark feature • Increase in lamina propria cellularity • ACR graded as mild, moderate, and severe ○ Similar to small intestine rejection grading ○ Severe rejection characterized by erosions and ulcers • Always look for chronic rejection in serosal vessels ○ Intimal proliferation with luminal narrowing • Antibody-mediated rejection (AMR) has similar features to small bowel ○ Mucosal erosions ○ Capillary dilatation with plump endothelial cells and neutrophils in capillaries ○ Capillary thrombi ○ Correlate with circulating donor specific antibodies • Mucosal regeneration characterized by Paneth cell and small intestinal metaplasia (villous metaplasia)

DIFFERENTIAL DIAGNOSIS EBV Colitis • Lamina propria infiltrate of activated lymphocytes and intraepithelial lymphocytes without crypt apoptosis • EBER highlights diffuse positive nuclei in lamina propria ○ Absence of mass lesion consisting of EBV-positive cells

Intestinal Transplantation

ETIOLOGY/PATHOGENESIS

Posttransplant Lymphoproliferative Disorder • Suspect when aggregates of EBV-positive cells extend/infiltrate into mass lesion

Viral Colitis, Non-EBV • Adenovirus colitis associated with epithelial proliferation and intranuclear inclusions that have smudged appearance ○ Adenovirus immunostain identifies inclusions • CMV colitis often associated with lamina propria neutrophils and epithelial injury including ulceration ○ CMV inclusions usually in endothelial cells

Clostridium difficile (Pseudomembranous) Colitis • Characterized by pseudomembranes composed of sloughed cells and neutrophils • Usually causes acute colitis with neutrophils and ulceration

Mycophenolate Mofetil Colitis • Morphologic overlap with ACR

Graft-vs.-Host Disease • Mimics ACR but involves native colon rather than allograft

Reperfusion Injury • Reactive mucosal changes, regeneration, erosions, hemorrhage, and crypt dropout

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • • • •

Crypt apoptosis, similar to small bowel ACR rejection Graded as mild, moderate, and severe rejection Mucosal ulceration: Hallmark of severe rejection Intimal proliferation of serosal vessels: Chronic rejection

SELECTED REFERENCES 1.

2.

3.

4. 5. 6.

Huard G et al: Comparative incidence of rejection occurring in small intestinal and colonic mucosal biopsies of patients undergoing intestinal transplantation. Histopathology. 69(4):600-6, 2016 Ruiz P: How can pathologists help to diagnose late complications in small bowel and multivisceral transplantation? Curr Opin Organ Transplant. 17(3):273-9, 2012 Selbst MK et al: Spectrum of histologic changes in colonic biopsies in patients treated with mycophenolate mofetil. Mod Pathol. 22(6):737-43, 2009 Kato T et al: Inclusion of donor colon and ileocecal valve in intestinal transplantation. Transplantation. 86(2):293-7, 2008 Sauvat F et al: Factors influencing outcome after intestinal transplantation in children. Transplant Proc. 38(6):1689-91, 2006 Sigurdsson L et al: Anatomic variability of rejection in intestinal allografts after pediatric intestinal transplantation. J Pediatr Gastroenterol Nutr. 27(4):403-6, 1998

445

Intestinal Transplantation

Colon Rejection

Reperfusion Injury, Colon

Reperfusion Injury, Colon

Severe Reperfusion Injury, Colon

Acute Cellular Rejection, Colon

Acute Cellular Rejection, Colon

Moderate Acute Cellular Rejection, Colon

(Left) A portion of colonic mucosa taken immediately post transplant (day 0) shows surface denudation, reactive epithelial injury, and apoptotic debris in crypts ſt, which are features of reperfusion injury. (Right) High-magnification image shows crypt damage with apoptosis ſt and cellular debris in the gland lumen in a day-0 biopsy that is suggestive of moderate reperfusion injury.

(Left) Colon biopsy shows mucosal erosion with exudate, hemorrhage, and necrotic debris as a result of severe reperfusion injury in a day-0 biopsy. (Right) The hallmark of acute cellular rejection in the colon is crypt apoptosis, as illustrated here, with adjacent epithelial cells showing nuclear fragmentation ſt and vacuolation typical of apoptotic cells.

(Left) Colon biopsy shows architectural distortion with lamina propria edema, crypt loss, and isolated apoptosis ﬈ in residual crypts, consistent with moderate acute cellular rejection. (Right) This colon biopsy demonstrates another example of moderate acute cellular rejection, consisting of areas with absent crypts and residual crypts with multiple confluent apoptoses ﬈. Note also the increase of inflammatory infiltrate within the lamina propria.

446

Colon Rejection

Antibody-Mediated Rejection, Colon (Left) Acute severe exfoliative rejection involves this colon with complete sloughing of mucosa ﬈, granulation tissue, absence of residual crypts, and increased inflammatory infiltrates in the lamina propria. (Right) The lamina propria has dilated capillary channels lined by plump endothelial cells ﬊ as well as occasional capillaries containing thrombi in the lumina ſt, which are features that should raise the consideration of antibodymediated rejection.

Antibody-Mediated Rejection, Colon

Intestinal Transplantation

Severe Exfoliative Rejection, Colon

Cytomegalovirus Colitis (Left) C4d immunohistochemical study highlights strong and granular capillary endothelial cell deposition ﬈ in this example of antibody-mediated rejection involving the colon and small bowel (not shown) from a patient with documented circulating donorspecific antibodies. (Right) This colonic biopsy shows an ulcer with granulation tissue and many endothelial cells demonstrating classic Cytomegalovirus inclusions ﬈ in an example of CMV allograft colitis.

EBV Colitis

EBV Colitis, EBER Stain (Left) Colonic biopsy of EBV colitis shows mucosal sloughing with increased lamina propria cellularity mainly due to activated lymphocytes and intraepithelial lymphocytes within crypts ﬈. No apoptosis is noted. (Right) In situ hybridization study for EBER in an allograft colon biopsy shows many positive nuclei ﬈ in an example of EBV colitis. There is no aggregation of EBV-positive cells into a mass lesion that would raise the suspicion of posttransplant lymphoproliferative disorder.

447

Intestinal Transplantation

Graft-vs.-Host Disease, Intestine KEY FACTS ○ Fatal in most cases as difficult to diagnose early

TERMINOLOGY • Process wherein alloreactive T cells in allograft mount cellular immune response against host cells and tissues

ETIOLOGY/PATHOGENESIS • Immunologic mismatch needs to be present between graft and donor • Small bowel transplant associated with potential transfer of large amount of immunogenic cells

CLINICAL ISSUES • Can involve multiple organ systems including ○ Gastrointestinal tract ○ Liver ○ Lungs ○ Skin ○ Bone marrow • Rejection and GVHD usually do not coexist at same time • Poor prognosis

MICROSCOPIC • Native colon or duodenum may show mucosal ulceration and granulation tissue • Hallmark: Residual crypts may show apoptoses • Liver shows bile duct injury with lymphocytes infiltrating ducts • Bone marrow suppression with pancytopenia • Spongiotic dermatitis with ○ Lymphocytic exocytosis ○ Vacuolar degeneration ○ Epithelial lifting ○ Frank ulceration

TOP DIFFERENTIAL DIAGNOSES • Enteric infections • Drug-induced enterocolitis • Acute cellular rejection

Grade 3 GVHD: Skin

Adnexal Involvement in GVHD

Grade 2 GVHD: Skin

GVHD: Esophagus

(Left) Scanning view of a skin biopsy shows a large blister with subepidermal splitting in a small bowel transplant patient. The few adnexa that are present also show some involvement. (Right) Highmagnification view of a skin biopsy shows an adnexal structure with multiple necrotic keratinocytes that have eosinophilic cytoplasm and rounded dark nuclei. Other cells may show apoptoses.

(Left) Skin biopsy shows multiple epithelial apoptoses along the basal layer of the epidermis ﬈ with evidence of early vacuolation at the dermal-epidermal junction ﬊. The superficial dermis shows a mild perivascular lymphocytic infiltrate with some lymphocytes infiltrating the epidermis. (Right) Native esophageal biopsy demonstrates spongiosis and scattered necrotic keratinocytes or dyskeratotic cells ﬈, raising the suspicion for graft-vs.-host disease (GVHD).

448

Graft-vs.-Host Disease, Intestine

Definitions

Endoscopic Findings

• Process wherein alloreactive T cells in allograft mount cellular immune response against host cells and tissues • Acute or chronic ○ Most following small bowel transplant are acute GVHD

• Erythema or small erosions • Deep ulcers in severe cases

Abbreviations

ETIOLOGY/PATHOGENESIS Mechanisms • Immunologic mismatch needs to exist between recipient and donor • Host must not react to allograft for certain time period for graft to mount immune reaction against host • Sufficient amount of lymphoid tissue within allograft must be present to mount response against host • Small bowel transplant associated with transfer of larger amounts of immunogenic cells than other solid organ transplants • Newer tolerogenic protocols result in lower doses of immunosuppressive agents for maintenance ○ Increases potential for GVHD • Normal transplant process results in local mixed lymphocyte chimerism (donor and recipient) that disappears over period of 3 weeks ○ Persistence of chimerism beyond this period may induce immune response against host

CLINICAL ISSUES Site • Can involve multiple organ systems, including ○ Gastrointestinal (GI) tract – Upper GI involvement may manifest as □ Oral and esophageal ulcers □ Gastric erosions or ulcers □ Duodenal ulceration, if severe – Extensive ulceration of native bowel – Does not affect allograft bowel □ No graft dysfunction ○ Liver – May be cholestatic and green □ Without cirrhosis ○ Lungs – May be heavy if involved by diffuse alveolar damage ○ Skin – May show extensive exfoliation in terminal stages ○ Bone marrow

Presentation • Can occur any time after transplantation ○ As early as 1st month • Clinical signs depend on organ involvement and include ○ Nausea, vomiting ○ Diarrhea with hematochezia ○ Jaundice ○ Hemolytic anemia ○ Respiratory distress

Laboratory Tests • Liver enzymes may be elevated with cholestatic pattern ○ Elevation of GGT ○ Elevation of alkaline phosphatase ○ Elevation of bilirubin

Intestinal Transplantation

• Graft-vs.-host disease (GVHD)

○ Fevers ○ Skin rash • Must be differentiated from organ rejection • Rejection and GVHD usually do not coexist at same time

TERMINOLOGY

Treatment • Steroids 1st line, though resistant to conventional immunosuppressive therapy • Graft enterectomy may be only therapeutic option ○ Removes dominant graft lymphocyte population, if done early

Prognosis • Poor ○ Fatal in most cases due to difficulty in early diagnosis

MACROSCOPIC General Features • Unusual to receive specimen ○ Except at autopsy • May show extensive ulceration and hemorrhage in gastrointestinal tract • Liver may appear green due to cholestasis • Lungs may be heavy and boggy due to diffuse alveolar damage

MICROSCOPIC Histologic Features • Native colon or duodenum ○ May show mucosal ulceration and granulation tissue ○ Hallmark: Residual crypts may show apoptoses ○ Lamina propria with mixed inflammation ○ Longstanding involvement may manifest with – Lamina propria fibrosis – Crypt loss □ Feature suggesting chronic GVHD – Epithelial regeneration • Native stomach or esophagus ○ May show erosions or ulcers ○ Glands may show prominent apoptoses ○ Esophagus with – Spongiosis – Epithelial apoptosis – Basal vacuolation and lifting of epithelium – Ulceration • Allograft small bowel ○ May show no evidence of rejection or inflammation ○ May have prominent lymphoid aggregates

449

Intestinal Transplantation

Graft-vs.-Host Disease, Intestine

•

• •

•

– Fluorescent in situ hybridization (FISH) testing of allograft with X and Y chromosome probes may determine donor origin of lymphoid population □ Donor and recipient must be of opposite genders Liver ○ Bile duct injury with lymphocytes infiltrating ducts ○ Progressive damage to duct epithelium with overlapping of epithelial cells and progressive loss of cells ○ Ductopenia, if present for long duration – Unlikely due to rapid progression of GVHD Bone marrow ○ Suppression with pancytopenia, hypocellular marrow Lungs ○ May show features of acute diffuse alveolar damage with hyaline membranes in alveoli Skin involvement, usually as grade III or IV disease ○ Spongiotic dermatitis with lymphocytic exocytosis ○ Vacuolar degeneration ○ Epithelial lifting ○ Frank ulceration ○ Adnexal involvement with apoptoses of adnexal epithelium is common ○ Chronic GVHD manifests by dense dermal sclerosis with morphea-like appearance

DIFFERENTIAL DIAGNOSIS Enteric Infections • Diarrhea suggests bacterial or viral infections, including ○ Clostridium difficile enterocolitis – Abundant neutrophils with pseudomembranes □ Rather than ulceration and crypt apoptoses seen in GVHD ○ Cytomegalovirus (CMV) infection – CMV enteritis shows ulcers with granulation tissue and multiple viral inclusions – Can coexist with GVHD ○ Other bacterial infections

Drug-Induced Enterocolitis • Some, such as mycophenolate mofetil, can mimic histologic changes of GVHD • Crypts may show regenerative changes with presence of apoptoses

Acute Cellular Rejection • Crypt apoptoses are hallmark of acute cellular rejection • Affects allograft and not native bowel

Lung Infections • Any bacterial or fungal pneumonia may be in differential diagnosis ○ Blood cultures and possible bronchoalveolar lavage needed to exclude infectious etiologies • Rapid onset of respiratory failure may preclude investigations for cause

Skin Lesions • Rash can mimic erythema multiforme, viral exanthem, or staphylococcal scalded skin syndrome ○ Typically not associated with adnexal involvement 450

○ May show spongiotic dermatitis with perivascular lymphohistiocytic infiltrate ○ Neutrophilic abscesses in staphylococcal scalded skin syndrome ○ Eosinophils may be prominent if drugs suspected etiology • Blisters may show herpes inclusions if cause

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Multiple organ system involvement post transplantation represents diagnostic clue • Skin involvement is most common ○ With generalized rash and histologic evidence of involvement • Apoptosis and basal vacuolation with involvement of adnexa are clues to skin GVHD • Gastrointestinal tract involvement manifested by ulceration on biopsies • Crypt apoptoses in nonallograft biopsies should prompt diagnosis of GVHD ○ If mycophenolate toxicity excluded • Consider coexisting infections, especially CMV • FISH with X and Y probes should be tested ○ If donor and recipient of different genders • Abnormal cholestatic liver disease should raise suspicion of liver GVHD with bile duct injury • Bone marrow may be hypocellular with pancytopenia • Chronic GVHD unusual in small bowel transplant recipients ○ Due to rapid fatal outcome of acute GVHD

SELECTED REFERENCES 1.

2.

3. 4.

5. 6. 7.

8. 9.

Cromvik J et al: Graft-versus-host disease after intestinal or multivisceral transplantation: A Scandinavian single-center experience. Transplant Proc. 48(1):185-90, 2016 Green T et al: Graft-versus-host disease in paediatric solid organ transplantation: A review of the literature. Pediatr Transplant. 20(5):607-18, 2016 Quirós-Tejeira RE: Immunological complications beyond rejection after intestinal transplantation. Curr Opin Organ Transplant. 17(3):268-72, 2012 Shin CR et al: Incidence of acute and chronic graft-versus-host disease and donor T-cell chimerism after small bowel or combined organ transplantation. J Pediatr Surg. 46(9):1732-8, 2011 Wu G et al: Graft-versus-host disease after intestinal and multivisceral transplantation. Transplantation. 91(2):219-24, 2011 Andres AM et al: Graft-vs-host disease after small bowel transplantation in children. J Pediatr Surg. 45(2):330-6; discussion 336, 2010 Mawad R et al: Graft-versus-host disease presenting with pancytopenia after en bloc multiorgan transplantation: case report and literature review. Transplant Proc. 41(10):4431-3, 2009 Mazariegos GV et al: Graft versus host disease in intestinal transplantation. Am J Transplant. 4(9):1459-65, 2004 Tryphonopoulos P et al: Epithelial and hematopoietic cell chimerism in intestinal allografts. Transplant Proc. 36(2):359-60, 2004

Graft-vs.-Host Disease, Intestine

GVHD: Duodenum (Left) Native stomach biopsy shows extensive gland destruction with apoptotic debris in the lumen ﬈. Note the dense mixed inflammation in the lamina propria and epithelial injury with epithelial lifting and regenerative changes ﬊. (Right) Native duodenum demonstrates crypt apoptosis with necrotic debris in the lumina ﬈ and mixed inflammation in the lamina propria with eosinophils. A single well-formed inclusion of Cytomegalovirus ﬊ is seen beneath the epithelium as a coexisting disease, which should always be considered.

GVHD: Duodenum

Intestinal Transplantation

Severe GVHD: Stomach

GVHD: Liver (Left) Duodenal biopsy shows crypt apoptosis ﬈ as well as necrotic debris in some crypt lumina. The presence of apoptoses in the native bowel is a diagnostic clue. (Right) Liver biopsy from a patient with small bowel transplant reveals portal areas with absence of bile duct profiles accompanying arteries ﬈. Ductopenia can be the end result of GVHD.

GVHD: Liver

Mycophenolate Mofetil Injury: Colon (Left) Cytokeratin 7 stain shows evidence of bile duct injury with irregular contour of bile ducts with no lumen ſt and irregular placement of nuclei of biliary epithelial cells. Note occasional lymphocytes abutting and infiltrating bile ducts ﬈. (Right) Colonic biopsy shows mucosal erosions with increased lamina propria cellularity and crypt injury with neutrophils, eosinophils, and necrotic debris in lumen. The crypt shows marked regenerative changes besides the damage seen on one side.

451

Intestinal Transplantation

Bacterial and Fungal Infections KEY FACTS

ETIOLOGY/PATHOGENESIS

MICROSCOPIC

• Most common infectious agents ○ Enterococcus ○ Staphylococcus ○ Enterobacter ○ Klebsiella • Clostridium difficile infection may manifest with pseudomembranous colitis • Severe systemic fungal infections are usually fatal ○ Causative agents include – Aspergillus – Candida – Less commonly Cryptococcus, Zygomycota, and Pneumocystis

• Usually manifest with increased lamina propria cellularity • Infiltrate composed of neutrophils predominantly but also lymphocytes and plasma cells • Villi appear swollen and may contain inflammatory cells • Pseudomembrane composed of fibrin and cellular debris, including neutrophils, may be evident in rare instances of unsuspected C. difficile infection • Presence of granulomas should warrant search for acid-fast bacilli with cultures and PPD testing • Fungal infections never seen in intestinal biopsies

CLINICAL ISSUES

TOP DIFFERENTIAL DIAGNOSES • Viral infections • Acute cellular rejection • Antibody-mediated rejection

• Can present at any time post transplant (days to weeks)

Bacterial Enteritis

Bacterial Enteritis

Zygomycetes Infection

Zygomycetes Infection

(Left) Ileal biopsy shows villous alterations with flattening and blunting ﬈. The lamina propria shows increased cellularity with a mixed infiltrate. These are features of bacterial enteritis. (Right) High-magnification view shows epithelial injury with vacuolation, intraepithelial neutrophils ﬈, and mixed infiltrate in the lamina propria ﬊. No viral inclusions are seen.

(Left) Image shows the presence of pale, broad hyphal forms of fungi that are aseptate ﬈. The presence of these structures on H&E staining should prompt a diagnosis of Zygomycetes infection. (Right) A Grocott (GMS) stain will not show the same staining of the hyphae as may be seen with Aspergillus or Candida. The hyphae appear as pale, negativestaining structures ﬈ that do not take up the silver stain. They may be easily missed.

452

Bacterial and Fungal Infections

Definitions • Patient with fevers and increased output following small bowel or multivisceral transplant ○ Confirmed by bacterial and fungal cultures or Clostridium difficile toxin documentation

ETIOLOGY/PATHOGENESIS Infectious Agents • Bacterial infections ○ Results from prolonged immunosuppression ○ May also be effect of bacterial translocation following reperfusion injury – Most common infectious agents: Enterococcus, Staphylococcus, Enterobacter, and Klebsiella ○ C. difficile infection may manifest with pseudomembranous colitis ○ Rare instances of Escherichia coli infection manifesting as malakoplakia • Fungal infections ○ Usually result of severe immunosuppression, e.g., following bouts of rejection ○ May be in form of oral thrush or candidal esophagitis ○ Causative agents – Aspergillus and Candida – Less commonly Cryptococcus, Zygomycota, and Pneumocystis ○ Severe systemic fungal infections usually fatal

CLINICAL ISSUES Presentation • Can present any time post transplant (days to weeks) • Multiple episodes may be seen • Symptoms include ○ Fevers ○ Irritability ○ Malaise ○ Decreased appetite ○ Wound dehiscence or drainage – Wound infection is early and important complication that can cause multiple immediate surgical issues ○ Increased stomal output • More systemic infections can result in signs of sepsis and multiorgan dysfunction • Systemic fungal infections may present with pulmonary symptoms, CNS symptoms, or severe abdominal symptoms, such as pain, evidence of peritonitis, increased stomal output, or discharge from wound • Line infections are frequent sources of systemic bacterial and fungal infections

Treatment • Systemic antibiotics and antifungals after cultures and sensitivity testing • Response to antifungals may not be optimal ○ May necessitate withdrawal of immune suppressive medications • Sometimes surgical resection of affected bowel or stoma

• Malakoplakia may warrant withdrawal of immune suppressive drugs

Prognosis • Usually good for bacterial and most candidal infections • Systemic fungal infections always cause death if widespread and not treated aggressively

IMAGING Radiographic Findings

Intestinal Transplantation

TERMINOLOGY

• Abdominal radiographs may show evidence of peritonitis • Lungs may show evidence of pneumonia in systemic infections • Rarely, fungal infections may manifest as lung abscess, seen as cystic lung lesions • Vascular thrombosis due to infected grafts or line infections may show abnormal vascular flow patterns

MACROSCOPIC General Features • Unusual to get resections for bacterial or fungal infections • Rare instances of fungal infections of stoma site can be resected ○ Show necrosis and ulceration of mucosa • Segmental or diffuse ulceration may be seen in severe infections in allograft ileum or colon if resected

MICROSCOPIC Histologic Features • Bacterial infections ○ Usually manifest with increased lamina propria cellularity – Infiltrate composed of neutrophils predominantly – Also lymphocytes and plasma cells ○ Villi appear swollen and may show inflammatory cells ○ Neutrophils frequently infiltrate epithelium with damage to epithelial cells – Scattered neutrophils in lamina propria – Warrants stool cultures and work-up for bacterial infections ○ Erosions or ulcers may be evident, though unusual, except for C. difficile – Pseudomembrane composed of fibrin and cellular debris, including neutrophils, may be evident in rare instances of unsuspected C. difficile infection ○ Biopsy of blind loops may frequently have neutrophils due to bacterial colonization – Crypt apoptosis suggests coexisting rejection process and is not feature of colonization – Bacterial colonization on surface epithelium may be evident ○ Mycobacterial infections uncommon but may be important depending on patient demographic – Presence of granulomas warrants search for acid-fast bacilli with cultures and PPD testing • Fungal infections ○ Not identified in allograft intestinal biopsies ○ Esophageal candidiasis may be evident on upper endoscopy in intestinal transplant patients

453

Intestinal Transplantation

Bacterial and Fungal Infections ○ Fungal infection of graft invariably associated with fatal outcome and diagnosed at autopsy ○ Early wound infection post transplant may have fungal etiology, especially Candida species ○ Wound debridement should warrant silver stains to look for fungal organisms ○ Can cause vascular thrombi and graft ischemic injury following wound infections ○ Fungal peritonitis manifests with serosal exudate with scant inflammation but with many fungal organisms ○ Autopsy may show fungal organisms in other organs, such as lungs and heart valves as vegetations ○ Autopsy fungal culture is mandatory for confirmation

Cytologic Features • Ascitic fluid rarely obtained for cytology ○ May show abundant neutrophils if peritonitis is cause ○ Warrants Gram and Grocott stain for bacterial and fungal organisms ○ Fluid should always be sent for microbiologic cultures

ANCILLARY TESTS PCR • Testing for 16s ribosomal RNA or PCR for fungal typing • PCR for mycobacteria may be necessary for confirmation

DIFFERENTIAL DIAGNOSIS Viral Infections • Cytomegalovirus (CMV) and adenovirus can mimic bacterial infections with neutrophils in lamina propria • Presence of viral inclusions for CMV in endothelial cells and adenovirus in epithelial cells would clinch diagnosis • Rotavirus can also cause increased lamina propria cellularity and reactive epithelial changes but no viral inclusions • Stool cultures for rotavirus and adenovirus helpful and viral PCR for CMV • Immunohistochemistry for CMV and adenovirus also help confirm diagnosis

Acute Cellular Rejection • Acute cellular rejection (ACR) should always be looked for in allograft intestinal biopsies • Can coexist with infections • Important to recognize, since treatment strategies would have to be balanced • Can cause increased lamina propria cellularity, including neutrophils and eosinophils • Crypt apoptosis is hallmark of ACR

Antibody-Mediated Rejection • Antibody-mediated rejection (AMR) usually evident in early period post transplant, when infections may coexist due to immune suppression • Can cause dilated and congested capillaries with neutrophil margination and fibrin thrombi • Neutrophils may be present in lamina propria • Severe cases can cause ulceration mimicking infections • C4d stain and correlation with circulating donor-specific antibodies helpful in diagnosis of AMR

454

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Presence of erosions or ulcers on endoscopy should warrant search for infectious etiology • Visible endoscopic exudates may be due to infection • Histological evidence of increased neutrophils in lamina propria or within epithelium should raise suspicion • Biopsy unusual for bacterial infections ○ Patient is treated with empirical antibiotics or based on stool cultures without biopsy • Need to differentiate from viral etiology with search for viral inclusions • ACR can coexist with infections

Pathologic Interpretation Pearls • Neutrophils in lamina propria • Ulcers with pseudomembrane formation should warrant correlation with C. difficile toxin study • Cryptitis not usual feature of bacterial infections ○ Should warrant search for other etiologies, such as viruses or ACR • Presence of granulomas should warrant search for mycobacteria, unusual in intestinal biopsies • Fungal infections cause severe inflammation with deep ulcers and peritonitis • Wound infections can result in peritonitis and dissemination of fungal infections • Any early vascular disease, such as thrombosis, should warrant search for fungi as cause of thrombi

SELECTED REFERENCES 1.

Suhr MJ et al: Epidemiological investigation of Candida species causing bloodstream infection in paediatric small bowel transplant recipients. Mycoses. 60(6):366-374, 2017 2. Silva JT et al: Infectious complications following small bowel transplantation. Am J Transplant. 16(3):951-9, 2016 3. Timpone JG Jr et al: Infections in intestinal and multivisceral transplant recipients. Infect Dis Clin North Am. 27(2):359-77, 2013 4. Akhter K et al: Six-month incidence of bloodstream infections in intestinal transplant patients. Transpl Infect Dis. 14(3):242-7, 2012 5. Florescu DF et al: Bloodstream infections during the first year after pediatric small bowel transplantation. Pediatr Infect Dis J. 31(7):700-4, 2012 6. Florescu DF et al: Incidence and outcome of fungal infections in pediatric small bowel transplant recipients. Transpl Infect Dis. 12(6):497-504, 2010 7. Pappas PG et al: Invasive fungal infections among organ transplant recipients: results of the Transplant-Associated Infection Surveillance Network (TRANSNET). Clin Infect Dis. 50(8):1101-11, 2010 8. Guaraldi G et al: Outcome, incidence, and timing of infectious complications in small bowel and multivisceral organ transplantation patients. Transplantation. 80(12):1742-8, 2005 9. Cicalese L et al: Bacterial translocation in clinical intestinal transplantation. Transplantation. 71(10):1414-7, 2001 10. Kusne S et al: Infectious complications after small bowel transplantation in adults. Transplant Proc. 26(3):1682-3, 1994 11. Reyes J et al: Infectious complications after human small bowel transplantation. Transplant Proc. 24(3):1249-50, 1992 12. Shabtai M et al: Malakoplakia in renal transplantation: an expression of altered tissue reactivity under immunosuppression. Transplant Proc. 21(4):3725-7, 1989

Bacterial and Fungal Infections

Clostridium difficile Infection (Left) An important clue to the diagnosis of enteritis is the lack of significant crypt injury, as seen here. No crypt apoptosis or intraepithelial neutrophils are seen. The lamina propria, however, shows increased cellularity. (Right) Low-magnification view of a small bowel allograft biopsy shows ulceration and granulation tissue ſt and a superficial exudate with cellular debris constituting a pseudomembrane ﬇. This should raise suspicion of C. difficile infection.

C. difficile Enteritis

Intestinal Transplantation

Acute Enteritis

C. difficile Enteritis (Left) Small intestinal biopsy shows dilated capillaries with neutrophils ﬈ in the lamina propria as well as foci of intraepithelial neutrophilic infiltration ﬊ in an example of C. difficile enteritis. (Right) High-magnification view of a pseudomembrane ﬈ shows fibrinous material, abundant neutrophils, mucinous material, cellular debris, and no significant bacterial colonies, all features of C. difficile infection.

Fungal Infection

Fungal Infection (Left) A necrotic segment of a small bowel allograft removed 1 month after surgery due to wound infection shows regenerative crypts with hemorrhage and fibrin in lamina propria. Fungal organisms are difficult to identify but should be looked for. (Right) Grocott stain highlights fungal hyphae that appear to be septate and narrow with branching ﬈, suggestive of Aspergillus species. Wound cultures were positive for Aspergillus.

455

Intestinal Transplantation

Adenovirus, Intestine KEY FACTS

ETIOLOGY/PATHOGENESIS • Different serotypes ○ Types 40 and 41 associated with GI disease • Disseminated disease in immunosuppressed patients

CLINICAL ISSUES • More common in pediatric transplant patients • Infection usually occurs ~ 25 days post transplant ○ Peak period extending to ~ 45 days • Increased stomal output • Usually self-limited

MICROSCOPIC

• Eosinophilic intranuclear inclusions with peripheral aggregation of chromatin along nuclear membrane and nuclear enlargement

ANCILLARY TESTS • Adenovirus immunostain sensitive in picking up infected cells and inclusions • Stool cultures confirm diagnosis of adenovirus enteritis

TOP DIFFERENTIAL DIAGNOSES • Other bacterial or viral infections • Acute cellular rejection • Reperfusion/ischemic injury

• Villous architecture may be altered with blunting and flattening of villi ○ Characteristic "piling up" or stratification of epithelial nuclei on surface • Viral infection causes increased nuclear basophilia with smudged appearance

Small Bowel Adenovirus Infection

Adenovirus Enteritis

Smudged Inclusions

Intranuclear Epithelial Inclusions

(Left) Low magnification of a small bowel allograft biopsy shows preserved villi with some loss of goblet cells and increased lamina propria cellularity. Biopsy may look normal or enteritic. (Right) Adenovirus immunohistochemistry is useful in highlighting the adenoviral nuclear inclusions ﬈ that could be easily missed by light microscopy.

(Left) High magnification shows the inclusions of Adenovirus in surface epithelial cell nuclei ſt with peripheral arrangement of chromatin and central nuclear homogeneous eosinophilic inclusion. (Right) Adenovirus immunostain shows many intraepithelial nuclear inclusions ſt with stratification and enlargement of nuclei.

456

Adenovirus, Intestine

IMAGING

Definitions

Radiographic Findings

• Infectious enteritis characterized by epithelial injury due to adenoviral infection, resulting in fevers and diarrhea or increased stomal output

• GI disease usually with no significant imaging findings • Pulmonary infection may show evidence of pneumonia

ETIOLOGY/PATHOGENESIS Adenovirus • Double-stranded DNA virus • Many serotypes ○ Serotypes 40 and 41 associated with gastrointestinal (GI) disease ○ Other subtypes cause pulmonary disease • Disseminated disease in immunosuppressed patients

CLINICAL ISSUES Epidemiology • Incidence ○ Infection usually occurs ~ 25 days post transplant – Peak period extending to ~ 45 days ○ Can occur anytime after transplantation ○ Seasonal infections and in clusters, such as transplant inpatient units or schools • Age ○ More common in pediatric transplant patients – Incidence ranges 9-50% in pediatric age group – Younger age at transplantation is possible risk factor

Site • • • •

Allograft intestine Native jejunum or colon Hepatic involvement may be seen Rare instances of small bowel involvement as part of systemic infection

Presentation • Fevers • Diarrhea • Increased stomal output ○ Severe disseminated infections may present with pneumonia and tracheobronchitis ○ Hepatic involvement may be associated with elevated enzymes, including GGTP, depending on type of infection – Ascending cholangiopathy vs. systemic disease with necrotic parenchymal foci

Treatment • Conservative management with modification of immunosuppression usually sufficient • Severe infections may require cidofovir or other antiviral agents

Prognosis • Usually self-limited • Can recur seasonally or in hospital setting due to clustering • Disseminated disease can lead to death ○ Rare cause of death in children

MACROSCOPIC Autopsy Organ Findings • GI tract ○ Extensive mucosal ulcerations • Lungs ○ May be consolidated with hemorrhagic, necrotic foci • Liver ○ May show congestion or foci of necrosis

Intestinal Transplantation

TERMINOLOGY

MICROSCOPIC Histologic Features • Increased cellularity in lamina propria with lymphocytes, plasma cells, and neutrophils • Blunting and flattening of villous architecture • Characteristic "piling up" or stratification of epithelial nuclei on surface • Reactive surface enterocytes ○ May be mistaken for regenerative mucosa • Increased basophilia of nuclei with smudged appearance • Eosinophilic intranuclear inclusions with peripheral aggregation of chromatin along nuclear membrane and nuclear enlargement ○ Often incidentally discovered ○ Requires high index of suspicion to identify – Frequently missed ○ Can be few to many inclusions ○ Can progressively increase in serial biopsies ○ Epithelial shedding on surface with inclusions ○ Rare instances of inclusions in stromal cells in early infection • Crypt involvement unusual except in overwhelming infections ○ Crypt apoptosis rarely seen – Often restricted to surface epithelium and superficial aspects of crypts ○ May predominantly involve crypts in hematogenous infections with associated crypt injury and apoptosis • Mucosal ulcerations are rare but seen in severe infections ○ Can be mistaken for rejection

ANCILLARY TESTS Immunohistochemistry • Adenovirus ○ Sensitive for detecting adenovirus-infected cells and inclusions – Sensitive even in autopsy material to determine disease dissemination ○ Prior biopsies with diagnosis of adenoviral enteritis should be immunostained to establish duration of disease

In Situ Hybridization • Adenovirus 457

Intestinal Transplantation

Adenovirus, Intestine

PCR • Testing available on stool specimen or blood • Does not differentiate acute infection from latent infection • Positive blood cultures harbinger of disseminated disease and needs aggressive treatment

Microbiology • Stool cultures confirm diagnosis of adenovirus enteritis • Positive blood cultures suggest disseminated disease

DIFFERENTIAL DIAGNOSIS Bacterial Infections • May have same inflammatory component and biopsy appearance as adenovirus • Bacterial infections usually diagnosed by stool cultures ○ May show surface exudate and pseudomembrane ○ No inclusions

Viral Infections • May have same inflammatory component and biopsy appearance as adenovirus • Presence of intraepithelial inclusions is pathognomonic • Epithelial piling is diagnostic clue • Rotavirus looks similar but without inclusions ○ Diagnosis usually based on stool cultures ○ Seasonal • CMV inclusions usually in endothelial cells and not epithelial cells ○ Usually associated with ulceration and granulation tissue ○ CMV may coexist with adenovirus as it colonizes ulcerated mucosa • Calicivirus infection rarely reported in pediatric patients ○ Can show piling up of epithelial cells and villous alterations ○ No inclusions are usually noted ○ Lamina propria with mainly lymphoplasmacytic infiltrate – No neutrophils ○ Diagnosis established by PCR testing

Acute Cellular Rejection

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Adenoviral enteritis typically more common in pediatric age group ○ Rare in adult small bowel transplantation • Infection can coexist with other infections or with ACR • Large number of inclusions suggest greater viral load ○ May warrant antiviral therapy • Lack of immunosuppression modulation may lead to disseminated disease ○ Would manifest with progressive enteritis that persists through several sequential biopsies • Recurrence of adenoviral infections can occur seasonally

Pathologic Interpretation Pearls • Villous alterations warrant search for infectious etiology • Neutrophils in lamina propria may indicate infection • Piling up of surface epithelial cells should prompt search &/or immunostain for viral inclusions • Light microscopy easily identifies inclusions except when rare in early infection • Characteristic smudged cells or intranuclear inclusions • Crypt apoptosis suggests coexisting ACR • Disseminated infection (hematogenous) may show mainly crypt injury and inclusions

SELECTED REFERENCES 1.

2. 3. 4. 5. 6. 7.

• Mixed inflammatory cell infiltrate noted in both • Neutrophils less likely in acute cellular rejection (ACR) unless ulceration present • Absence of classic intranuclear inclusions of adenoviral enteritis • Crypt apoptosis, including multiple large apoptoses, diagnostic of ACR ○ ACR can coexist with infection in same biopsy ○ Adenovirus immunostain confirms viral infection in suspected cases

8.

Reperfusion/Ischemic Injury

14.

• Usually early event in course of transplantation ○ Most often seen in biopsies within first 2 weeks, as opposed to later presentation (1 month) for adenovirus • Reactive-appearing, small epithelial cells common to both • More hemorrhage and crypt necrosis/apoptosis in ischemic injury • Eosinophilic nuclei of epithelial cells due to cell injury 458

○ Negative immunostain for adenovirus excludes viral infection in difficult cases

○ Technically challenging compared to immunohistochemistry

9.

10. 11.

12. 13.

Mehta V et al: Adenovirus disease in six small bowel, kidney and heart transplant recipients; pathology and clinical outcome. Virchows Arch. 467(5):603-8, 2015 Timpone JG Jr et al: Infections in intestinal and multivisceral transplant recipients. Infect Dis Clin North Am. 27(2):359-77, 2013 Florescu DF et al: Adenovirus infections in pediatric small bowel transplant recipients. Transplantation. 90(2):198-204, 2010 Adeyi OA et al: Posttransplant adenoviral enteropathy in patients with small bowel transplantation. Arch Pathol Lab Med. 132(4):703-5, 2008 Hoffman JA: Adenoviral disease in pediatric solid organ transplant recipients. Pediatr Transplant. 10(1):17-25, 2006 Ozolek JA et al: Adenovirus infection within stromal cells in a pediatric small bowel allograft. Pediatr Dev Pathol. 9(4):321-7, 2006 Humar A et al: A surveillance study of adenovirus infection in adult solid organ transplant recipients. Am J Transplant. 5(10):2555-9, 2005 Morotti RA et al: Calicivirus infection in pediatric small intestine transplant recipients: pathological considerations. Hum Pathol. 35(10):1236-40, 2004 McLaughlin GE et al: Adenovirus infection in pediatric liver and intestinal transplant recipients: utility of DNA detection by PCR. Am J Transplant. 3(2):224-8, 2003 Pinchoff RJ et al: Adenovirus infection in pediatric small bowel transplantation recipients. Transplantation. 76(1):183-9, 2003 Kaufman SS et al: Discrimination between acute rejection and adenoviral enteritis in intestinal transplant recipients. Transplant Proc. 34(3):943-5, 2002 Parizhskaya M et al: Enteric adenovirus infection in pediatric small bowel transplant recipients. Pediatr Dev Pathol. 4(2):122-8, 2001 Berho M et al: Adenovirus enterocolitis in human small bowel transplants. Pediatr Transplant. 2(4):277-82, 1998 Levy MF et al: Adenovirus infection of the human intestinal allograft: a case report. Transplant Proc. 28(5):2786-7, 1996

Adenovirus, Intestine

Adenovirus Enteritis (Left) The classic morphologic features of an ileal biopsy include marked reactive surface epithelial changes with piling up of epithelial cell nuclei ﬊ and presence of eosinophilic to amphophilic intranuclear inclusions ﬈. This change should not be mistaken for regenerative bowel mucosa. (Right) High magnification of epithelium shows numerous intranuclear inclusions ﬈ giving a smudged appearance to the cell nucleus. Also note the piling up of nuclei.

Inclusions of Adenovirus

Intestinal Transplantation

Adenovirus Enteritis

Adenovirus Stain (Left) Another example of a small bowel allograft shows many intraepithelial neutrophils ﬇ that may lead to a misdiagnosis of bacterial infection if the intranuclear inclusions ſt are not identified. (Right) Low magnification of an adenoviral immunostain highlights the numerous inclusions ﬈ in the surface epithelium that may extend into the crypts if abundant. Typically, inclusions are only seen in surface epithelium or shed cells in lumen.

Crypt Involvement in Disseminated Infection

Predominant Crypt Inclusions (Left) High magnification shows crypt disruption with piling of cells, prominent apoptoses ſt, and smudged nuclei ﬈. This patient died of pneumonia with extensive lung involvement by adenovirus. (Right) Adenovirus immunostain confirms extensive crypt involvement illustrated in the previous image. Note the extensive crypt damage and numerous inclusions ſt not readily evident on H&E stain.

459

Intestinal Transplantation

Rotavirus, Cytomegalovirus, and Herpes Simplex Virus KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Cytomegalovirus (CMV) • Herpes simplex virus (HSV)

• Rotavirus infection ○ Mild to moderate villous blunting ○ Increased lamina propria and surface epithelial cellularity • CMV infection ○ Intraepithelial neutrophils in surface epithelium ○ Erosions ○ Ulcers ○ Plump endothelial cells in lamina propria vessels with CMV inclusions ○ Inclusions are classically intranuclear and cytoplasmic ○ Correlation with PCR or immunohistochemistry helpful • HSV infection ○ Cause ulceration of mucosa ○ Eosinophilic intranuclear inclusions or multinucleated cells with nuclear molding

CLINICAL ISSUES • Rotavirus: Any time post transplant, including months or years ○ Most common cause of diarrhea in children – Self-limited • CMV: Most common cause of morbidity and mortality ○ Increased stomal output ○ Diarrhea ○ Fevers • HSV: Least common viral infection following small bowel transplant ○ Can manifest as – Skin rash – Oral lesions – Esophagitis with ulcers

Rotavirus Infection

Rotavirus Enteritis

Rotavirus Infection

Rotavirus Enteritis

(Left) Small intestine allograft biopsy shows rotaviral enteritis with villous blunting, reactive epithelial changes ﬈, and increased lamina propria cellularity. No crypt apoptosis is seen. (Right) Highmagnification view of rotaviral enteritis shows the absence of epithelial inclusions and presence of mixed infiltrate with neutrophils ſt in the lamina propria. Note the epithelial piling ﬇.

(Left) Allograft biopsy shows expanded lamina propria with mixed infiltrate and villous blunting, a clue to rotavirus infection. The crypt architecture is maintained, and no crypt apoptoses are appreciated at this magnification. The surface epithelium shows loss of goblet cells. (Right) Allograft biopsy shows reactive changes of surface epithelium in rotaviral enteritis. The lamina propria is expanded with a mixed infiltrate that includes lymphocytes, plasma cells, and neutrophils.

460

Rotavirus, Cytomegalovirus, and Herpes Simplex Virus

Abbreviations • Cytomegalovirus (CMV) • Herpes simplex virus (HSV)

ETIOLOGY/PATHOGENESIS

• HSV ○ Skin rash – Examination of skin blister base an easy technique for diagnosis ○ Oral lesions or esophagitis with ulcers ○ Fevers ○ Difficulty swallowing

Infectious Agents

Treatment

• Rotavirus ○ Double-stranded RNA virus ○ Reoviridae family • CMV ○ DNA virus of Herpesviridae family • HSV1 and HSV2 ○ Herpesviridae family

• Drugs ○ Rotavirus – Usually self-limited and treated with conservative management ○ CMV – Ganciclovir – Decrease immune suppression ○ HSV – Acyclovir – Decrease immune suppression ○ ACR can follow or precede viral infections – Over-treatment of rejection may lead to dissemination of CMV or HSV

CLINICAL ISSUES Epidemiology • Incidence ○ Rotavirus – Any time post transplant, including months or years – Seasonal variation – No specific known relationship to transplant immune suppression ○ CMV – Decreased incidence due to prophylaxis and PCR monitoring – Most common cause of morbidity and mortality – Can occur any time post transplant – Occurs after episodes of acute cellular rejection (ACR) – Increased risk if seropositive donor and seronegative recipient ○ HSV – Least common viral infections following small bowel transplant – More common after bone marrow transplantation – Unusual to involve allograft □ Almost never diagnosed on biopsies • Age ○ Rotavirus – Common childhood presentation □ Also occur in adults ○ CMV – More common in pediatric patients

Presentation • Rotavirus ○ Diarrhea – Most common cause in children, but self-limited – No generalized systemic symptoms ○ Infection confers strong immunity that may immunize children for life • CMV ○ Increased stomal output, diarrhea, fevers ○ Systemic infections manifest as hepatitis, pneumonitis, pancreatitis, encephalitis, and retinitis – Usually fatal ○ Presentation may mimic ACR and hence important to exclude

Intestinal Transplantation

TERMINOLOGY

Prognosis • Rotavirus self-limited • CMV usually responsive to therapy ○ Can persist for some time especially with concomitant ACR • HSV responsive to antivirals ○ Can be fatal if systemic

MICROSCOPIC Histologic Features • Rotavirus infection ○ Mild to moderate villous blunting ○ Increased lamina propria and surface epithelial cellularity – Infiltrate includes plasma cells, lymphocytes, eosinophils, and neutrophils ○ Reactive surface epithelial appearance with intraepithelial neutrophil loss of goblet cells ○ Crypt apoptosis usually absent – Apoptosis suggests coexisting rejection ○ No viral inclusions • CMV infection ○ Early allograft infections cause increased lamina propria cellularity with lymphocytes and plasma cells – May show very few inclusions or transformed cells and may be missed ○ Acute infection with neutrophils in lamina propria ○ Intraepithelial neutrophils on surface, erosions, or ulcers ○ Plump endothelial cells in lamina propria vessels with CMV inclusions – Inclusions are classically intranuclear and cytoplasmic – Intranuclear inclusions appear eosinophilic to basophilic surrounded by halo with peripheral marginating chromatin – Coarse and granular cytoplasmic inclusions appear eosinophilic (initially) to basophilic (later) ○ Rare crypt apoptoses may be present – Do not meet criterion for ACR 461

Intestinal Transplantation

Rotavirus, Cytomegalovirus, and Herpes Simplex Virus ○ Correlation with PCR or immunohistochemistry helpful ○ Rare CMV involvement of donor intestine – 1st biopsy may reveal inclusions • HSV infection ○ Causes ulceration of mucosa ○ Infiltrate usually mixed with many neutrophils in granulation tissue ○ Inclusions seen exclusively in surface epithelial cells – Eosinophilic intranuclear inclusions or multinucleated cells with nuclear molding

ANCILLARY TESTS Immunohistochemistry • CMV or HSV IHC sensitive for identifying inclusions or infection

PCR • Sensitive blood test to determine active infection or latency

Microbiology • Direct or indirect immunofluorescent antigen testing may be good screening test • ELISA test for rotavirus • Viral cultures can be used but are cumbersome

DIFFERENTIAL DIAGNOSIS Other Infections • Overlap of histology with other infections ○ Bacterial ○ Fungal ○ Other viral infections • Usually no exudate with viral infections, so bacterial infections ruled out • Presence of typical inclusions of CMV or HSV clinch diagnosis • Rotavirus can mimic adenovirus infection but lacks epithelial inclusions of adenovirus

• CMV inclusions may persist for long time in allograft biopsies ○ Despite treatment and decreasing immune suppression • Knowledge of donor and recipient CMV status critical ○ Especially in children

Pathologic Interpretation Pearls • • • •

Typical viral inclusions in CMV and HSV infections CMV infects endothelial and stromal cells HSV infects only epithelial cells Early CMV infections that only show rare transformed cells with cytomegaly require immunohistochemistry ○ Enlarged endothelial cells may reveal more inclusions in serial sections • CMV can frequently colonize ulcerated mucosa and prevent healing ○ Any ulcer should warrant search for CMV • Rotaviral infection can be easily missed on biopsy unless stool testing reported as positive ○ Seasonal testing for rotavirus helpful in this diagnosis

SELECTED REFERENCES 1.

2. 3.

4.

5.

6.

7. 8.

Acute Cellular Rejection • All infections can mimic ACR • Crypt apoptoses not characteristic of viral infections • Ulcerated mucosa makes it difficult to evaluate for coexisting rejection ○ Presence of crypt apoptoses helpful • CMV inclusions sometimes persist on allograft biopsies even after PCR has normalized ○ May cause confusion in interpretation of ACR • Important to distinguish because ACR treatment is direct opposite of viral infection ○ Treating ACR aggressively can lead to viral dissemination

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Viral infections occur any time post transplant • Viral infections can precede, occur simultaneously, or follow ACR episodes • Increased crypt apoptoses suggests ACR • Clinical monitoring of viral PCR for CMV or HSV critical in following patients off prophylaxis ○ Especially during rejection episodes 462

9. 10.

11. 12. 13.

14.

Bonatti H et al: Use of cidofovir for cytomegalovirus disease refractory to ganciclovir in solid organ recipients. Surg Infect (Larchmt). 18(2):128-136, 2017 Timpone JG et al: Resistant cytomegalovirus in intestinal and multivisceral transplant recipients. Transpl Infect Dis. 18(2):202-9, 2016 Troxell ML et al: Practical applications in immunohistochemistry: evaluation of rejection and infection in organ transplantation. Arch Pathol Lab Med. ePub, 2016 Florescu DF et al: Incidence, risk factors, and outcomes associated with cytomegalovirus disease in small bowel transplant recipients. Pediatr Transplant. 16(3):294-301, 2012 Adeyi OA et al: Rotavirus infection in adult small intestine allografts: a clinicopathological study of a cohort of 23 patients. Am J Transplant. 10(12):2683-9, 2010 Petrisli E et al: Early and late virological monitoring of cytomegalovirus, Epstein-Barr virus, and human herpes virus 6 infections in small bowel/multivisceral transplant recipients. Transplant Proc. 42(1):74-8, 2010 Talmon GA: Histologic features of cytomegalovirus enteritis in small bowel allografts. Transplant Proc. 42(7):2671-5, 2010 Eisengart LJ et al: Rotavirus infection in small bowel transplant: a histologic comparison with acute cellular rejection. Pediatr Dev Pathol. 12(2):85-8, 2009 Stelzmueller I et al: Rotavirus enteritis in solid organ transplant recipients: an underestimated problem? Transpl Infect Dis. 9(4):281-5, 2007 Pascher A et al: CMV, EBV, HHV6, and HHV7 infections after intestinal transplantation without specific antiviral prophylaxis. Transplant Proc. 36(2):381-2, 2004 Ziring D et al: Infectious enteritis after intestinal transplantation: incidence, timing, and outcome. Transplant Proc. 36(2):379-80, 2004 Kaufman SS et al: Calicivirus enteritis in an intestinal transplant recipient. Am J Transplant. 3(6):764-8, 2003 Tzakis AG: Cytomegalovirus prophylaxis with ganciclovir and cytomegalovirus immune globulin in liver and intestinal transplantation. Transpl Infect Dis. 3 Suppl 2:35-9, 2001 Bueno J et al: Cytomegalovirus infection after intestinal transplantation in children. Clin Infect Dis. 25(5):1078-83, 1997

Rotavirus, Cytomegalovirus, and Herpes Simplex Virus

Herpes Simplex Infection (Left) An oral mucosal biopsy shows ulceration and granulation tissue with surface necrosis and occasional multinucleate cells ﬈ suggestive of herpes viral inclusions. (Right) An HSV 1/2 immunohistochemical stain shows positive staining of surface cells ſt and some serum staining, a frequent finding in herpes infection.

Acute Enteritis

Intestinal Transplantation

Herpes Simplex Infection

CMV Enteritis (Left) Allograft biopsy shows extensive ulceration with granulation tissue and fibrinopurulent exudate. This should prompt a search for viral inclusions, especially CMV. (Right) Highmagnification view of granulation tissue demonstrates an endothelial cell with nuclear and cytoplasmic granular eosinophilic inclusions of CMV ﬈. Inclusions may be few or many depending on the stage. Note the mixed inflammatory infiltrate with many neutrophils.

CMV Infection

CMV Enteritis (Left) Another biopsy reveals better preserved mucosa with no ulceration but with some increase in lamina propria cellularity and a single distinct large cell with an intranuclear inclusion of CMV ﬈. (Right) Immunohistochemical stain for CMV is extremely helpful in cases of recent infections as it highlights many small nontransformed cells ﬈ that would not be recognizable on H&E. This stain detects the early antigen.

463

Intestinal Transplantation

Epstein-Barr Virus, Intestine KEY FACTS

ETIOLOGY/PATHOGENESIS • Common complication post transplant

CLINICAL ISSUES • Epstein-Barr virus (EBV) affects small bowel allograft, causing enteritis • Monitoring of EBV PCR greatly reduces this complication • Treatment by decreasing immune suppression • Posttransplant lymphoproliferative disease (PTLD) may need chemotherapy, depending on subtype

MACROSCOPIC • Graft resection usually done for PTLD masses but not for EBV enteritis • Masses may ulcerate mucosa and extend transmurally

MICROSCOPIC

• Lymphocytes can appear enlarged and very activated with prominent nucleoli resembling immunoblasts • Ulcer unusual, unless due to PTLD ○ Frequent cause of bowel perforation, which may lead to resection of allograft • EBV-encoded RNA (EBER) staining required for diagnosis in most

ANCILLARY TESTS • EBER stain critical in diagnosis of EBV disease • Immunohistochemistry needed for tumors • EBV PCR very helpful to correlate with involvement and follow-up

TOP DIFFERENTIAL DIAGNOSES • Acute cellular rejection • Other viral infections

• Can involve allograft or native intestine • Increased lamina propria cellularity, predominantly lymphocytes and plasma cells

EBV: Latent Infection

EBV: Latency Pattern

Polymorphous PTLD

EBER Stain: Polymorphous PTLD

(Left) Duodenal biopsy shows preserved villous architecture with lymphoid aggregates in the lamina propria and no increase in inflammatory cells in the epithelium or lamina propria [Epstein-Barr virus (EBV) latency]. (Right) This portion of duodenum demonstrates EBV in situ hybridization (EBER) (+) nuclei ﬈ mainly restricted to lymphoid follicles in the lamina propria, consistent with an EBV latency pattern.

(Left) Section of a tonsil shows effacement of architecture and infiltration by sheets of activated lymphocytes of varying sizes with foci of necrosis consistent with a posttransplant lymphoproliferative disease (PTLD). (Right) EBER stain in the same case shows numerous EBER(+) nuclei of large activated lymphocytes ﬈ as well as smaller lymphocytes ſt suggestive of a polymorphous PTLD picture.

464

Epstein-Barr Virus, Intestine

Abbreviations • Epstein-Barr virus (EBV)

ETIOLOGY/PATHOGENESIS

Endoscopic Findings • Early infections may show erythema of mucosa • Occasional small ulcers may be noted • Mass lesions or large, deep ulcers are indicators of PTLD

MACROSCOPIC

Infectious Agents

Resected Graft

• EBV ○ B-lymphotrophic herpesvirus 4, DNA virus ○ Common complication post transplant ○ Primary infection in children often subclinical – Older children present with infectious mononucleosis (IM) ○ Secondary in children and adults ○ Both lytic (productive) and latent phase ○ Rarely, patients show recurring bouts of infection or have low-level infection as high viral load carriers – PCR monitoring shows steady or only minimally fluctuating EBV viral load of 50,000-100,000 □ Usually not in millions

• Graft resection usually done for PTLD masses but not EBV enteritis ○ Masses may ulcerate mucosa and extend transmurally ○ Size can vary from 1 cm to large necrotic masses that perforate bowel • Rare nodules and small masses found incidentally at routine surgeries, such as stoma closure

CLINICAL ISSUES Presentation • Increased stomal output or diarrhea with fevers if systemic infection • Systemic constitutional symptoms usually sign of IM • EBV PCR monitoring has greatly reduced complications of infection • Level of immune suppression is critical in determining likelihood of EBV infection

Treatment • Decreasing immune suppression ○ Complete withdrawal of immune suppression may be necessary for severe infections or posttransplant lymphoproliferative disease (PTLD) – PTLD may need chemotherapy, depending on subtype • Antiviral treatment possible with varying effects • Anti-CD20 monoclonal antibody, rituximab, used in severe infections, latent infection, and PTLD management

Prognosis • Primary infection usually responds to therapy • Balance between decreasing immune suppression and preventing rejection necessary • Difficult to predict with high-load carriers who can progress to PTLD ○ Polymorphic PTLD responds with good prognosis ○ Monomorphic PTLD may require graft removal ○ Some cases of disseminated PTLD with CNS involvement prove fatal

IMAGING Radiographic Findings

Intestinal Transplantation

TERMINOLOGY

MICROSCOPIC Histologic Features • EBV enteritis ○ Involves either allograft or native intestine with similar morphology ○ Increased lamina propria cellularity – Predominantly lymphocytes and plasma cells □ Lymphocytes appear enlarged and highly activated with prominent nucleoli, resembling immunoblasts – Increased intraepithelial lymphocytes may be noted ○ Lymphoid aggregates usually in latent infections but not acute infection – Usually indicate decreased immune suppression – May be prominent in EBV carriers ○ Ulcer unusual, unless due to PTLD • PTLD ○ Difficult to diagnose on allograft mucosal biopsies due to incomplete representation of lesion ○ Small bowel allograft is most common site for PTLD ○ Other organ involvement or nodal involvement may be seen ○ Can be polymorphic or monomorphic PTLD ○ Frequent cause of bowel perforation – May lead to resection of allograft – Usually associated with monomorphic PTLD, including posttransplant Burkitt lymphoma ○ High index of suspicion and use of ancillary techniques needed for diagnosis • Posttransplant smooth muscle (spindle cell) tumor ○ Rare instances of spindle cell proliferation on biopsy, more often on resection specimen ○ EBV in situ hybridization (EBER) shows diffuse nuclear staining of smooth muscle cells ○ Can involve mucosa and ulcerate and hence can be biopsied ○ Storiform proliferation of spindle cells with pale cytoplasm and uniform spindled nuclei characteristic ○ Prominent vascularity and perivascular proliferation seen ○ Can be multifocal and involve liver or mesentery

• IM may show enlarged lymph nodes or tonsils as well as spleen • PTLD may show nodal enlargement or mass lesions in abdomen 465

Intestinal Transplantation

Epstein-Barr Virus, Intestine EBV Disease Characteristics in Small Bowel Allografts Pattern of Graft Involvement by EBV

EBV Levels

Histologic Features

EBER Staining Pattern

Latent infection

PCR(-)

Lymphoid aggregates, no increased cellularity, no activated lymphocytes, no epithelial damage

Scattered positive nuclei mainly in lymphoid follicles; usually 1-5 cells per HPF

Epstein-Barr virus (EBV) chronic carrier status

PCR usually 50500,000

Similar to latency; may have some increased lamina propria cellularity

Mainly lymphoid aggregates, few outside, but no clustering; 5-15 per HPF

EBV infection/enteritis

PCR rising and Diffuse lamina propria cellularity; usually in 100,000s intraepithelial lymphocytes and villus alterations present; crypt injury may be seen; mixed lymphoid cells, including few activated forms

More diffusely positive within lamina propria; varying sizes of positive cells suggesting no clonal population; usually < 15 EBER(+) cells per HPF

Polymorphic posttransplant lymphoproliferative disease (PTLD)

PCR usually elevated or rising

Nodular aggregation of lymphoid cells of varying sizes with expansion of lamina propria; may be associated with visible nodule or ulcer; plasma cells frequent

Usually > 15 EBER(+) cells per HPF; staining of different-sized nuclei evident, usually no single clonal population

Monomorphic PTLD

PCR high early in transplant; low or negative late post transplant

Diffuse or nodular proliferation of monotonous large cells (usually immunoblasts) ulcerating mucosa and infiltrating wall; usually B-cell phenotype but can rarely be T-cell or plasma-cell type; common types are diffuse large B-cell lymphoma and Burkitt lymphoma

Diffusely positive in clonal population of large or small cells; numerous cells usually positive; plasma-cell type or T-cell lymphoma may be negative for EBV; small subset of Bcell PTLD is EBV negative

ANCILLARY TESTS Flow Cytometry • Indicated in PTLD, especially if mass lesion seen on resection specimen or in lymph node biopsy or tonsillectomy

In Situ Hybridization • EBER indicated in all suspected EBV cases, including PTLD ○ May serve as clonal marker in PTLD

Other Viral Infections • CMV infection usually associated with neutrophilic infiltrate ○ Characterized by presence of CMV inclusions or positive immunostain ○ Usually activated lymphocytes are not feature of CMV • Rotavirus infection may mimic with increased lamina propria cellularity ○ Needs correlation with stool cultures • Adenovirus infection usually associated with intraepithelial inclusions

PCR • EBV PCR helps to correlate with involvement and follow-up • Very high blood viral load suggests infectious stage ○ May associate with many positive EBER-staining nuclei – Diagnosis of EBV infection or mononucleosis • Progressive decreasing PCR may correlate with treated infection • Serial levels of EBV PCR in range of 100,000 copies would suggest high-load carrier status ○ Warrant close monitoring for development of PTLD • EBV PCR does not correlate well with PTLD ○ PCR may be low or even absent at time of PTLD development

DIFFERENTIAL DIAGNOSIS Acute Cellular Rejection • Both associated with increased lymphocytes, including blastic forms • Presence of crypt apoptoses is critical differentiating point ○ Many apoptoses mean acute cellular rejection ○ EBV causes only rare apoptoses, if at all • Ulcerations may be seen with both but typical of PTLD and not EBV infection

466

SELECTED REFERENCES 1.

Nassif S et al: Clinicopathologic features of post-transplant lymphoproliferative disorders arising after pediatric small bowel transplant. Pediatr Transplant. 17(8):765-73, 2013 2. Perry AM et al: Early onset, EBV(-) PTLD in pediatric liver-small bowel transplantation recipients: a spectrum of plasma cell neoplasms with favorable prognosis. Blood. 121(8):1377-83, 2013 3. Gulley ML et al: Using Epstein-Barr viral load assays to diagnose, monitor, and prevent posttransplant lymphoproliferative disorder. Clin Microbiol Rev. 23(2):350-66, 2010 4. Lau AH et al: Chronic high Epstein-Barr viral load carriage in pediatric small bowel transplant recipients. Pediatr Transplant. 14(4):549-53, 2010 5. Swerdlow SH et al: Post-transplant lymphoproliferative disorders. In: WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. Lyon: International Agency for Research on Cancer. 343-49, 2008 6. Pascher A et al: CMV, EBV, HHV6, and HHV7 infections after intestinal transplantation without specific antiviral prophylaxis. Transplant Proc. 36(2):381-2, 2004 7. Green M et al: Predictive negative value of persistent low Epstein-Barr virus viral load after intestinal transplantation in children. Transplantation. 70(4):593-6, 2000 8. Finn L et al: Epstein-Barr virus infections in children after transplantation of the small intestine. Am J Surg Pathol. 22(3):299-309, 1998 9. Lee ES et al: The association of Epstein-Barr virus with smooth-muscle tumors occurring after organ transplantation. N Engl J Med. 332(1):19-25, 1995 10. Ho M et al: The frequency of Epstein-Barr virus infection and associated lymphoproliferative syndrome after transplantation and its manifestations in children. Transplantation. 45(4):719-27, 1988

Epstein-Barr Virus, Intestine

EBV Infection (Left) Allograft duodenal biopsy reveals expanded lamina propria with increased cellularity and no significant crypt damage. Villous architecture is blunted in this example of EBV duodenitis. (Right) High-magnification view of an allograft duodenal biopsy shows increased lymphocytes in the lamina propria and some increase within crypts. Vague lymphoid aggregates are also noted. This should prompt correlation with EBV PCR and an EBER stain.

EBV Infection

Intestinal Transplantation

EBV Infection

EBV Gastritis (Left) EBER stain demonstrates an increased number of EBER(+) lymphocytes extending from a lymphoid aggregate into the surrounding lamina propria ﬈ in an example of EBV duodenitis. The EBV PCR was rising in this small bowel transplant patient. (Right) EBV gastritis affecting the native stomach in a small bowel transplant patient is shown. Note the markedly dense lamina propria lymphocytic infiltrate with intraepithelial lymphocytes ſt.

EBV Gastritis

EBV Gastritis: EBER Stain (Left) High-magnification view of EBV gastritis highlights the dense lamina propria lymphocytic infiltrate that shows small and intermediate to larger activated lymphocytes ﬇ with intraepithelial inflammatory cells ſt. There was no ulcer, and a mass lesion was not seen. (Right) Native stomach biopsy demonstrates increased numbers of EBER(+) lymphocytes in the lamina propria in this example of EBV gastritis.

467

Intestinal Transplantation

Epstein-Barr Virus, Intestine

EBV Infection: Colon

EBV Infection: Colon

EBV Infection: Colon

EBV Infection: CD20 Stain

EBV Infection: CD3 Stain

EBV Infection: EBER Stain

(Left) Colonic mucosal biopsy from a patient with a small bowel transplant shows expanded lymphoid follicles in the submucosa extending focally into the lamina propria. Although this biopsy suggests nodularity, an endoscopic mass lesion was not noted. These findings should prompt an EBER stain. (Right) Highmagnification view of a colonic biopsy reveals a submucosal lymphoid aggregate without crypt destruction and some increase in lamina propria eosinophils.

(Left) High-magnification view of a colonic biopsy demonstrates the expanded lamina propria with mixed lymphoid infiltrates, including larger lymphocytes, suspicious for EBV disease. Note damage to crypts ﬈ and some injury to surface epithelium, though no definite ulcer is seen. (Right) Immunohistochemical stain for CD20 highlights the B-cell population in the lymphoid aggregate and scant B cells in the lamina propria.

(Left) The T-lymphocyte marker CD3 is also present in abundance in the same lymphoid aggregate and lamina propria infiltrate. This mixed population of B and T cells with high T-cell content would suggest EBV infection rather than PTLD. (Right) EBER stain on a colonic biopsy with EBV colitis shows many positive nuclei in the lymphoid aggregate and in the lamina propria in a diffuse fashion with no aggregation into nodular aggregates to suggest a PTLD.

468

Epstein-Barr Virus, Intestine

EBER Stain: PTLD (Left) H&E shows a diffuse proliferation of lymphoid cells in the mucosa and submucosa ﬈ without ulceration. The absence of germinal centers should prompt staining to identify a PTLD. This case represents a polymorphic PTLD. (Right) EBER stain in an example of polymorphic PTLD shows numerous EBV(+) nuclei that stain different subtypes of lymphocytes, from small ﬈ to large ﬊ lymphoid cells. This pattern suggests a polymorphic PTLD. An occasional cell may resemble a Reed-Sternberg cell.

Monomorphic PTLD

Intestinal Transplantation

PTLD Affecting Graft

Monomorphic PTLD: EBER Stain (Left) H&E shows a proliferation of large blastic cells associated with apoptosis of tumor cells and mitoses, suggestive of high turnover and suspicious for a lymphomatous proliferation. This is an example of a monomorphic PTLD. (Right) EBER stain shows diffuse strong staining of a uniform large cell population in this example of a diffuse large Bcell lymphoma-type PTLD.

Smooth Muscle Tumor

Smooth Muscle Tumor: EBER Stain (Left) H&E shows a spindle cell proliferation composed of fascicles of cells with spindled to ovoid nuclei and prominent vascularity. This appearance resembles a leiomyoma and should warrant testing for EBV by EBER stain to confirm a posttransplant smooth muscle tumor. (Right) EBER stain shows strong, diffuse nuclear positivity in the spindle cells, confirming the diagnosis of posttransplant smooth muscle (spindle cell) tumor.

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SECTION 9

Pancreas Transplantation

Pathologic Classification of Pancreas Allograft Diseases Clinical Considerations in Pancreas Transplant Evaluation History of Pancreas Transplantation

472 474 480

Surgical Complications Surgical Aspects and Complications, Pancreas

482

Allograft Rejection Acute Cellular Rejection, Pancreas Antibody-Mediated Rejection, Pancreas Chronic Allograft Rejection/Graft Sclerosis

488 492 496

Graft Dysfunction Recurrent Diabetes Mellitus Islet Cell Toxicity and Islet Amyloid Deposition

498 502

Infections Intraabdominal and Opportunistic Infections

504

Pancreas Transplantation

Pathologic Classification of Pancreas Allograft Diseases

TERMINOLOGY Pathologic Classification

Cell-Mediated Rejection

• Based on pathogenesis, divided into broad categories of alloimmune, drug-related, and nonalloimmune (including surgical complications and recurrent diabetes)

• ACR ○ Acinitis and lobular inflammation ○ Inflammation of septa, ducts, and veins ○ Arterial endothelialitis or transmural inflammation • CR ○ Acinar inflammation and lobular atrophy ○ Chronic septal inflammation and fibrosis ○ Transplant arteriopathy (intimal inflammation, fibrosis)

Abbreviations • Acute rejection ○ Acute T-cell-mediated rejection ○ Acute antibody-mediated rejection • Chronic rejection (CR) ○ Cell-mediated type ○ Antibody-mediated type • Islet cell toxicity (ICT) • Recurrent autoimmune diabetes mellitus • Islet amyloid (IA)

Definitions • Cell-mediated rejection ○ Target antigens expressed on cell surfaces of endothelial and parenchymal cells, e.g., acinar and ductal cells ○ Alloimmune reaction of T cells to donor alloantigens, mainly against HLA classes I and II • Antibody-mediated rejection ○ Graft dysfunction due to alloantibody-mediated injury to cells expressing donor alloantigens, HLA classes I and II – Hyperacute rejection caused by preformed antibodies – Acute antibody-mediated rejection caused by de novo antibodies – Accessory mechanisms include activation of complement • C4d deposition ○ Fragment of C4 produced by activation of classic pathway that binds to endothelial cell and capillary basement membranes • Lobular/acinar inflammation ○ Infiltration by activated lymphocytes and macrophages ○ Epithelial cell lysis or necrosis ○ Disruption of acinar architecture • Ductitis/ductal inflammation ○ Infiltration by activated T lymphocytes ○ Periductal and septal inflammation • Venulitis ○ Infiltration of venular walls by inflammatory cells • Interacinar capillaritis ○ Neutrophil &/or mononuclear infiltrates • Endothelialitis/arteritis ○ Mononuclear infiltration under endothelium or transmural inflammation • Isletitis ○ Infiltration of mononuclear cells causing cellular injury – Isolated isletitis causing primarily β-cell injury – Islet inflammation as part of T-cell or antibodymediated rejection – Nonimmune inflammation due to ischemia or infectious pancreatitis • Types of pancreas transplantation ○ Simultaneous pancreas-kidney (SPK) ○ Pancreas after kidney ○ Pancreas transplant alone 472

ALLOIMMUNE RESPONSES

Antibody-Mediated Rejection • Hyperacute rejection ○ Edema, congestion – Occurs immediately ○ Confluent hemorrhagic necrosis of parenchyma ○ Fibrinoid vascular necrosis and thrombosis • Acute antibody-mediated rejection ○ Acinar and interacinar capillary inflammation ○ Monocytic &/or neutrophil infiltrates ○ C4d(+) interacinar capillaries • Chronic active antibody-mediated rejection ○ Features of antibody-mediated rejection ○ C4d positive or negative ○ Acinar atrophy and septal fibrosis

Indeterminate • Septal inflammation that is active but does not fulfill criteria for mild acute rejection

DRUG TOXICITY Islet Cell Toxicity • Caused by cyclosporine, tacrolimus ○ Dose and duration dependent • Hyperglycemia and low insulin levels • Histologic features of ICT ○ Islet cell swelling and vacuolization ○ Nuclear and cytoplasmic features of apoptosis ○ Normal pancreatic lobules

NONALLOIMMUNE DISEASES Acute Ischemic Pancreatitis • Hemorrhagic or coagulative necrosis

Acute Infectious Peripancreatitis • Purulent inflammatory exudate on surface or invading capsule with fat necrosis

Viral Infections • • • •

Cytomegalovirus Adenovirus Herpesvirus Others (rare)

Bacterial Infections • • • •

Escherichia coli Pseudomonas species Enterococcus faecalis Due to surgical complications and type of exocrine drainage

Pathologic Classification of Pancreas Allograft Diseases

• Mainly Candida species

Major Blood Vessel Disease • Arterial thrombosis • Venous thrombosis

Surgical Complications • Bladder drainage ○ Reflux pancreatitis ○ Urinary tract infections ○ Dehydration, chronic metabolic acidosis • Enteric drainage ○ Duodenal fistula ○ Anastomotic bleeding ○ Source of infection/peritonitis • Systemic venous drainage ○ Peripheral hyperinsulinemia ○ Increased atherosclerosis • Portal venous drainage ○ Physiologic state of insulin metabolism – Insulin partly inactivated by passage through liver

• Uncertainty of optimal immunosuppression • Irregular rate of loss of insulin-producing cells over time

NOTES Comments • Pancreatic dysfunction due to any cause ○ Rejection, toxicity, autoimmune mechanisms ○ Often asymptomatic and sometimes symptomatic, particularly with infections • Laboratory testing for pancreatic enzymes, blood glucose levels, hormone levels, and antibody titers are helpful • Pancreatic transplant biopsy useful to identify underlying disease process • Kidney transplant biopsy may show rejection process in patients with SPK transplants ○ May act as surrogate for pancreas biopsy

SELECTED REFERENCES 1.

2.

Reflux (Aseptic) Pancreatitis • Intestinal obstruction • Pancreatic ductal obstruction

Recurrent Autoimmune Diabetes Mellitus • Rising titers of antiislet cell antibodies • Autoimmune T-cell mediated isletitis with loss of β cells

3.

4.

5.

Neoplasia

6.

• Posttransplant lymphoproliferative disease ○ Epstein-Barr virus (EBV) and non-EBV related

7.

Islet Amyloid

8.

• Accumulation of interstitial pale eosinophilic amyloid mainly in islets • Amyloid deposits proportional to hyperglycemia • Relatively preserved α- and β-cell populations, initially • Progressive β-cell dysfunction and loss • Due to IA polypeptide secretion by β cells, particularly with hyperinsulinemia

9.

10. 11.

12.

ISLET CELL TRANSPLANTATION Choice of Technique and Location

13.

• Mostly liver via portal vein infusion • Other potential sites: Kidney capsule, skin

14.

Sources of Islet Cells

Pancreas Transplantation

Fungal Infections

15.

Loupy A et al: The Banff 2015 Kidney meeting report: current challenges in rejection classification and prospects for adopting molecular pathology. Am J Transplant. 17(1):28-41, 2017 Kumar R et al: Current principles and practice in autologous intraportal islet transplantation: a meta-analysis of the technical considerations. Clin Transplant. 30(4):344-56, 2016 Troxell ML et al: Practical applications in immunohistochemistry: evaluation of rejection and infection in organ transplantation. Arch Pathol Lab Med. 140(9):910-25, 2016 León Fradejas M et al: Islet amyloid in whole pancreas transplants for type 1 diabetes mellitus (DM): possible role of type 2 DM for graft failure. Am J Transplant. 15(9):2495-500, 2015 de Kort H et al: Pancreas transplantation, antibodies and rejection: where do we stand? Curr Opin Organ Transplant. 18(3):337-44, 2013 Mengel M et al: Banff 2011 Meeting report: new concepts in antibodymediated rejection. Am J Transplant. 12(3):563-70, 2012 Papadimitriou JC et al: Distinctive morphological features of antibodymediated and T-cell-mediated acute rejection in pancreas allograft biopsies. Curr Opin Organ Transplant. 17(1):93-9, 2012 Drachenberg CB et al: Guidelines for the diagnosis of antibody-mediated rejection in pancreas allografts-updated Banff grading schema. Am J Transplant. 11(9):1792-802, 2011 Wee AC et al: Pancreas transplantation: surgical techniques. In Srinivas TR et al: Kidney and Pancreas Transplantation: A Practical Guide. New York City: Springer Science+Business Media. 249-58, 2011 Sollinger HW et al: One thousand simultaneous pancreas-kidney transplants at a single center with 22-year follow-up. Ann Surg. 250(4):618-30, 2009 Drachenberg CB et al: Banff schema for grading pancreas allograft rejection: working proposal by a multi-disciplinary international consensus panel. Am J Transplant. 8(6):1237-49, 2008 Humar A et al: Technical failures after pancreas transplants: why grafts fail and the risk factors--a multivariate analysis. Transplantation. 78(8):1188-92, 2004 Papadimitriou JC et al: Histological grading of chronic pancreas allograft rejection/graft sclerosis. Am J Transplant. 3(5):599-605, 2003 Sutherland DE et al: Lessons learned from more than 1,000 pancreas transplants at a single institution. Ann Surg. 233(4):463-501, 2001 Knight RJ et al: Risk factors for intra-abdominal infection after pancreas transplantation. Am J Surg. 179(2):99-102, 2000

• Living or deceased donor, single or pooled • Autotransplantation

Advantages • • • •

Less invasive procedure, low complications Sustained insulin independence (> 5 years) Beneficial for microvascular complications, e.g., retinopathy Amelioration of hypoglycemia-unaware events

Disadvantages • Unpredictability in yield and engraftment • Toxic or autoimmune islet cell damage 473

Pancreas Transplantation

Clinical Considerations in Pancreas Transplant Evaluation

TERMINOLOGY Types of Pancreas Transplantation • Simultaneous pancreas-kidney (SPK) ○ Advanced diabetic nephropathy ○ Uremia/end-stage renal disease ○ 75% of cases • Pancreas after kidney (PAK) ○ Functioning kidney transplant in end-stage kidney disease followed by pancreas transplant (PTx) ○ 18% of cases • Pancreas transplant alone (PTA) ○ Nonuremic diabetic patients ○ Other end-stage pancreatic diseases ○ 7% of cases – Segmental – Whole pancreas • Pancreas and liver • Islet cell transplant

CLINICAL IMPLICATIONS Indications for Pancreas Transplantation • C-peptide-deficient, insulin-dependent type 1 diabetes mellitus • Advanced or end-stage kidney disease or diabetic nephropathy • Severe systemic complications of diabetes • Loss of exocrine and endocrine pancreas function ○ Chronic pancreatitis with total pancreatectomy ○ Cystic fibrosis ○ Pancreatic tumors with pancreatectomy • Unconventional recipients ○ Hepatitis C virus and HIV positive ○ Obesity ○ Type 2 ( C-peptide-positive) diabetes

Benefits of Pancreas Transplantation • Overall quality of life improvement • Metabolic response

○ Glycemic control and glucose homeostasis ○ Prevention of recurrent hypoglycemic reactions ○ Improved lipid profile • Hormonal response ○ Improved glucagon response ○ Improved catecholamine response ○ Source of pancreatic polypeptides • Improvement in end-organ diabetic complications ○ Nephropathy ○ Retinopathy ○ Neuropathy ○ Cardiovascular morbidity ○ Other microvascular and macrovascular complications ○ Overall survival rates • Replacement of exocrine and endocrine pancreas function following ○ Chronic pancreatitis ○ Pancreas resection due to benign tumor

Challenges of Pancreas Transplantation • Surgical complications ○ Exocrine drainage ○ Endocrine drainage • Chronic hyperglycemia ○ Graft failure – Acute or chronic rejection – Pancreatitis – Thrombosis ○ Insulin resistance with new-onset type 2 diabetes – Weight gain – Coexistent obesity – Family history of diabetes ○ Adverse effects of immunosuppression – Calcineurin inhibitor-induced islet cell toxicity – Dyslipidemia due to medications ○ Immune-mediated islet cell damage – Recurrent type 1 diabetes – Antiislet and antiglutamic acid decarboxylase antibodies

Anatomy of Pancreas Transplant (Left) Graphic shows pancreas transplant preparation with a donor duodenal segment and donor iliac artery joined to donor superior mesenteric and splenic arteries. (Right) Axial CECT shows a pancreatic allograft ſt with mild gland swelling and peripancreatic infiltration, which are common in the postoperative period.

474

Pancreatic Allograft and Adjacent Tissue

Clinical Considerations in Pancreas Transplant Evaluation

OUTCOMES Factors Determining Outcome of PTx • Donor selection ○ Donors ≤ 50 years old (favorable) – Deceased: Segmental, whole pancreas – Living: Segmental ○ Body mass index (BMI) ≤ 30 kg/m² • Optimal organ preservation techniques ○ Medium of preservation ○ Duration of cold ischemia time • Type of pancreas transplantation ○ SPK – 1-year graft survival: 86% – 1-year patient survival: 95% – Risk of graft loss: 2% ○ PAK – 1-year graft survival: 80% – 1-year patient survival: 97% – Risk of graft loss: 8% ○ PTA – 1-year graft survival: 78% – 1-year patient survival: 99% – Risk of graft loss: 10% • Recipient selection with relative contraindications ○ Significant cardiovascular disease – Requires appropriate invasive and noninvasive testing ○ Severe peripheral vascular disease ○ BMI > 35 kg/m² ○ Substance abuse ○ Poorly controlled psychiatric illness ○ Noncompliance ○ Evidence of malignancy • Human leukocyte antigen matching ○ No significant correlation in SPK ○ Matching relevant in PAK and PTA • Surgical techniques and complications • Induction and immunosuppressive protocols ○ Considerable reduction in rejection rate • Prevention or early identification of intraabdominal infections ○ Bacterial/fungal • Serologic monitoring and early identification of opportunistic infections ○ Viral: e.g., BK virus, cytomegalovirus, Epstein-Barr virus, adenovirus ○ Bacterial ○ Fungal • Pancreatic dysfunction ○ Allograft rejection – Acute cell-mediated rejection – Acute antibody-mediated rejection – Chronic allograft rejection

○ Recurrent diabetes mellitus or islet cell disease ○ Medication-induced islet cell or pancreatic diseases

ASSESSMENT Pancreas Transplant Dysfunction • Pancreatic enzymes ○ Amylase in serum and urine ○ Lipase ○ Glucose, HbA1c, glucose tolerance test ○ C-peptide levels, insulin • Transplant renal function as surrogate markers ○ Creatinine ○ Proteinuria • Serology ○ Immune monitoring for rejection – Donor-specific antibodies ○ Anti-islet cell antibodies – Anti-glutamic acid decarboxylase 65 – Anti-protein tyrosine phosphatase-like protein IA2 – Cytoplasmic islet cell antibodies • Imaging studies ○ Peripancreatic swelling and fluid collection ○ Acute pancreatitis ○ Duct obstruction ○ Perfusion defects ○ Abscess or pseudocysts ○ Anastomotic leaks • Graft failure ○ Varies by definitions by different programs – C-peptide deficiency (< 0.5 ng) – Insulin dependence

Pancreas Transplantation

• Hypertension ○ Immunosuppression effect ○ Preexisting vascular disease • Infections ○ Local: e.g., bacterial, fungal ○ Systemic: e.g., viral, bacterial

DIAGNOSTIC UTILITY OF PANCREAS TRANSPLANT BIOPSY 1992 • Nakhleh ○ Histopathologic findings with implications for classification of rejection

1995 • Nakhleh et al ○ Graft duodenum and pancreas biopsy for therapy

1997 • Drachenberg et al ○ Evaluation of pancreas transplant biopsy ○ Reproducibility and revision of histologic grading • Gill et al ○ Correlation of serologic and urinary tests with PTx biopsy for diagnosis of rejection

2003 • Papadimitriou ○ Histologic grading of chronic PTx rejection

2008 • Drachenberg et al ○ Banff schema for grading pancreas allograft rejection

475

Pancreas Transplantation

Clinical Considerations in Pancreas Transplant Evaluation – Pancreatic tumors

2011 • Drachenberg et al ○ Guidelines for diagnosis of antibody-mediated rejection in PTx ○ Updated Banff grading schema

2013-2015 • DeKort et al, Loupy/Drachenberg et al ○ Updated Banff schema and grading of antibodymediated rejection in PTx

COMPREHENSIVE HISTOLOGIC EXAMINATION Pancreas Transplant Biopsy Work-Up • PTx biopsy is standard for diagnosis of rejection • Specimen procurement and preparation ○ Ultrasound or computed tomographic guidance used ○ Common needle gauge is 18 or 20 ○ Complications are rare (2-3%) ○ Multiple levels of paraffin sections – Hematoxylin and eosin (3 levels) and trichrome stains – Multiple unstained sections for additional immunohistochemistry • Morphologic evaluation of biopsies ○ Assess adequacy of pancreatic tissue ○ Pancreas lobular architecture and acini ○ Interlobular septa and blood vessels ○ Islet quantity and quality ○ Immunohistochemistry – C4d by immunofluorescence or immunoperoxidase stains – Insulin and glucagon in cases of hyperglycemia – Immunophenotyping infiltrating inflammatory cells ○ Classification and grading of rejection using accepted schema ○ Diagnosis of other pancreas allograft diseases – Electron microscopy of islet diseases • Systematic examination of failed pancreatic explants ○ Gross study: Arterial vessels and veins ○ Microscopic study: Adequate sampling of parenchyma; 410 sections for accurate diagnosis

Advantages • Less invasive procedure ○ Decreased patient morbidity • Can be combined with kidney and liver transplantation • Significant cost savings • Indicated in patients with brittle diabetes and severe hypoglycemia

Disadvantages • Relatively lower yield and quality of islets harvested from potential pancreas allografts • Difficulty in initial engraftment and function • Majority (80%) of patients require exogenous insulin • Defective glucagon responses to hypoglycemia with intrahepatic islet transplantation • Islet cell toxicity following immunosuppression • Immune-mediated islet damage • Poor long-term islet graft function: 20-27% 1-year survival • Islet amyloid deposition

SELECTED REFERENCES 1. 2.

3. 4. 5. 6. 7. 8.

9.

10. 11. 12.

HUMAN ISLET TRANSPLANTATION Technique and Location • • • •

Portal vein infused for implantation in liver Placed under kidney capsule Injected intraperitoneally Future strategy: insulin-producing stem cells as surrogate for human islet transplantation

13.

14. 15. 16.

Sources of Islet Cells • Allotransplantation ○ Deceased donor without suitable recipients – Single pancreas islets – Pooled islets from several donors ○ Living donors • Autotransplantation ○ From total pancreatectomy – Chronic pancreatic diseases 476

17. 18. 19. 20. 21.

Kandaswamy R et al: OPTN/SRTR 2016 annual data report: pancreas. Am J Transplant. 18 Suppl 1:114-171, 2018 Loupy A et al: The Banff 2015 Kidney meeting report: current challenges in rejection classification and prospects for adopting molecular pathology. Am J Transplant. 17(1):28-41, 2017 Lo DJ et al: Pancreas transplantation in unconventional recipients. Curr Opin Organ Transplant. 21(4):393-8, 2016 Stratta RJ et al: Pancreas transplantation in C-peptide positive patients: does "type" of diabetes really matter? J Am Coll Surg. 220(4):716-27, 2015 Mittal S et al: Pancreas transplantation: a treatment option for people with diabetes. Diabet Med. 31(5):512-21, 2014 Rogers J et al: Pancreas transplantation with portal venous drainage with an emphasis on technical aspects. Clin Transplant. 28(1):16-26, 2014 de Kort H et al: Pancreas transplantation, antibodies and rejection: where do we stand? Curr Opin Organ Transplant. 18(3):337-44, 2013 Drachenberg CB et al: Guidelines for the diagnosis of antibody-mediated rejection in pancreas allografts-updated Banff grading schema. Am J Transplant. 11(9):1792-802, 2011 Gruessner AC: 2011 update on pancreas transplantation: comprehensive trend analysis of 25,000 cases followed up over the course of twenty-four years at the International Pancreas Transplant Registry (IPTR). Rev Diabet Stud. 8(1):6-16, 2011 Sollinger HW et al: One thousand simultaneous pancreas-kidney transplants at a single center with 22-year follow-up. Ann Surg. 250(4):618-30, 2009 White SA et al: Pancreas transplantation. Lancet. 373(9677):1808-17, 2009 Dean PG et al: Long-term benefits of pancreas transplantation. Curr Opin Organ Transplant. 13(1):85-90, 2008 Drachenberg CB et al: Banff schema for grading pancreas allograft rejection: working proposal by a multi-disciplinary international consensus panel. Am J Transplant. 8(6):1237-49, 2008 Andreoni KA et al: Kidney and pancreas transplantation in the United States, 1996-2005. Am J Transplant. 7(5 Pt 2):1359-75, 2007 Cohen DJ et al: Kidney and pancreas transplantation in the United States, 1995-2004. Am J Transplant. 6(5 Pt 2):1153-69, 2006 Gruessner AC et al: Pancreas transplant outcomes for United States (US) and non-US cases as reported to the United Network for Organ Sharing (UNOS) and the International Pancreas Transplant Registry (IPTR) as of June 2004. Clin Transplant. 19(4):433-55, 2005 Larsen JL: Pancreas transplantation: indications and consequences. Endocr Rev. 25(6):919-46, 2004 Papadimitriou JC et al: Histological grading of chronic pancreas allograft rejection/graft sclerosis. Am J Transplant. 3(5):599-605, 2003 Hariharan S et al: Pancreas after kidney transplantation. J Am Soc Nephrol. 13(4):1109-18, 2002 Sutherland DE et al: Lessons learned from more than 1,000 pancreas transplants at a single institution. Ann Surg. 233(4):463-501, 2001 Alejandro R et al: Long-term function (6 years) of islet allografts in type 1 diabetes. Diabetes. 46(12):1983-9, 1997

Clinical Considerations in Pancreas Transplant Evaluation Normal Perfusion of Kidney-Pancreas Transplant (Left) Axial CECT shows a normal pancreatic allograft ſt in the immediate postoperative period with mild, diffuse swelling of the allograft and soft tissue in the adjacent fat planes st. These findings are frequently seen immediately post transplant. (Right) Coronal scan from an MRA of a kidney-pancreas transplant reveals that the renal st and pancreatic allografts ſt show normal perfusion and parenchymal enhancement.

Normal Architecture of Pancreas Tissue

Pancreas Transplantation

Post Pancreas Transplant With Swelling

Normal Pancreas Acinar Structure (Left) Low-magnification view of a normal pancreas transplant biopsy demonstrates lobular architecture with thin septa and ducts ﬈ and scattered islets within the lobules ﬉. (Right) High-magnification view of a normal pancreas transplant biopsy reveals a preserved acinar structure surrounded by delicate fibrous septa with arterioles and capillaries.

Normal Islet Cell Aggregate

Normal Pancreas Ductal Elements (Left) High-magnification view of a normal pancreatic lobule shows an islet aggregate containing occasional intraislet capillaries with red cells. (Right) Highmagnification view shows the periphery of normal pancreatic lobules containing ductal elements in the septa ﬈.

477

Pancreas Transplantation

Clinical Considerations in Pancreas Transplant Evaluation

Normal Septal Vasculature

Insulin (+) Cells in Normal Pancreatic Islet

Glucagon (+) Cells in Normal Pancreatic Islet

Somatostatin (+) Cells in Normal Pancreatic Islet

Acute Cellular Rejection

Acute Cellular Rejection With Septal and Ductal Inflammation

(Left) These normal pancreatic lobules are arranged with adjacent septal arterioles ﬈ and a venule ﬉. (Right) A normal islet in a pancreas transplant biopsy shows strong immunostaining for insulin with occasional negative cells ﬊.

(Left) This normal islet in a pancreas transplant biopsy demonstrates positive immunostaining for glucagon in a few cells ﬈, while the majority of them are negative. (Right) This normal islet in a pancreas transplant biopsy reveals positive immunostaining for somatostatin in occasional cells ﬊.

(Left) Acute cellular rejection in a pancreatic transplant biopsy is shown involving the acini, which are infiltrated by activated lymphocytes and a few macrophages with focal disruption and loss of acinar cells ﬈. (Right) H&E shows acute cellular rejection in a pancreatic transplant biopsy with septal inflammation and ductal infiltration by lymphocytic inflammatory cells (ductitis) ſt.

478

Clinical Considerations in Pancreas Transplant Evaluation Acute Cellular Rejection With Lobular and Perineural Inflammation (Left) Acute cellular rejection in a pancreatic transplant biopsy demonstrates marked venulitis and perivenular inflammation ﬊ with the periphery of a pancreatic lobule on the right side showing an inflammatory infiltrate ﬈. (Right) Acute cellular rejection in a pancreatic transplant biopsy reveals lobular inflammation, septal inflammation, and perivenular ﬈ and perineural ﬉ lymphocytic inflammatory infiltrate.

Acute T-Cell-Mediated Rejection

Pancreas Transplantation

Acute Cellular Rejection With Venulitis and Perivenular Inflammation

Acute Cellular Rejection Localizing Macrophages (Left) In this example of acute cellular rejection in a pancreatic transplant biopsy with lobular infiltration and acinitis ﬈, many CD3-positive T lymphocytes are highlighted by immunoperoxidase stain. (Right) CD68 immunoperoxidase study highlights many macrophages in a pancreatic transplant biopsy with acute cellular rejection showing septal and lobular infiltration ﬊.

Chronic Pancreas Rejection With Septal Fibrosis

C4d Deposition of Capillaries in AntibodyMediated Rejection (Left) Low-magnification view of a pancreatic transplant biopsy using trichrome stain demonstrates chronic rejection with expanded and fibrosing intervening septa (blue staining), which surrounds fragmented lobules with acini ﬈. (Right) Immunofluorescence microscopy for C4d confirms antibody-mediated rejection and shows diffuse, linear staining of the interacinar capillaries and a small artery ſt.

479

Pancreas Transplantation

History of Pancreas Transplantation

TERMINOLOGY Abbreviations

•

• Pancreas transplantation (PTx)

Synonyms • • • •

Pancreatic homotransplantation Allotransplantation of pancreas Pancreaticoduodenal allograft Pancreatic transplantation

•

Definitions • Transplantation of vascularized pancreas to effectively treat human type 1 diabetes mellitus ○ Establish long-term normoglycemia ○ Normalize HbA1c ○ Prevent consequences of metabolic abnormalities

CHRONOLOGY AND EVOLUTION Timeline • 4th documented organ allograft ○ Following kidney, liver, and heart • 1928-1929 ○ Gayet et al, Houssay – PTx as physiologic experiment to normalize glucose homeostasis • 1959-1962 ○ Brooks et al, DeJode et al – Experimental PTx to treat diabetes in laboratory animals • 1966 ○ Kelly et al – 1st PTx in human diabetic patient □ University of Minnesota, Minneapolis □ Duct-ligated segmental graft from deceased donor □ Simultaneously with kidney transplant □ Achieved insulin independence for 6 days ○ Lillehei et al – Whole pancreaticoduodenal transplant – Stoma of donor duodenum for drainage of exocrine secretions – Improved to provide enteric drainage via Roux-en-Y loop • 1969-1980 ○ Impediments to initial and long-term successful PTx – Experimental and clinical techniques □ Complications of surgical techniques – Lack of adequate organ procurement and graft preservation techniques – Immunologic failure; lack of optimal immunosuppression protocols – Infectious complications – Lack of diagnostic and histopathologic monitoring of allograft ○ Poor transplant outcomes • 1980 ○ Sutherland – International Pancreas Transplant Registry • 1985 ○ Starzl 480

•

•

– Reintroduced enteric-drained whole-organ pancreaticoduodenal transplants 1985-1994 ○ Sutherland et al, Sollinger et al – Urinary bladder or enteric drainage of exocrine pancreas secretions – Improved immunosuppression and prophylaxis for infections – Histopathologic assessment for allograft rejection 1989 ○ Bilous et al – Successful PTx associated with less severe diabetic glomerulopathy in kidney transplants 1998 ○ Fioretto et al – PTx reverses lesions of diabetic nephropathy – Pathologic changes observed 5-10 years later 2001 ○ Gruessner et al – Living-donor laparoscopic distal pancreatectomy

PROGRESS IN TREATMENT Immunosuppressive Protocols in PTx • 1967-1973 ○ Lillehei et al – Azathioprine, high-dose steroids, and antilymphocyte globulin – Complications included delayed wound healing, sepsis, and diabeticogenicity of steroids • 1978-1986 ○ Sutherland et al, Sollinger et al – Use of cyclosporine in PTx – Minnesota antilymphocyte globulin for induction • 1981 ○ Cosimi et al – Use of OKT3 (antibody to T lymphocytes) for rejection – Later used in PTx • 1986-1994 ○ Sutherland et al – Cyclosporine, azathioprine, and prednisone for maintenance therapy – Antithymocyte globulin (ATGAM) for induction in 1993 • 1989 ○ Starzl et al – Introduced tacrolimus (FK506) in organ transplant – Potential dose-related diabetogenicity recognized – Steroid-sparing effect – Superior control of rejection • 1994-1998 ○ Clinical use of tacrolimus in combination with mycophenolate mofetil for maintenance • 1999 ○ Thymoglobulin (rabbit antithymocyte) for induction therapy

PANCREAS TRANSPLANT BIOPSY Timeline • 1986

History of Pancreas Transplantation

•

•

•

•

•

•

HUMAN ISLET TRANSPLANTATION Definition • Transplantation of islet cells harvested from human pancreas in patients with type 1 diabetes mellitus, resulting in insulin independence and normoglycemia

Timeline • 1967 ○ Lacy et al – Successful isolation of pancreatic islets • 1972 ○ Ballinger et al – Islet transplant partially corrected diabetes in rats • 1980 ○ Largiader et al – Allotransplantation led to normoglycemia and insulin independence in human – Required 200,000 islets from young donor • 1980 ○ Najarian et al – Successful islet autotransplantation in humans – Treatment of chronic pancreatitis • 1990 ○ Scharp et al, Tzakis et al – Insulin independence and nearly normal glucose levels after islet allotransplantation – C-peptide secretion in response to glucose • 1992 ○ Warnock et al – Freshly prepared and cryopreserved islet allotransplantation – Insulin independence and normoglycemia, up to 1 year

• 1995 ○ Wahoff et al – Islet autotransplantation in 48 patients after pancreatectomy for chronic pancreatitis – Number of islets transplanted (> 300,000) predicted long-term insulin independence • 1997 ○ Alejandro et al – Allogeneic islet transplant with long-term function in type 1 diabetes • 2000 ○ Shapiro et al – Successful high-quality islet transplant using 2 or more islet infusions – Steroid-free regimen with tacrolimus and sirolimus • 2005 ○ Shapiro et al – Edmonton protocol: Combination of sirolimus, tacrolimus, and daclizumab • 2007 ○ Shapiro et al – Identified chronic graft dysfunction in islets – Immune rejection, recurrence of autoimmunity, exposure to diabetogenic drugs • 2017 ○ Smink et al – Prevascularized subcutaneous scaffold as transplant site for pancreas islets

Pancreas Transplantation

•

○ Steiniger et al – Histologic patterns of rejection in rat PTx 1987 ○ Sutherland et al, Sibley et al – Percutaneous PTx biopsy for diagnosis – Histologic and immunohistochemical studies 1990 ○ Bernardino et al – CT-guided PTx biopsy 1991 ○ Allen et al – Percutaneous biopsy of bladder-drained PTx 1994 ○ Jones et al – Cystoscopic transduodenal PTx biopsy 1997 ○ Egidi et al – Fine-needle aspiration biopsy to monitor entericdrained PTx 2001 ○ Stegall – Surveillance biopsies in solitary PTx 2002 ○ Klassen et al – Safety of percutaneous PTx biopsy

SELECTED REFERENCES 1.

2. 3. 4. 5.

6. 7. 8. 9. 10. 11. 12.

13. 14.

15. 16.

Smink AM et al: The efficacy of a prevascularized, retrievable poly(D,L,lactide-co-ε-caprolactone) subcutaneous scaffold as transplantation site for pancreatic islets. Transplantation. 101(4):e112-e119, 2017 Pepper AR et al: Current status of clinical islet transplantation. World J Transplant. 3(4):48-53, 2013 Sollinger HW et al: One thousand simultaneous pancreas-kidney transplants at a single center with 22-year follow-up. Ann Surg. 250(4):618-30, 2009 White SA et al: Pancreas transplantation. Lancet. 373(9677):1808-17, 2009 Andreoni KA et al: Kidney and pancreas transplantation in the United States, 1996-2005. Am J Transplant. 2007;7(5 Pt 2):1359-75. Erratum in: Am J Transplant. 7(9):2214, 2007 Cohen DJ et al: Kidney and pancreas transplantation in the United States, 1995-2004. Am J Transplant. 6(5 Pt 2):1153-69, 2006 Hariharan S et al: Pancreas after kidney transplantation. J Am Soc Nephrol. 13(4):1109-18, 2002 Becker BN et al: Simultaneous pancreas-kidney and pancreas transplantation. J Am Soc Nephrol. 12(11):2517-27, 2001 Sutherland DE et al: Lessons learned from more than 1,000 pancreas transplants at a single institution. Ann Surg. 233(4):463-501, 2001 Robertson RP et al: Pancreas and islet transplantation for patients with diabetes. Diabetes Care. 23(1):112-6, 2000 Fioretto P et al: Reversal of lesions of diabetic nephropathy after pancreas transplantation. N Engl J Med. 339(2):69-75, 1998 Gores PF et al: Insulin independence in type I diabetes after transplantation of unpurified islets from single donor with 15-deoxyspergualin. Lancet. 341(8836):19-21, 1993 Pyzdrowski KL et al: Preserved insulin secretion and insulin independence in recipients of islet autografts. N Engl J Med. 327(4):220-6, 1992 Warnock GL et al: Long-term follow-up after transplantation of insulinproducing pancreatic islets into patients with type 1 (insulin-dependent) diabetes mellitus. Diabetologia. 35(1):89-95, 1992 Sutherland DE: International human pancreas and islet transplant registry. Transplant Proc. 12(4 Suppl 2):229-36, 1980 Lillehei RC et al: Pancreatico-duodenal allotransplantation: experimental and clinical experience. Ann Surg. 172(3):405-36, 1970

481

Pancreas Transplantation

Surgical Aspects and Complications, Pancreas

TERMINOLOGY Definitions

•

• Whole-organ pancreas transplantation ○ Pancreas allograft reconstruction ○ Y graft from donor iliac artery to donor superior mesenteric and splenic arteries ○ Arterial anastomosis to common iliac artery in right iliac fossa • Exocrine bladder drainage ○ Pancreatic secretions drained via duct and duodenal segment anastomosed to recipient bladder • Exocrine enteric drainage ○ Pancreatic secretions drained using duodenal segment anastomosed to recipient jejunum via Roux-en-Y loop • Endocrine systemic venous drainage ○ Hormonal secretions delivered to common or right iliac vein • Endocrine portal venous drainage ○ Hormonal secretions delivered to portal vein • Graft pancreatitis ○ Develops secondary to ischemic injury caused by dissolution of parenchymal structures with leakage of cellular content evoking inflammatory reaction

•

•

•

•

HISTORY Evolution of Surgical Techniques • 1968 ○ Lillehei et al – Whole pancreaticoduodenal transplant – Enteric drainage with donor duodenum • 1973 ○ Gliedman et al – Anastomosis of pancreatic duct from segmental graft to recipient ureter • 1978 ○ Dubernard JM et al, Sutherland et al – Multiple surgical techniques using segmental pancreatic grafts

•

– Experimental in dogs and humans – Transplant outcomes poor due to complications 1980 ○ Sutherland et al – Segmental pancreas transplant (PTx) from living donors 1983-1985 ○ Sollinger et al, Cook – Pancreaticocystostomy for exocrine bladder drainage in dogs – Segmental and pancreaticosplenic transplant with pancreaticocystostomy in humans 1987 ○ Ngheim and Corry – Whole pancreaticoduodenal grafts used with bladder drainage 1992 ○ Rosenlof et al, Shokouh-Amiri et al – Portal drainage of pancreas endocrine secretions in enteric-drained, whole-organ pancreaticoduodenal transplants 1998-2007 ○ Sutherland, Sollinger – Primarily enteric drainage of exocrine secretions – Increased enteric conversion in bladder-drained pancreas allografts – Improved PTx outcomes [simultaneous pancreaskidney (SPK), pancreas after kidney (PAK), pancreas transplant alone (PTA)] 2013 ○ Rogers et al – Choice of surgical technique dictated by donor and recipient anatomy and surgeon experience

ETIOLOGY/PATHOGENESIS Complications • Graft thrombosis ○ Higher with segmental grafts ○ Dependent on type of surgical procedure

Infarcted Pancreatic Allograft (Left) Gross pathologic specimen shows an infarcted pancreatic allograft. (Right) Transverse color Doppler US in a patient with an allograft infarction shows the allograft ſt as an enlarged, hypoechoic structure with virtually no evidence of blood flow within.

482

Pancreatic Allograft Infarction by Doppler US

Surgical Aspects and Complications, Pancreas

• • •

•

CLINICAL IMPLICATIONS Epidemiology • Pancreas transplantation is gold standard endocrine replacement in diabetic patients • > 30,000 pancreas transplants performed according to International Pancreas Transplant Registry • Surgical and technical complication rate (15-30%) higher than other solid organ transplants • > 10% of PTx lost due to technical reasons

Risk Factors for Surgical Complications • Donor factors (pancreas donor risk index) ○ Age, obesity/high body mass index ○ Cause of death • Recipient factors ○ Age ○ Obesity ○ Cardiac disease • Type of pancreatic transplant ○ SPK ○ PTA ○ PAK • Increased preservation times (cold ischemia times) • Surgical technique ○ Exocrine bladder drainage ○ Exocrine enteric drainage ○ Implantation site • Infections ○ Intraabdominal ○ Systemic • Imaging studies sensitive for graft dysfunction but lack specificity to differentiate etiologies • Readmission and laparotomy for technical failures ○ Higher in SPK ○ Higher in cases with bladder drainage

Presentation • Vascular graft thrombosis

○ Common cause of early graft dysfunction/relaparotomy ○ Graft tenderness ○ Decreased or absent urinary amylase levels ○ Hyperglycemia with increasing insulin requirement ○ Dark hematuria with bladder drainage ○ Leucocytosis/thrombocytopenia • Anastomotic and duodenal stump leak ○ Abdominal pain ○ Abdominal distension ○ Fever ○ Symptoms of peritonitis • Graft pancreatitis and peripancreatitis ○ Abdominal pain, distension, and symptoms of peritonitis

Pancreas Transplantation

•

○ Relative hypercoagulable state in PAK ○ Associated with acute rejection – Cell-mediated vascular rejection – Antibody-mediated vascular rejection Anastomotic and duodenal stump leak ○ Early postoperative period – Reperfusion/ischemic damage of donor duodenum – Impaired wound healing – Technical complications during surgery ○ Late postoperative period – Infections – Rejection – Ischemia Postoperative bleeding ○ Surgical complication Portal venous thrombosis Graft pancreatitis ○ Poor perfusion or pancreatic necrosis ○ Pancreatic duct injury or reflux ○ Complicated by subsequent infection Sterile pancreatic and peripancreatic fluid collections

Preventive Strategies and Treatment • Appropriate donor selection • Recipients with fewer risk factors • Meticulous surgical technique during ○ Graft procurement ○ Preparation ○ Implantation • Postoperative anticoagulation therapy • Successful PTx despite technical difficulties depends on prevention of allograft rejection ○ Optimal induction and maintenance immunosuppressive therapy • Pancreas transplant salvage techniques ○ Reanastomosis ○ Thrombectomy ○ Pancreatic graft explant ○ Duodenoduodenostomy and caval drainage • Prophylactic therapy for potential infections ○ Drainage of intraabdominal abscesses

Prognosis • Technical complications are common cause of graft dysfunction/failure ○ Graft thrombosis most frequent cause of graft failure ○ High patient morbidity – < 10% mortality ○ Increased potential for infections ○ Higher rate of all complications in SPK • Readmission and laparotomy for technical failures ○ Higher rate in cases with bladder drainage

IMAGE FINDINGS Vascular Graft Thrombosis • Color duplex Doppler ultrasonography ○ Infarcted pancreas is enlarged, hypoechoic, and heterogeneous ○ Vascular thrombosis – Absence of arterial or venous tracings – Presence of intraluminal echogenic material • Pancreas perfusion scintigraphy ○ Tc-99m DPTA or Tc-99m MAG3 used ○ Delay in peak graft flow, changes in intensity and homogeneity • CT scan ○ Defines arterial vessels and patency • Arteriography 483

Pancreas Transplantation

Surgical Aspects and Complications, Pancreas ○ Rarely used for diagnosis ○ Vascular narrowing or occlusion is detected • Contrast-enhanced magnetic resonance (MR) imaging ○ Noninvasive assessment of graft complications

Anastomotic Leaks and Intraabdominal Infections • CT scan ○ Differentiates anastomotic and duodenal stump leaks • Cystogram with bladder drainage

Graft Pancreatitis and Fluid Collection • CT scan • Ultrasonography

MACROSCOPIC

– Destruction of ductal and vascular structures

DIFFERENTIAL DIAGNOSIS Acute T-Cell-Mediated Rejection • Acinar and duodenal inflammation • Arterial and venular inflammation

Acute Antibody-Mediated Rejection • Donor-specific antibody positive • Interacinar capillary inflammation • C4d-positive staining in capillaries

Acute Infectious Pancreatitis and Abscess • Mostly neutrophils • Areas of necrosis and purulent exudate

Hemorrhagic Infarction • Extensive dark discoloration • Diffuse swelling • Soft and friable due to necrosis

Ischemic Infarction • Pale and somewhat shrunken • Vascular thrombi

Donor Duodenal Integrity

DIAGNOSTIC CHECKLIST Hemorrhagic or Ischemic Infarction • • • • • •

Vascular graft thrombosis Poor blood flow by imaging techniques Abdominal pain or distension Marked elevation of amylase, lipase, and glucose levels Symptoms of pancreatitis or peritonitis Pancreatic transplant biopsy or explant showing coagulative necrosis ± parenchymal hemorrhage

• Hemorrhagic and friable

MICROSCOPIC Histologic Features • Mild ischemic pancreatitis ○ Causes – Prolonged cold ischemia time – Ischemia-reperfusion injury ○ Delay in primary graft function – Elevated serum pancreatic enzymes – Hyperglycemia ○ Microscopic pathology – Focal acinar cell lysis – Occasional apoptosis – Flattening of acinar cells – Minimal inflammation – Islet cell swelling/vacuolization • PTx infarction and ischemic pancreatitis ○ Ischemic/coagulative lobular necrosis, patchy/diffuse – Extent dependent on degree of vascular occlusion – Focal interstitial hemorrhage ○ Septal edema and inflammation – Predominantly neutrophils ○ Septal and peripancreatic fat necrosis – Presence of foamy macrophages • Peripancreatitis ○ Active and chronic inflammation – Polymorphous infiltrate including lymphocytes, histiocytes, and plasma cells – Admixture of neutrophils and eosinophils – Peripancreatic, septal, and perilobular distribution ○ Focal fat necrosis ○ Septal fibroblastic proliferation and collagen deposition – Lobular dissection by bands of fibrosis – Relative preservation of lobular centers 484

Ischemic Pancreatitis: Peripancreatitis With Fat Necrosis • Similar clinical symptoms and imaging findings • Significant peripancreatic inflammation, fat necrosis, focal hemorrhage, and focal involvement of lobular/acinar tissue and ducts

SELECTED REFERENCES 1.

2. 3. 4. 5.

6.

7. 8. 9.

10. 11. 12.

13.

14.

Ryu JH et al: Pancreas transplant with duodeno-duodenostomy and caval drainage using a diamond patch graft: a single-center experience. Ann Transplant. 22:24-34, 2017 Laurence JM et al: Techniques of pancreas graft salvage/indications for allograft pancreatectomy. Curr Opin Organ Transplant. 21(4):405-11, 2016 Liu Y et al: Value of magnetic resonance imaging in evaluating the pancreatic allograft transplant complications. Abdom Imaging. 40(7):2384-90, 2015 Rogers J et al: Pancreas transplantation with portal venous drainage with an emphasis on technical aspects. Clin Transplant. 28(1):16-26, 2014 Tiong HY et al: Selection and preparation of the pancreas transplant recipient. In Srinivas T et al: Kidney and Pancreas Transplantation: A Practical Guide. Totowa: Humana. 201-9, 2011 Wee A et al: Pancreas transplantation: surgical techniques. In Srinivas T et al: Kidney and Pancreas Transplantation: A Practical Guide. Totowa: Humana. 249-58, 2011 Han DJ et al: Pancreas transplantation. Gut Liver. 4(4):450-65, 2010 Sollinger HW et al: One thousand simultaneous pancreas-kidney transplants at a single center with 22-year follow-up. Ann Surg. 250(4):618-30, 2009 Drachenberg CB et al: Banff schema for grading pancreas allograft rejection: working proposal by a multi-disciplinary international consensus panel. Am J Transplant. 8(6):1237-49, 2008 Federle MP et al: Radiology aspects. In Corry RJ et al: Pancreatic Transplantation. New York: Informa Healthcare USA Inc. 211-27, 2007 Larsen JL: Pancreas transplantation: indications and consequences. Endocr Rev. 2004 Dec;25(6):919-46. Review. Erratum in: Endocr Rev. 26(5):661, 2005 Humar A et al: Technical failures after pancreas transplants: why grafts fail and the risk factors--a multivariate analysis. Transplantation. 78(8):1188-92, 2004 Larson TS et al: Pancreas-after-kidney transplantation: an increasingly attractive alternative to simultaneous pancreas-kidney transplantation. Transplantation. 77(6):838-43, 2004 Orsenigo E et al: Urological complications after simultaneous renal and pancreatic transplantation. Eur J Surg. 168(11):609-13, 2002

Surgical Aspects and Complications, Pancreas

Bladder Drainage

Enteric Drainage

Complications Reflux pancreatitis

Duodenal fistula

Bladder fistula

Anastomotic bleeding

Anastomotic bleeding

Intestinal perforation

Chronic metabolic acidosis

Intestinal obstruction

Dehydration

Peritonitis

Pancreas Transplantation

Complications and Benefits of Exocrine Drainage of Pancreas Allograft

Urologic Complications Chemical cystitis

Lower incidence of urinary tract infections than bladder drainage

Recurrent hematuria Bladder calculi Urinary tract infections Prostatitis, urethritis, urethral structure Advantages > 90% of all PTx up to 1995

Majority of all PTx after 1997

Valuable in PTA and PAK for urinary amylase monitoring

Normal physiologic state

Leaks managed conservatively

Lower incidence of opportunistic infections Low metabolic complications

PAK = pancreas after kidney; PTA = pancreas transplant alone; PTx = pancreas transplant.

Comparison of Systemic vs. Portal Venous Endocrine Drainage Systemic/Iliac Vein

Portal Vein

Simple surgical technique

Creates physiologic state of insulin metabolism

Peripheral hyperinsulinemia

50% of insulin metabolized in liver

Increased atherosclerosis

Lower free cholesterol levels

Peripheral insulin resistance

Lower levels of very low-density lipoproteins

Normoglycemia

Normoglycemia

15. Becker BN et al: Simultaneous pancreas-kidney and pancreas transplantation. J Am Soc Nephrol. 12(11):2517-27, 2001 16. Sutherland DE et al: Lessons learned from more than 1,000 pancreas transplants at a single institution. Ann Surg. 233(4):463-501, 2001 17. Humar A et al: Decreased surgical risks of pancreas transplantation in the modern era. Ann Surg. 231(2):269-75, 2000 18. Stratta RJ et al: A prospective comparison of systemic-bladder versus portalenteric drainage in vascularized pancreas transplantation. Surgery. 127(2):217-26, 2000 19. Del Pizzo JJ et al: Urological complications of bladder-drained pancreatic allografts. Br J Urol. 81(4):543-7, 1998 20. Sugitani A et al: Surgical complications in 123 consecutive pancreas transplant recipients: comparison of bladder and enteric drainage. Transplant Proc. 30(2):293-4, 1998 21. Stratta RJ et al: Analysis of early readmissions after combined pancreaskidney transplantation. Am J Kidney Dis. 28(6):867-77, 1996 22. Gaber AO et al: Results of pancreas transplantation with portal venous and enteric drainage. Ann Surg. 221(6):613-22; discussion 622-4, 1995 23. Gruessner RW et al: Recipient risk factors have an impact on technical failure and patient and graft survival rates in bladder-drained pancreas transplants. Transplantation. 57(11):1598-606, 1994 24. Marsh CL et al: Combined hepatic and pancreaticoduodenal procurement for transplantation. Surg Gynecol Obstet. 168(3):254-8, 1989 25. Nghiem DD et al: Technique of simultaneous renal pancreatoduodenal transplantation with urinary drainage of pancreatic secretion. Am J Surg. 153(4):405-6, 1987

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Surgical Aspects and Complications, Pancreas Combined Kidney and Pancreas Transplant With Pancreatic Enteric Drainage

Pancreatic Transplant With Pancreatic Urinary Bladder Drainage via Donor Duodenum

Acute Ischemic Infarct of Pancreas Allograft

Pancreatic Allograft Infarction With Arterial Thrombosis

CT of Posttransplant Pancreatitis With Perigraft Fluid

Posttransplant Peripancreatitis

(Left) Graphic illustrates the surgical anatomy of a combined pancreas-kidney transplant with pancreaticenteric drainage. The donor iliac artery is anastomosed to the donor superior mesenteric and splenic arteries (inset). The venous drainage is to the recipient iliac vein. (Right) Graphic shows a pancreatic transplant with pancreaticurinary bladder drainage and donor duodenal segment. The recipient iliac artery is anastomosed to the donor iliac artery. The venous drainage is to the iliac vein.

(Left) H&E of an acute pancreatic transplant ischemic infarction demonstrates arterial occlusion ﬈ with diffuse eosinophilic staining and loss of nuclear staining due to cell death. (Courtesy B. Fyfe, MD.) (Right) Highmagnification view reveals a pancreatic transplant infarction with arterial thrombus formation. (Courtesy B. Fyfe, MD.)

(Left) Axial CECT shows diffuse swelling of the pancreatic allograft ſt and perigraft fluid st, consistent with posttransplant pancreatitis. (Right) Posttransplant peripancreatitis shows preserved underlying lobular acini ﬈ surrounded by partly hemorrhagic inflammation and fat necrosis.

486

Surgical Aspects and Complications, Pancreas CT With Peripancreatic Graft Fluid Collection (Left) Axial CECT demonstrates a pancreatic allograft st and renal allograft ﬇ in the iliac fossae. Both show normal parenchymal enhancement and no sign of infarction or rejection. There are several small and large pseudocysts ſt near the pancreatic allograft. These were aspirated and drained with US guidance. (Right) Axial CECT shows diffuse pancreatic allograft heterogeneity with decreased enhancement ſt surrounding perigraft fluid collections st.

Severe Posttransplant Peripancreatitis

Pancreas Transplantation

CT of Normal Renal and Pancreas Allograft With Few Pseudocysts

Total Ischemic Infarction of Pancreas Allograft (Left) Severe posttransplant peripancreatitis is characterized by edema, severe inflammation, and focal fat necrosis. (Right) H&E section of a pancreas transplant biopsy with dysfunction manifesting elevated enzymes and hyperglycemia is shown. Total ischemic infarction is visible with coagulative, lobular acinar necrosis with loss of nuclei and homogenized cytoplasm.

Ischemic and Focally Hemorrhagic Pancreatic Infarction

Total Hemorrhagic Pancreas Allograft Infarction (Left) H&E shows an ischemic and focally hemorrhagic infarction of attached donor duodenal mucosa ﬈ and submucosal duodenal glands below the hemorrhagic mucosal layer ﬊. (Courtesy B. Fyfe, MD.) (Right) H&E shows a total hemorrhagic infarction of attached donor duodenal mucosa with evidence of gastrointestinal bleeding. (Courtesy B. Fyfe, MD.)

487

Pancreas Transplantation

Acute Cellular Rejection, Pancreas KEY FACTS

ETIOLOGY/PATHOGENESIS • Alloreactive T cells against donor antigens

CLINICAL ISSUES • • • •

Acute graft failure Increased serum amylase Increased serum lipase Treating higher grades of acute cellular rejection (ACR) often requires anti-T-cell therapy in addition to corticosteroids

MICROSCOPIC

• Presence of intimal arteritis indicates at least grade II (moderate) ACR • Transmural arterial inflammation/fibrinoid necrosis seen in grade III (severe) ACR • C4d(-) in pure ACR ○ Positive if combined with AMR

TOP DIFFERENTIAL DIAGNOSES • • • •

Acute antibody-mediated rejection Peripancreatitis Posttransplant lymphoproliferative disorder Cytomegalovirus-associated pancreatitis

• Mixed mononuclear infiltrate within septal areas ○ With involvement of septal structures – Ducts – Veins – Nerves • Increasing degrees of acinar inflammation accompanied by acinar cell injury/necrosis

Ductulitis

Venulitis

Intimal Arteritis

Acinar and Septal Inflammation

(Left) Scattered lymphocytes ﬈ are intimately associated with ductal epithelium (ductulitis) in this case of grade I (mild) acute cellular rejection (ACR). (Right) Peripheral cuffing of a septal venule by mononuclear cells is shown with disruption ﬈ and inflammation ﬉ of the overlying endothelial cells (venulitis). Active venulitis &/or ductulitis indicate at least grade I (mild) ACR, according to Banff criteria.

(Left) H&E shows an intimal arteritis lesion with mononuclear cells lifting up the endothelium (endothelialitis) ﬉ of this muscular artery. Intimal arteritis correlates with at least grade II (moderate) ACR, according to Banff criteria. (Right) Active mononuclear inflammation fills the fibrous septa ﬇ and extends into the acinar tissue st, as shown in this example of grade II (moderate) ACR.

488

Acute Cellular Rejection, Pancreas

MICROSCOPIC

Abbreviations

Histologic Features

• Acute cellular rejection (ACR)

• Septa ○ Edema ○ Mononuclear cell infiltrate – Activated/blast-like lymphocytes [mostly CD4(+) and CD8(+) T cells], eosinophils (common), macrophages – Eosinophil-rich variants can occur – Late rejection: Plasma cells, B cells ○ Ductulitis: Inflammation of duct epithelium – Small intralobular ducts and larger septal branches – May have associated epithelial cell injury with cytoplasmic swelling, cell sloughing, loss of polarity, reactive nuclei ○ Venulitis: Inflammation of endothelium within septal veins ± endothelial injury ○ Neural/perineural infiltrates – Rare finding in needle biopsies • Acinar tissue ○ Acinitis: Inflammation of acinar lobules with permeation of basement membrane and close association with acinar epithelium – Similar cell composition as septal infiltrates – Focal: No more than 2 foci per lobule – Multifocal: ≥ 3 foci per lobule – Diffuse: Widespread, extensive ○ Acinar cell injury/necrosis – Vacuolization and swelling of cell cytoplasm – Cell dropout – Focal to multifocal to diffuse/confluent • Arteries ○ Intimal arteritis: Mononuclear cell infiltrates beneath arterial endothelium – Indicates ≥ grade II (moderate) ACR – Minimal: Focal process, usually involving small area of vessel lumen on cross section – Moderate to severe: Multiple vessels involved, or majority of vessel lumen involved on cross section – a.k.a. endothelialitis, endarteritis ○ May see marginated mononuclear cells along vascular endothelial surface – Often associated with active intimal arteritis elsewhere ○ Endothelial injury/activation – Swelling of cytoplasm – Reactive nuclear enlargement ○ Transmural inflammation – Indicates grade III (severe) ACR – Severe cases may have fibrinoid necrosis • Islets ○ May contain mononuclear infiltrates (isletitis) – Seen in higher grades of ACR

Synonyms • Acute T-cell-mediated rejection

Definitions • Active immunologic rejection to pancreatic allografts mediated by T cells ○ Grade I: Mild ○ Grade II: Moderate ○ Grade III: Severe

ETIOLOGY/PATHOGENESIS T-Cell-Mediated Rejection • Alloreactive T cells targeting donor antigens ○ MHC (HLA) ○ Non-MHC • Targets include following ○ Acinar epithelium ○ Duct epithelium ○ Venous and arterial endothelium

CLINICAL ISSUES Epidemiology • Incidence ○ More frequent in pancreas transplant alone than simultaneous pancreas-kidney transplant ○ Absolute incidence unknown in patients with simultaneous renal allografts – Diagnosis often based on presence of renal allograft rejection (surrogate for pancreas) and laboratory data

Presentation • Usually asymptomatic • Laboratory evidence of acinar cell injury ○ Increased serum amylase ○ Increased serum lipase • Abnormalities in exocrine/endocrine function ○ Decreased urine amylase – Bladder-drained grafts ○ Hyperglycemia – Severe rejection, late marker – Due to involvement of islets by inflammation • Graft tenderness ○ Severe cases

Treatment • Drugs ○ Pulse corticosteroid therapy ○ Anti-T-cell agents – Antithymocyte globulin

Prognosis • Better for grade I (mild) • Worse for grades II (moderate) and III (severe)

Pancreas Transplantation

TERMINOLOGY

ANCILLARY TESTS Immunohistochemistry • C4d(-) in interacinar capillaries (IAC) in pure T-cell-mediated rejection

489

Pancreas Transplantation

Acute Cellular Rejection, Pancreas ○ Cellular rejection cases with C4d likely represent superimposed acute or chronic antibody-mediated rejection (AMR) • Markers to evaluate cellular composition of inflammation ○ T-cell marker (CD3) – Useful to detect low-level infiltrates within acinar tissue, duct epithelium, vascular endothelium ○ Macrophage markers (CD68, CD163) – Variable degrees of macrophage infiltration of lobules may be present • Antiinsulin/antiglucagon ○ May be used to identify islet destruction by inflammatory cells

Immunofluorescence • C4d negative in IAC • No specific immunoglobulin or complement deposition • Requires frozen tissue

Electron Microscopy • Not generally performed

DIFFERENTIAL DIAGNOSIS Antibody-Mediated Rejection • • • •

Inflammation of IAC with neutrophils and macrophages C4d stains positive in IAC Minimal T cells within infiltrates Minimal acinitis

Peripancreatitis • Active fibroblastic response within associated connective tissue • Large numbers of macrophages

Posttransplant Lymphoproliferative Disorder • Nodular or expansile infiltrates, randomly distributed ○ No specific targeting of acinar tissue, minimal acinar cell injury • Atypical plasmacytoid B cells • No intimal arteritis (unless there is concomitant rejection) • Epstein-Barr virus associated (positive EBER)

Cytomegalovirus Pancreatitis • Epithelial or endothelial cells with characteristic cytopathic changes ○ Cellular enlargement ○ Intranuclear and cytoplasmic inclusions • Confirmatory cytomegalovirus immunohistochemistry

• Biopsies meeting criteria for ACR and AMR do occur and represent mixed rejection

Banff Grading of Acute Cellular Rejection • Normal ○ Absent inflammation or inactive septal, mononuclear inflammation not involving ducts, veins, arteries, or acini ○ No graft sclerosis ○ No atrophy/injury to acinar parenchyma • Indeterminate for ACR ○ Septal inflammation appears active – Activated/blastic lymphocytes, ± eosinophils – May see reactive stromal cells within septa ○ Overall features do not fulfill criteria for grade I (mild) ACR – No involvement of septal structures (no venulitis or ductulitis) – No acinar inflammation • Grade I: Mild ACR ○ Active septal inflammation (activated blastic lymphocytes and/or eosinophils) involving septal structures (venulitis, ductulitis) ○ Focal acinar inflammation – No more than 2 inflammatory foci per lobule ○ Absent or minimal acinar cell injury • Grade II: Moderate ACR ○ Multifocal acinar inflammation and spotty acinar cell injury/dropout – ≥ 3 inflammatory foci per lobule – Not confluent or diffuse ○ Minimal intimal arteritis (< 25% luminal compromise) ○ Requires differentiation from AMR • Grade III: Severe ACR ○ Diffuse (widespread/extensive) acinar inflammation with multicellular/confluent acinar cell injury and necrosis/dropout ○ Moderate to severe intimal arteritis (> 25% luminal compromise) &/or transmural arterial inflammation or fibrinoid necrosis ○ Requires differentiation from AMR

SELECTED REFERENCES 1.

2.

3.

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Multiple level sections should be examined to identify focal lesions ○ Ductulitis, intimal arteritis • Diagnosis/grading based primarily on routine H&E morphology ○ Immunohistochemistry for T cells (CD3) useful adjunct to detect focal or mild infiltrates • Severe acinar inflammation usually paired with marked acinar cell injury/necrosis 490

4.

5. 6. 7.

Loupy A et al: The Banff 2015 Kidney meeting report: current challenges in rejection classification and prospects for adopting molecular pathology. Am J Transplant. 17(1):28-41, 2016 Haas M et al: Banff 2013 meeting report: inclusion of C4d-negative antibodymediated rejection and antibody-associated arterial lesions. Am J Transplant. 14(2):272-83, 2014 Papadimitriou JC et al: Distinctive morphological features of antibodymediated and T-cell-mediated acute rejection in pancreas allograft biopsies. Curr Opin Organ Transplant. 17(1):93-9, 2012 Drachenberg CB et al: Banff schema for grading pancreas allograft rejection: working proposal by a multi-disciplinary international consensus panel. Am J Transplant. 8(6):1237-49, 2008 Drachenberg CB et al: The inflamed pancreas transplant: histological differential diagnosis. Semin Diagn Pathol. 21(4):255-9, 2004 Sutherland DE et al: Lessons learned from more than 1,000 pancreas transplants at a single institution. Ann Surg. 233(4):463-501, 2001 Papadimitriou JC et al: Histologic grading scheme for pancreas allograft rejection: application in the differential diagnosis from other pathologic entities. Transplant Proc. 30(2):267, 1998

Acute Cellular Rejection, Pancreas

Perineural Inflammation (Left) The infiltrate within the septa appears "active" with enlarged, blastic lymphocytes ſt and scattered plasma cells and rare eosinophils ﬈. Active septal inflammation indicates at least indeterminate ACR. (Right) Edema and active inflammation with prominent lymphocytes and eosinophils fill this septal area and surround a peripheral nerve segment ﬊. May be a rare finding in needle biopsies.

Acinar Inflammation

Pancreas Transplantation

Active Septal Inflammation

Acinar Inflammation and Cell Injury (Left) Dense mononuclear inflammation is shown involving acinar tissue with expansion of the interacinar space ﬇, close approximation with acinar epithelium st, and reactive nuclear enlargement of acinar epithelial cells ſt. (Right) Inflammation of acinar tissue with associated injury of acinar epithelium is shown, including vacuolization ﬉ and swelling ﬊ of cell cytoplasm. There are scattered areas of acinar cell dropout st.

Eosinophil-Rich Acinar Inflammation

CD3 (T-Cell) Immunohistochemical Stain (Left) H&E shows an eosinophil-rich infiltrate in this case of grade III (severe) ACR. Eosinophils are commonly seen in acute cellular rejection of the pancreas and can predominate in some cases. (Right) CD3 immunohistochemistry highlights several T lymphocytes ﬈ interacting with duct epithelium (ductitis). Although diagnosis/grading of ACR should be performed predominantly on routine morphology, CD3 stain can help identify low-level infiltrates.

491

Pancreas Transplantation

Antibody-Mediated Rejection, Pancreas KEY FACTS

ETIOLOGY/PATHOGENESIS • Donor-specific antibody (DSA) directed against HLA classes I and II antigens on endothelium • Activation of complement cascade • Microvascular inflammation with endothelial injury/activation • Ischemic tissue damage

CLINICAL ISSUES • Graft dysfunction manifested by ○ Elevated serum amylase ○ Elevated serum lipase • DSA detected in patient serum • Graft loss may be precipitous in severe antibody-mediated rejection (AMR) • Hyperglycemia in chronic AMR

• Interacinar edema and inflammation (neutrophilic or mixed) • Inflammation of interacinar capillaries (acute capillaritis) • Lobular interacinar capillaries ○ Positive for C4d by immunohistochemistry &/or immunofluorescence

TOP DIFFERENTIAL DIAGNOSES • Acute cellular rejection • Nonrejection-related graft thrombosis • Infections

MICROSCOPIC • Hemorrhage and necrosis in hyperacute AMR

Interacinar Inflammation With Capillaritis

Positive C4d Immunohistochemistry

Positive C4d Immunofluorescence

Intravascular Thrombus

(Left) Neutrophils are shown within interacinar capillaries ﬇ and between acini st in acute antibody-mediated rejection (AMR). Acinar cells are well preserved with only focal cytoplasmic vacuolization ſt. (Courtesy M. Troxell, MD, PhD.) (Right) Strong, diffuse staining of interacinar capillaries ﬈ for C4d is shown by immunohistochemistry. Diffuse staining is defined as involving > 50% of interacinar capillaries (IAC) within the biopsy tissue and is most strongly correlated with acute AMR.

(Left) Immunofluorescence for C4d shows strong, diffuse staining of the interacinar capillaries in this case of acute AMR. Only granular/linear interacinar capillaries staining is considered positive; staining of larger vessels (arteries and veins) is nonspecific. (Right) Vascular thrombosis may occur in hyperacute and acute AMR, representing an important cause of graft failure. Thrombosis associated with AMR should be distinguished from nonrejection thrombosis.

492

Antibody-Mediated Rejection, Pancreas

Abbreviations

• Serum amylase and lipase • Serum glucose • Urine amylase

• Antibody-mediated rejection (AMR)

Treatment

Synonyms

• • • •

• Humoral rejection

Definitions • Graft dysfunction and failure caused by antibodies directed toward donor antigens ○ Hyperacute AMR – Immediate dysfunction due to preformed antibodies ○ Acute AMR – Sudden dysfunction due to de novo antibodies ○ Chronic active AMR – Progressive dysfunction due to de novo antibodies with graft sclerosis

ETIOLOGY/PATHOGENESIS Donor-Specific Antibodies • Usually directed to HLA classes I and II antigens on endothelium • Other antibodies to ○ ABO blood group isoagglutinins ○ MHC class I-related chain A • Binding of antibody to antigens on endothelial surface ○ Fixation and activation of complement cascade – Classic complement pathway ○ Recruitment of leukocytes into microvasculature (interacinar capillaritis) ○ Initiation of clotting cascade ○ Impaired blood flow and ischemic tissue injury

CLINICAL ISSUES Epidemiology • Incidence ○ Unknown

Presentation • Hyperacute rejection ○ Immediate (≤ 1 hour) ○ Severe graft dysfunction – Coagulopathy – Rapid rise in serum amylase and lipase – Decreased urine amylase • Acute AMR ○ May be asymptomatic ○ Elevated serum amylase and lipase ○ Decreased urine amylase ○ Endocrine function variable • Chronic active AMR ○ Progressive graft dysfunction ○ Loss of glycemic control (hyperglycemia) ○ Decreased serum amylase and lipase – Levels may be undetectable

Laboratory Tests • Detection of donor-specific antibodies (DSA) in recipient serum

Plasma exchange (plasmapheresis) Increased immunosuppression Intravenous immune globulin Rituximab (anti-CD20 antibody)

Prognosis

Pancreas Transplantation

TERMINOLOGY

• Graft failure ○ Presence of DSA and positive C4d in biopsy associated with poor graft survival ○ Precipitous graft loss may occur in severe, acute AMR • Graft sclerosis with decreasing graft function

MACROSCOPIC General Features • Hyperacute rejection ○ Rapid graft hemorrhage and necrosis ○ Vascular thrombosis often prominent • Acute AMR ○ Swollen graft ○ May have areas of hemorrhage ○ May have vascular thrombi • Chronic active AMR ○ Graft often decreased in size ○ Fat necrosis ○ Fibrosis

MICROSCOPIC Histologic Features • Hyperacute rejection ○ Immediate changes (minutes) – Interacinar edema – Vascular congestion, hyperemia – Spotty acinar cell injury ○ Later changes (hours) – Confluent hemorrhagic necrosis of acini, islets, and ducts – Prominent neutrophilic infiltrates – Fibrinoid vascular necrosis – Vascular thrombosis • Acute AMR ○ Acinar and interacinar inflammation – Monocytic &/or neutrophilic infiltrates ○ Inflammation of interacinar capillaries – Early: Neutrophils often predominate – Later: Mixed or monocytic infiltrates ○ Interstitial edema ○ Interstitial hemorrhage ○ Acinar cell injury, necrosis – Mild (grade 1): Spotty – Moderate (grade 2): Multifocal – Severe (grade 3): Diffuse • Chronic active AMR ○ Features of acute AMR ○ Graft sclerosis 493

Pancreas Transplantation

Antibody-Mediated Rejection, Pancreas – Expansion of fibrous septa – Fibrous tissue between acini ○ Atrophy: Progressive loss of acinar cell epithelium

DIFFERENTIAL DIAGNOSIS Acute T-Cell-Mediated (Cellular) Rejection • Increased monocytic septal infiltrates • Increased acinar cell inflammation • Inflammation of septal structures ○ Ductulitis ○ Venulitis • Minimal interacinar capillaritis • No C4d capillary deposition • Hemorrhagic necrosis and thrombosis rare • Mixed (antibody- and cell-mediated) rejection can occur

Graft Thrombosis • Early vascular thrombosis: Primary graft failure ○ No capillary inflammation (capillaritis) – No active endothelialitis or vascular necrosis ○ No acinar inflammation ○ No C4d capillary deposition • Late vascular thrombosis ○ May be associated with acute T-cell-mediated rejection ○ Minimal interacinar capillaritis ○ No C4d capillary deposition

Infections • Constitutional symptoms may include ○ Fever ○ Nausea ○ Vomiting • Positive cultures or PCR for organisms (blood, pancreatic secretions) • Positive serology (viral pancreatitis) • Immunohistochemistry (i.e., CMV)

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Graft dysfunction ranging from immediate/severe (hyperacute) to progressive loss of function over time (chronic active AMR)

Pathologic Interpretation Pearls • Acinar and interacinar inflammation ○ Predominantly neutrophilic • Interacinar capillaritis • Positive C4d staining of interacinar capillaries • Hemorrhagic necrosis • Vascular thrombosis

Banff Grading of Acute/Active Antibody-Mediated Rejection • Diagnostic components: Histologic evidence of tissue injury, C4d positivity in interacinar capillaries, serologic evidence of DSA ○ 1 of 3 components: Requires exclusion of AMR ○ 2 of 3 components: Consider acute AMR ○ 3 of 3 components: Definite acute AMR • Histologic evidence of acute tissue injury 494

○ Grade I (mild acute AMR) – Architecture preserved – Mild interacinar infiltrates – Rare acinar cell damage ○ Grade II (moderate acute AMR) – Architecture preserved – Interacinar infiltrates – Dilated interacinar capillaries with capillaritis – Intimal arteritis – Acinar cell dropout ○ Grade III (severe acute AMR) – Architectural disarray – Interstitial hemorrhage – Multifocal/confluent necrosis – Transmural/necrotizing arteritis – Vascular thrombosis • C4d positivity in interacinar capillaries ○ ≤ 1% of acinar lobular surface for C4d immunohistochemistry

SELECTED REFERENCES 1.

Loupy A et al: The Banff 2015 Kidney meeting report: current challenges in rejection classification and prospects for adopting molecular pathology. Am J Transplant. 17(1):28-41, 2016 2. Becker LE et al: A single-center experience on the value of pancreas graft biopsies and HLA antibody monitoring after simultaneous pancreas-kidney transplantation. Transplant Proc. 47(8):2504-12, 2015 3. de Kort H et al: Diagnosis of early pancreas graft failure via antibodymediated rejection: single-center experience with 256 pancreas transplantations. Am J Transplant. 14(4):936-42, 2014 4. Haas M et al: Banff 2013 meeting report: inclusion of C4d-negative antibodymediated rejection and antibody-associated arterial lesions. Am J Transplant. 14(2):272-83, 2014 5. de Kort H et al: Pancreas transplantation, antibodies and rejection: where do we stand? Curr Opin Organ Transplant. 18(3):337-44, 2013 6. Papadimitriou JC et al: Distinctive morphological features of antibodymediated and T-cell-mediated acute rejection in pancreas allograft biopsies. Curr Opin Organ Transplant. 17(1):93-9, 2012 7. Drachenberg CB et al: Guidelines for the diagnosis of antibody-mediated rejection in pancreas allografts-updated Banff grading schema. Am J Transplant. 11(9):1792-802, 2011 8. de Kort H et al: Pancreas allograft biopsies with positive c4d staining and anti-donor antibodies related to worse outcome for patients. Am J Transplant. 10(7):1660-7, 2010 9. Rangel EB et al: Antibody-mediated rejection (AMR) after pancreas and pancreas-kidney transplantation. Transpl Int. 23(6):602-10, 2010 10. Drachenberg CB et al: Banff schema for grading pancreas allograft rejection: working proposal by a multi-disciplinary international consensus panel. Am J Transplant. 8(6):1237-49, 2008 11. Torrealba JR et al: C4d-positive interacinar capillaries correlates with donorspecific antibody-mediated rejection in pancreas allografts. Transplantation. 86(12):1849-56, 2008

Antibody-Mediated Rejection, Pancreas

Interstitial Hemorrhage (Left) Extensive tissue necrosis is shown in a case of severe, acute AMR. A central artery shows fibrinoid necrosis and an intraluminal thrombus ﬊. (Right) Interstitial hemorrhage ﬇ associated with loss of acinar epithelium is shown in acute AMR. On the left, the interacinar space is expanded by edema and mild inflammation ﬈.

Interacinar Capillaritis

Pancreas Transplantation

Graft Necrosis

Interacinar Inflammation With Atrophy (Left) Mononuclear infiltrates within interacinar capillaries (capillaritis) ﬉ are shown in acute AMR. The inflammatory infiltrate may consist of neutrophils, lymphocytes, or a mixture of these inflammatory cells. (Right) Mononuclear cells and neutrophils within capillaries ﬇ and interacinar connective tissue st are shown. This case of chronic active AMR shows marked loss of acinar epithelium with only scattered acinar cells remaining ﬈. (Courtesy M. Troxell, MD, PhD.)

Weakly Positive C4d Immunofluorescence

C4d Staining in Chronic Active AntibodyMediated Rejection (Left) Scattered interacinar capillaries show weak positivity for C4d by immunofluorescence in chronic, active AMR ſt. C4d may be falsely negative in chronic active AMR due to loss of interacinar capillaries and corresponding fibrosis. (Right) Although the acinar parenchyma appears atrophic, C4d immunohistochemistry highlights multiple positive capillaries ﬈ in this case of chronic active AMR. Staining of connective tissue fibers (lower right image st) is nonspecific.

495

Pancreas Transplantation

Chronic Allograft Rejection/Graft Sclerosis KEY FACTS

CLINICAL ISSUES

DIAGNOSTIC CHECKLIST

• Serum enzymes often remain normal • Loss of glycemic control (hyperglycemia) • Degree of fibrosis and atrophy correlate with risk for graft loss

• Chronic cellular and antibody-mediated rejection may coexist • Graft sclerosis grading ○ Mild ○ Moderate ○ Severe ○ Based on amount of fibrosis and degree of acinar cell loss • Chronic arteriopathy (similar to renal and cardiac allografts)

MICROSCOPIC • • • •

Progressive fibrosis of septal areas and acinar lobules Proportional acinar cell atrophy Allograft arteriopathy Islet cell fibrosis (late)

ANCILLARY TESTS • Trichrome stain to estimate amount of fibrosis • C4d stain to detect active antibody-mediated rejection

TOP DIFFERENTIAL DIAGNOSES • Recurrent diabetes mellitus • Chronic pancreatitis

Chronic Cellular Rejection

Moderate Graft Sclerosis

Severe Graft Sclerosis

Severe Chronic Allograft Arteriopathy

(Left) Loss of acinar epithelium is coupled with increased fibrosis ﬊ in this case of chronic cellular rejection. A sparse lymphocytic infiltrate is present ﬉. (Right) Trichrome stain highlights expansion of septal areas ﬊ and increased fibrous tissue within acinar parenchyma ﬉, corresponding to moderate (grade II) graft sclerosis.

(Left) Severe graft sclerosis with marked loss of acinar epithelium is shown. Mononuclear inflammation indicates ongoing cellular rejection. (Right) Severe chronic allograft arteriopathy is shown with marked fibrous intimal hyperplasia ﬇, disruption of the internal elastic lamina st, and an organizing thrombus ſt. (Courtesy A. Farris, MD.)

496

Chronic Allograft Rejection/Graft Sclerosis

Synonyms • Chronic T-cell-mediated rejection • Chronic antibody-mediated rejection

Definitions • Persistent T-cell- &/or antibody-mediated rejection, leading to chronic changes in allograft

ETIOLOGY/PATHOGENESIS T-Cell-Mediated Injury • Alloresponse to HLA antigens • Involves acini, ducts, and vasculature • Macrophages and eosinophils also participate

Antibody-Mediated Injury • Donor-specific antibody (usually against HLA class II antigens) • Involves interacinar capillaries, larger vessels • Antigen-antibody binding activates complement ○ C4d deposition in interacinar capillaries indicates ongoing activation • Repeated cycles of antibody-mediated endothelial injury and repair

○ Presence of endotheliitis indicates active rejection • Islets ○ Progressive fibrosis with collagen bands between cells (late feature) ○ Loss of endocrine cells, nonselective for cell type ○ No significant inflammation • Banff grading: Chronic allograft sclerosis ○ Grade I (mild) – Fibrosis occupies < 30% of tissue core – Expansion of fibrous septa – Acinar lobules have irregular contours ○ Grade II (moderate) – Fibrosis occupies 30-60% of tissue core – Atrophy involves periphery and center of acinar lobules ○ Grade III (severe) – Fibrosis occupies > 60% of tissue core – Only isolated acinar cells and islets remain

ANCILLARY TESTS Immunohistochemistry • C4d stain may be positive in interacinar capillaries with active antibody rejection component • Glucagon and insulin stains show identifiable α and β cells

Stimulation of Fibrosis

Trichrome Histochemistry

• Expansion of septal fibrous tissue • Collagen deposition around acini and within islets • Proportional loss of glandular components

• Used to estimate degree of fibrosis

CLINICAL ISSUES Presentation • Loss of glycemic control (hyperglycemia) ○ Due to progressive loss of β cells from pancreas, which secrete insulin • Serum amylase, lipase often remain within normal limits

Pancreas Transplantation

TERMINOLOGY

DIFFERENTIAL DIAGNOSIS Recurrent Diabetes Mellitus • No significant fibrosis of islets; may have isletitis • Selective loss of β cells with preservation of α cells

Chronic Pancreatitis • Differentiation from chronic cellular rejection problematic (overlapping features) • May have more mixed inflammation with PMNs if active

Prognosis • Higher grades of fibrosis and atrophy correlate with increased risk of graft loss • Chronic vascular changes (allograft arteriopathy) also impact graft survival

MICROSCOPIC Histologic Features • Septa ○ Expansion of fibrous tissue, loss of septal structures • Acini ○ Early: Encroachment of fibrosis at periphery of lobules ○ Late: Fibrosis progresses to center of lobules with collagen strands separating individual acini ○ Proportionate atrophy of acinar epithelium ○ Loss of interacinar capillaries ○ Variable mononuclear inflammation • Arteries ○ Allograft arteriopathy: Fibrous intimal hyperplasia with progressive luminal narrowing ○ Mononuclear cells within intima ○ May have superimposed vascular thrombosis

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Chronic cellular and antibody-mediated rejection may coexist • Fibrosis, atrophy occur 1st at periphery of acinar lobules and later involve central lobules • Grading of graft sclerosis (mild, moderate, severe) based on amount of fibrosis and acinar loss • Chronic arteriopathy similar to renal and cardiac allografts, may have superimposed thrombosis

SELECTED REFERENCES 1.

2.

3. 4.

Drachenberg CB et al: Banff schema for grading pancreas allograft rejection: working proposal by a multi-disciplinary international consensus panel. Am J Transplant. 8(6):1237-49, 2008 Drachenberg CB et al: Spectrum of histopathological changes in pancreas allograft biopsies and relationship to graft loss. Transplant Proc. 39(7):23268, 2007 Papadimitriou JC et al: Histological grading of chronic pancreas allograft rejection/graft sclerosis. Am J Transplant. 3(5):599-605, 2003 Drachenberg CB et al: Chronic pancreas allograft rejection: morphologic evidence of progression in needle biopsies and proposal of a grading scheme. Transplant Proc. 31(1-2):614, 1999

497

Pancreas Transplantation

Recurrent Diabetes Mellitus KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Recurrent autoimmune isletitis • Isolated pancreatic islet inflammation and β-cell loss in pancreas allotransplantation, mediated by autoantibodies, causing hyperglycemia

• Inflammatory pancreatic islet damage • Gradual destruction and selective loss of β cells • Predominantly infiltrated by CD4 &/or cytotoxic T-cell subset lymphocytes

ETIOLOGY/PATHOGENESIS

ANCILLARY TESTS

• Pure cell-mediated mechanisms not documented • Antibodies against glutamic acid decarboxylase (GAD65), tyrosine phosphatase-like molecule (IA-2), and zinc transporter 8 (ZnT8)

• Loss of insulin-positive/β cells • T-lymphocyte subset staining, CD3, CD8 • Loss of intracellular granulation in β cells

CLINICAL ISSUES • 50-70% of type 1 diabetes mellitus patients have 1 or more circulating autoantibodies to islet cells • History of autoimmune/type 1 diabetes • Elevated glucose levels • Low or absent insulin levels

TOP DIFFERENTIAL DIAGNOSES • Acute/active cellular rejection ○ Inflammation of septal and acinar tissue • Duct obstruction pancreatitis ○ Septal and lobular inflammation and fibrosis • Infections

Mononuclear Cell Inflammation

Insulin Immunohistochemistry

T-Cell-Rich Islet Inflammation

Moderate Lymphocytic Islet Cell Inflammation

(Left) Pancreatic transplant biopsy shows an islet infiltrated by mononuclear inflammatory cells with adjacent mild infiltration of the acini, associated with hyperglycemia and circulating IA-2 antibodies. (Right) The same pancreatic islet in a consecutive section from the same patient shows only occasional cells positive for insulin immunostaining ﬈, suggesting significant destruction of the insulinproducing cells.

(Left) Pancreatic transplant with detectable antiislet cell antibody, hyperglycemia, and marked infiltration by T lymphocytes is shown. (Right) Pancreatic transplant biopsy from a patient with autoimmune diabetes mellitus and positive antiislet cell antibody shows moderate inflammation of the islet located at the edge.

498

Recurrent Diabetes Mellitus

Abbreviations • Recurrent diabetes mellitus (rDM)

Synonyms • Recurrent autoimmune isletitis • Recurrent autoimmune diabetes mellitus (DM) • Recurrent diabetic insulitis

Definitions • Isolated pancreatic islet inflammation of whole-organ pancreas or islet allotransplantation ○ Mediated primarily by autoantibodies ○ Result in selective β-cell loss and insulin-dependent hyperglycemia despite immunosuppression

ETIOLOGY/PATHOGENESIS Animal Models • Autoimmune isletitis first shown in rats that spontaneously developed diabetes • Recurrent autoimmune islet cell inflammation also observed in experimental animals • Methods of immunologic tolerance prevented recurrence

Autoimmune Mechanisms • Primarily mediated by detectable circulating islet cell autoantibodies, often fixing complement ○ Represent active or memory autoimmune response to islet autoantigens ○ Autoantibodies precede onset of hyperglycemia – Partially or completely blocked by immunosuppression ○ Evoke progressive lymphocytic inflammation restricted to islets • T-cell-mediated mechanisms ○ Circulating islet autoreactive CD4(+) T cells detected ○ Autoantigen-specific T cells localized in tissue – CD8(+) cytotoxic T cells – Increased CD4(+) T cells • Autoimmune destruction of β cells is major histocompatibility complex restricted ○ Common in HLA identical transplant recipients without immunosuppression – Twins, siblings, parents ○ Partially or mostly prevented with full-dose antirejection therapy ○ Occurs less often in cadaveric donors or HLAmismatched grafts – Mainly with complement-fixing antibodies • Expression of HLA I or II antigens may be necessary for inflammatory response

Putative Autoantibodies • Islet cell-specific autoantibodies ○ Cytoplasmic islet cell antibodies (ICAs) – Occasionally positive in prepancreatic transplant patients – May be elevated following transplantation – Responds to immunosuppression after pancreas transplantation (PTx)

○ Antibodies against glutamic acid decarboxylase (GAD65) – Frequently present pretransplantation with rising titers after PTx – May persist after immunosuppression – Useful to monitor rDM and prognosis of graft ○ Antibodies against tyrosine phosphatase-like molecule (IA-2) – Rise in titers post transplantation – Represents late-stage marker of immune response – Signifies spreading of humoral response against β cells • Antiinsulin antibody • Serve as specific markers for antiimmune type 1 diabetes and rDM • Detectable autoantibodies pretransplant predict graft failure ○ Islet cell transplantation ○ Whole-pancreas allograft

Pancreas Transplantation

TERMINOLOGY

CLINICAL ISSUES Epidemiology • Incidence ○ Exact incidence of rDM is not known ○ 50-70% of type 1 DM patients have 1 or more circulating autoantibodies to islet cells – Present in pretransplant sera – Rising titers post transplant ○ Rarely, de novo autoimmune type diabetes with autologous islet cell transplant • Risk factors ○ History of autoimmune/type 1 diabetes ○ Low or no immunosuppression

Presentation • Develops as early as 6 weeks post transplant; can take years ○ Acute hyperglycemia ○ Chronic hyperglycemia • Laboratory testing ○ Detectable or rising titers of anti-ICAs – Radioligand assay (35 S-methionine) – Indicate recurrence of islet cell autoimmunity – Marker to monitor susceptibility to insulitis ○ Elevated glucose levels ○ Low or absent insulin levels

Treatment • No specific treatment • Continue routine immunosuppression ○ Nonspecific depletion of T cells

Prognosis • Islet cell inflammation leads to pancreas graft and islet allograft failure with rapid course

Causes of Chronic Hyperglycemia After PTx • Intermittent hyperglycemia ○ Acute rejection – Cases treated with high doses of immunosuppression • Chronic hyperglycemia ○ Graft failure – Acute or chronic rejection 499

Pancreas Transplantation

Recurrent Diabetes Mellitus – Pancreatitis – Ischemic damage of pancreas (e.g., thrombosis) ○ Insulin resistance with new-onset type 2 diabetes – Weight gain – Coexistent obesity – Genetic predisposition to diabetes ○ Immunosuppression-induced islet cell toxicity, leading to new-onset type 2 diabetes ○ Immune-mediated islet cell damage, leading to recurrent type 1 diabetes

MICROSCOPIC Histologic Features • Pancreatic islet damage, early biopsies ○ Progressive inflammatory infiltrate – Minimal: < 10 cells per islet – Mild to moderate: 11-55 cells per islet – Severe: > 55 cells per islet ○ Gradual destruction and selective loss of β cells – β-cell degranulation and lysis ○ Predominantly mononuclear cells – Majority cytotoxic T cells – Smaller population of macrophages ○ Prominent vascularity of islets • Pancreatic islet damage, late biopsies ○ Resolution of inflammation after β-cell loss ○ Intraislet capillaries not apparent ○ No significant residual fibrosis seen ○ α- and polypeptide-cell-rich islets remain • Generally no exocrine pancreatic tissue affected ○ Cell-mediated rejection &/or vascular endotheliitis can rarely coexist ○ May require positive serologic testing for definitive diagnosis of autoimmune isletitis • Vasculature is normal

ANCILLARY TESTS

• Focal, mild, or no isletitis • Preserved integrity of islets • Both α and δ cells present

Duct Obstruction Pancreatitis • Septal and lobular inflammation and fibrosis • Minimal or no isletitis

Infections Involving Mainly Acinar Tissue • Bacterial ○ Diffuse neutrophil-rich inflammation ○ Focal necrosis or abscess formation • Viral ○ Patchy lymphoplasmacytic infiltrate ○ Intranuclear viral inclusions ○ Immunohistochemistry confirms specific viruses • Fungal ○ Diffuse, acute, or granulomatous inflammation ○ Focal necrosis ○ Periodic acid-Schiff and silver stains localize fungal elements

SELECTED REFERENCES 1.

2. 3.

4.

5.

6.

7.

Immunohistochemistry • Loss of insulin-positive/β cells • Normal staining pattern for glucagon in islets • T-lymphocyte subset staining ○ Mainly CD3(+), CD8(+) cells with granzyme B ○ CD68(+) macrophages • Localization of HLA I antigens ○ Islet cells ○ Endothelial cells ○ Duct cells • ICAs not localized in islets

8.

9. 10.

11.

12. 13.

Electron Microscopy • Degenerative cytoplasmic changes in β cells • Loss of intracellular granulation • β-cell granules in macrophage lysosomes

DIFFERENTIAL DIAGNOSIS

14.

15. 16.

Acute/Active Cellular Rejection • Inflammation of septal and acinar tissue • Evidence of arterial endotheliitis 500

17.

Bellin MD et al: Development of autoimmune-mediated β cell failure after total pancreatectomy with autologous islet transplantation. Am J Transplant. 15(7):1991-4, 2015 Pugliese A et al: Recurrence of autoimmunity in pancreas transplant patients: research update. Diabetes Manag (Lond). 1(2):229-238, 2011 Ishida-Oku M et al: A case of recurrent type 1 diabetes mellitus with insulitis of transplanted pancreas in simultaneous pancreas-kidney transplantation from cardiac death donor. Diabetologia. 53(2):341-5, 2010 Vendrame F et al: Recurrence of type 1 diabetes after simultaneous pancreas-kidney transplantation, despite immunosuppression, is associated with autoantibodies and pathogenic autoreactive CD4 T-cells. Diabetes. 59(4):947-57, 2010 Velthuis JH et al: Accumulation of autoreactive effector T cells and allospecific regulatory T cells in the pancreas allograft of a type 1 diabetic recipient. Diabetologia. 52(3):494-503, 2009 Laughlin E et al: Recurrence of autoreactive antigen-specific CD4+ T cells in autoimmune diabetes after pancreas transplantation. Clin Immunol. 128(1):23-30, 2008 Xiang Z et al: CD4+ T cells are sufficient to elicit allograft rejection and major histocompatibility complex class I molecule is required to induce recurrent autoimmune diabetes after pancreas transplantation in mice. Transplantation. 85(8):1205-11, 2008 Lohmann T et al: Islet cell-specific autoantibodies as potential markers for recurrence of autoimmune type 1 diabetes after simultaneous pancreaskidney transplantation. Transplant Proc. 34(6):2249-50, 2002 Da Silva M et al: Combined analysis of autoantibodies against beta-cells for prediction of pancreas allograft failure. Transplant Proc. 32(8):2773, 2000 Thivolet C et al: Serological markers of recurrent beta cell destruction in diabetic patients undergoing pancreatic transplantation. Transplantation. 69(1):99-103, 2000 Jaeger C et al: Glutamic acid decarboxylase antibodies are more frequent than islet cell antibodies in islet transplanted IDDM patients and persist or occur despite immunosuppression. J Mol Med (Berl). 77(1):45-8, 1999 Roep BO et al: Auto- and alloimmune reactivity to human islet allografts transplanted into type 1 diabetic patients. Diabetes. 48(3):484-90, 1999 Esmatjes E et al: Recurrence of immunological markers for type 1 (insulindependent) diabetes mellitus in immunosuppressed patients after pancreas transplantation. Transplantation. 66(1):128-31, 1998 Drachenberg CB et al: Histologic findings in islets of whole pancreas allografts: lack of evidence for recurrent cell-mediated diabetes mellitus. Transplantation. 62(12):1770-2, 1996 Tydén G et al: Recurrence of autoimmune diabetes mellitus in recipients of cadaveric pancreatic grafts. N Engl J Med. 335(12):860-3, 1996 Bonifacio E et al: Identification of protein tyrosine phosphatase-like IA2 (islet cell antigen 512) as the insulin-dependent diabetes-related 37/40K autoantigen and a target of islet-cell antibodies. J Immunol. 155(11):541926, 1995 Sibley RK et al: Recurrent diabetes mellitus in the pancreas iso- and allograft. A light and electron microscopic and immunohistochemical analysis of four cases. Lab Invest. 53(2):132-44, 1985

Recurrent Diabetes Mellitus

Partial Loss of Insulin-Positive Cells in Islet (Left) Pancreatic transplant biopsy from a patient with positive antiislet cell antibody, hyperglycemia, and moderate elevation of pancreatic enzymes reveals marked infiltration by lymphocytes of an islet and periislet inflammation. (Right) Insulin immunostaining is partially present within the islet cells ﬈.

Variable T-Cell [CD3(+)] Infiltration of Islet in Recurrent Diabetes Mellitus

Pancreas Transplantation

Islet and Periislet Inflammation in Recurrent Autoimmune Diabetes

Partial Destruction of Islet by Inflammation (Left) Islet area contains abundant infiltrating inflammatory cells, including T lymphocytes that are positive for CD3 immunoperoxidase staining. (Right) There is intense lymphocytic infiltration within and around the islet with partial destruction of cells involving adjacent septal and acinar tissue ﬈.

Chronic Destruction of Islet With Recurrent Autoimmune Diabetes

Pancreas Allograft Rejection With Abundant Insulin-Positive Cells in Islets (Left) Pancreatic transplant failure in a patient with antiislet cell antibody and pancreas explantation specimen shows chronic fibrosing parenchymal inflammation, rare lymphocytes ﬈, and loss of islet cells. (Right) Pancreas transplant biopsy with grade 2 cellular rejection shows strong immunostaining for insulin and minimal islet infiltration by lymphocytes ﬈ for comparison.

501

Pancreas Transplantation

Islet Cell Toxicity and Islet Amyloid Deposition KEY FACTS

ETIOLOGY/PATHOGENESIS • • • •

Tacrolimus (FK506), cyclosporine A Inhibition of insulin synthesis by β cells Decreased insulin release by β cells Drug-induced peripheral resistance to insulin

CLINICAL ISSUES • Cause of postpancreatic transplant hyperglycemia • High levels of immunosuppressive agents needed for pancreas transplants

MICROSCOPIC • Islet cell toxicity ○ Islet cell swelling and vacuolization without inflammation ○ Nuclear hyperchromasia and fragmentation ○ No alterations in acinar, septal, and vascular tissue ○ Morphology correlates with functional changes and drug levels ○ Recovery of cells with normoglycemia

• Islet amyloid deposition in whole pancreas and clinical islet transplants ○ β-cell dysfunction and selective apoptotic loss ○ Confirmation by Congo red (+) stain

ANCILLARY TESTS • Loss of insulin-containing cells with normal glucagon staining in islets by immunohistochemistry • Electron microscopic findings in islet cell toxicity ○ Intracytoplasmic vacuolization, variable loss of insulinsecretory granules ○ Nuclear pyknosis/condensation ○ Focal cytoplasmic lipid inclusions

TOP DIFFERENTIAL DIAGNOSES • Ischemic islet cell vacuolization with concomitant focal acinar necrosis • Superimposed infections or inflammatory reactions of islets

Normal Acinus

Cytoplasmic Vacuolization

Amyloid Deposits

Congo Red-Positive Amyloid Deposits

(Left) Normal pancreatic acinar tissue contains a preserved islet cell aggregate ﬈ with pale or amphophilic cytoplasm and vesicular nuclei. (Right) Islet cells show variable cytoplasmic vacuolization and focal clearing ﬈ and rare pyknosis ﬉ in a pancreatic transplant biopsy with high tacrolimus levels (> 12 mg). Adjacent acini are preserved.

(Left) H&E in a patient with type 2 diabetes mellitus and simultaneous pancreas kidney transplant with islet amyloid deposition shows extensive replacement of islet cells (top) and focal deposits separating islet cells (bottom) without an inflammatory reaction. The surrounding acinar and septal tissue are uninvolved. (Courtesy B. Fyfe, MD.) (Right) Simultaneous pancreas kidney transplant with islet amyloid deposition was confirmed by Congo red stain ﬊. (Courtesy B. Fyfe, MD.)

502

Islet Cell Toxicity and Islet Amyloid Deposition

Abbreviations • Islet cell toxicity (ICT)

Definitions • Structural and functional abnormalities in islets of pancreatic transplantation following immunosuppressive therapy, commonly using cyclosporine or tacrolimus resulting in hyperglycemia

ETIOLOGY/PATHOGENESIS Potential Immunosuppressive Agents • Tacrolimus (FK506), cyclosporine A ○ Dose and duration dependent, tacrolimus > cyclosporine • Mycophenolate • Sirolimus (rapamycin) • Daclizumab (IL-2 receptor antagonist) • Glucocorticoids ○ Potentiate action of cyclosporine, tacrolimus

Pathogenetic Mechanisms • Inhibition of insulin synthesis by β cells ○ Reduction in synthesis of DNA, RNA, and protein ○ Drug binding to FK-binding protein-12 to inhibit calcineurin ○ Reduction in insulin mRNA transcription ○ Inhibition of NFAT2 in β cells • Decreased insulin release by β cells ○ Defective conversion of proinsulin to insulin ○ Interference in Ca²⁺ influx via voltage-dependent calcium channel • Drug-induced peripheral resistance to insulin ○ Changes in insulin-specific binding rates of RBC ○ Changes in affinities of insulin receptors ○ Loss of transcription factors for glucose sensing – e.g., PDX-11, MafA, NeuroD, FoxO1

CLINICAL ISSUES Epidemiology • Incidence ○ Exact incidence of ICT not known ○ Posttransplant diabetes and insulin resistance in other solid organ transplants range from 7-25% – Immunosuppressive protocols – Weight gain – Preexisting obesity – Genetic predisposition to diabetes ○ High immunogenicity of pancreatic transplant – Requires higher immunosuppression than kidney, liver, or heart transplants • Risk factors for ICT ○ Older donor age ○ Lower islet cell mass in donor pancreas ○ Preexisting glucose intolerance/diabetes ○ Concomitant glucocorticoid administration

Laboratory Tests • • • •

Elevated serum levels of immunosuppressants Elevated blood glucose levels, glucose intolerance Variable C-peptide levels Low serum insulin levels

Treatment • Safe minimization of cyclosporine or tacrolimus dosage • Lower steroid doses or steroid-free protocol • Change cyclosporine/tacrolimus to rapamycin

Pancreas Transplantation

TERMINOLOGY

Prognosis • Often reversible in early stages ○ Normalization of glucose levels ○ Resolution of pathologic changes in islets • Slower recovery with long-term therapy

MICROSCOPIC Islet Cell Toxicity • Islet cell damage toxicity: Mild, moderate, or severe ○ Varied swelling and vacuolization, cytoplasmic clearing in center of islets ○ Focal cell "drop-out" leaving empty space ○ Nuclear pyknosis and cytoplasmic features of apoptosis • No changes in pancreatic lobules, septa, or vasculature • Morphologic findings of islets correlate with functional changes and drug levels • Cellular recovery following lowered drug levels and achievement of normoglycemia

Islet Amyloid Deposition • Variable accumulation of interstitial acellular amorphous Congo red-positive deposit restricted to islets ○ Preserved α and β cells, initially ○ β-cell dysfunction and selective apoptotic loss ○ Increased islet amyloid polypeptide (IAPP) secretion, during hyperinsulinemia ○ Amyloid deposits proportional to hyperglycemia ○ Also develops in clinical islet cell transplants – Heparin use implicated to promote IAPP amyloid fibril formation

SELECTED REFERENCES 1. 2.

3. 4.

5.

6. 7.

Triñanes J et al: Deciphering tacrolimus-induced toxicity in pancreatic β cells. Am J Transplant. 17(11):2829-2840, 2017 León Fradejas M et al: Islet amyloid in whole pancreas transplants for type 1 diabetes mellitus (DM): possible role of type 2 DM for graft failure. Am J Transplant. 15(9):2495-500, 2015 Potter KJ et al: Amyloid formation in human islets is enhanced by heparin and inhibited by heparinase. Am J Transplant. 15(6):1519-30, 2015 Paty BW et al: Inhibitory effects of immunosuppressive drugs on insulin secretion from HIT-T15 cells and Wistar rat islets. Transplantation. 73(3):3537, 2002 Drachenberg CB et al: Islet cell damage associated with tacrolimus and cyclosporine: morphological features in pancreas allograft biopsies and clinical correlation. Transplantation. 68(3):396-402, 1999 Neto AB et al: Metabolic and ultrastructural effects of cyclosporin A on pancreatic islets. Transpl Int. 12(3):208-12, 1999 Fioretto P et al: Cyclosporine associated lesions in native kidneys of diabetic pancreas transplant recipients. Kidney Int. 48(2):489-95, 1995

Presentation • Acute or chronic hyperglycemia • Native or transplant renal dysfunction 503

Pancreas Transplantation

Intraabdominal and Opportunistic Infections KEY FACTS • Safe reduction in immunosuppression • Prophylaxis • Intraabdominal fungal infections associated with increased morbidity and mortality rates

ETIOLOGY/PATHOGENESIS • Wound and intraabdominal infection ○ Bacterial infections • Bloodstream infection • Postoperative urinary tract infection • Opportunistic infections ○ Bacterial ○ Fungal: > 90% Candida albicans ○ Viral – Cytomegalovirus (CMV) – Epstein-Barr virus (EBV) – Herpes ○ Parasitic: Strongyloides, Giardia

IMAGING • Ultrasonographic and CT findings ○ Abscess or fluid collection

MICROSCOPIC

CLINICAL ISSUES • High infection rate in pancreas transplant, up to 46% • Abdominal infections common in first 3 months • Treat with appropriate antibiotic therapy

• Acute purulent peripancreatitis • Acute infectious pancreatitis ○ Predominantly neutrophil inflammatory infiltrate • CMV infection ○ Eosinophilic intranuclear inclusions ○ Immunohistochemical stains positive for CMV ○ Donor duodenal or gastrointestinal CMV infection • Posttransplant lymphoproliferative disorder ○ Differentiate from acute cell rejection, test for EBV

CMV Infection

CMV Immunohistochemistry

Abscess

Posttransplant Lymphoproliferative Disorder and EBV(+) IHC

(Left) Pancreas allograft biopsy shows dysfunction 6 months post transplant. There are elevated IgM cytomegalovirus (CMV) antibody titers, acinar inflammation, and endothelial cell nuclei containing darkstaining inclusions ﬈. (Right) Septal capillary endothelial cell nucleus and cytoplasm ﬊ are positive for CMV by immunoperoxidase staining in a pancreatic transplant biopsy with known CMV infection.

(Left) Pancreas transplant explant shows a large parenchymal abscess with extensive necrotic material and focal peripheral hemorrhage at the abscess wall ﬉. (Right) Pancreas transplant explant in a 53year-old woman with graft dysfunction with standard immunosuppressive therapy, exclusively involved by extensive posttransplant lymphoproliferative disorder (top, H&E), is EBV(+) (bottom, IHC). Pancreatectomy was curative with no further transplantation. (Courtesy Cinthia Drachenberg, MD.)

504

Intraabdominal and Opportunistic Infections

Definitions • Surgical site and intraabdominal infections: Mainly bacterial &/or fungal infections involving superficial and deep abdominal locations • Postoperative blood stream infections are associated with intraabdominal infections, urinary tract infections (UTIs), or transplant pyelonephritis • Postoperative UTIs may involve upper or lower urinary tract and associate with immunosuppression, urologic instrumentation • Opportunistic infections primarily associated with immunosuppression caused by reactivation of virus or acquired infection • Epstein-Barr virus (EBV)-associated posttransplant lymphoproliferative disorder

ETIOLOGY/PATHOGENESIS Wound and Intraabdominal Infection • Bacterial infections ○ Escherichia coli ○ Pseudomonas aeruginosa ○ Enterococcus faecalis ○ Coagulase-negative staphylococci ○ Antimicrobial-resistant strains • Posttransplant period range: 2-76 days • Susceptible patients ○ Diabetes mellitus ○ Overweight • Intraabdominal infection ○ 54% bacterial ○ 15% fungal, mainly Candida species ○ 21% combined bacterial and fungal

○ ○ ○ ○

EBV Herpes simplex virus Adenovirus BK virus, in relation to kidney transplantation

Potential Sources of Infection • Hospital/nosocomial • Community acquired • Opportunistic infections ○ Activation of latent viruses

Modes of Infection • Surgical complication • Anastomotic leakage ○ Donor duodenum to bladder drainage ○ Donor duodenum to enteric drainage • Acute bacterial pancreatitis • Infection of fluid collection

CLINICAL ISSUES Epidemiology • Overall high infection rate in PTx, up to 46% • Common cause of relaparotomy • Abdominal infections in 12-27% of PTx ○ Common in first 3 months post transplantation ○ Can be localized (abscess) or occasionally generalized (peritonitis) ○ Bacterial &/or fungal

Presentation

• Catheter-related infection ○ Long-term indwelling intravenous catheter ○ Short-term central venous catheter • Bacterial infections ○ Staphylococcus aureus – Coagulase-negative staphylococci ○ Gram-negative bacilli ○ Usually 1st month after transplantation ○ Higher incidence in enteric drained pancreas transplant (PTx)

• General ○ Fever ○ Purulent wound drainage ○ Wound dehiscence ○ Septicemia, if severe • Abdominal findings ○ Diarrhea ○ Distension/ileus ○ Acute abdomen ○ Symptoms of peritonitis • Viral infection ○ Single or multisystemic ○ Elevation of viral antibody titers ○ Elevation of serum viral load by PCR ○ Elevation of pancreatic enzymes ○ Gastrointestinal hemorrhage

Postoperative UTI

Laboratory Tests

• Commonly seen with simultaneous pancreas-kidney transplantation (SPK) • Often in bladder-drained PTx • Bacterial infection ○ E. coli

• Leukocytosis • Microbiologic study ○ Wound/abscess culture ○ Blood culture ○ Urine culture

Opportunistic Infections

Risk of Infections

• Bacterial • Fungal: > 90% Candida albicans • Viral ○ Cytomegalovirus (CMV) – Common in mismatched patients (e.g., CMV-negative patient/CMV-positive donor)

• Similar risks in SPK, pancreas after kidney, pancreas transplant alone • Older donor age • Recipient obesity • Diabetes mellitus • Concomitant renal failure

Bloodstream Infection

Pancreas Transplantation

TERMINOLOGY

505

Pancreas Transplantation

Intraabdominal and Opportunistic Infections • • • •

Long cold ischemia time Contamination of donor duodenum Increased in enteric drainage Graft thrombosis/ischemia

Treatment and Prevention of Infections • Appropriate antibiotic therapy • Safe reduction in immunosuppression • Prophylaxis ○ Perioperative ○ Postoperative ○ Antibacterial, fungal, and viral • Majority require pancreatic graft explant to preserve patient survival

Fungal Pancreatitis

Ultrasonographic Findings

• Common sites are surgical wounds and abdominal cavity • Factors contributing to candidiasis ○ Older donor age less favorable ○ Enteric drainage (21%) • Associated with higher morbidity and mortality • Pathologic findings ○ Acute peripancreatitis or necrotizing pancreatitis ○ Acute inflammation with neutrophils ○ Focal granulomatous inflammation ○ Commonly yeast and hyphal forms of Candida species – Best identified by PAS and GMS stains

• Abscess formation or fluid collection

Chronic Pancreatitis

CT Findings

• Chronic pancreatitis (CP) results from recurrent or persistent acute pancreatitis ○ Chronic duct inflammation ○ Epithelial proliferation ± metaplastic changes ○ Periductal fibrosis ○ Compressed or distorted ducts ○ Focal dystrophic calcification ○ Extensive lobular or acinar atrophy ○ Septal and lobular fibrosis ○ Usually islet cell hypertrophy and preservation ○ Can have focal islet cell destruction or atrophy ○ Islet cells definitively localized by immunohistochemistry

Prognosis • Intraabdominal fungal infections associated with increased morbidity and mortality rates • Recurrent infection in retransplant • Most result in transplant failure or removal of PTx

IMAGING

• • • •

Location and extent of infection Presence of abscess Peripancreatic fluid collection Guidance for aspiration of fluids/abscess

MACROSCOPIC General Features • Acute infectious peripancreatitis ○ Partly or completely covered by purulent exudate • Acute necrotizing pancreatitis ○ Focal abscess formation • Hemorrhagic or ischemic infarction complicated by infection

MICROSCOPIC Acute Purulent Peripancreatitis • • • •

Pyemic exudate on surface ± bacterial or fungal colonies Inflammatory cell invasion of capsule Focal perilobular and septal inflammation Primarily composed of neutrophils

Posttransplant Lymphoproliferative Disorder • Involves 1-3% of PTx recipients ○ Develops few weeks to several years after transplantation ○ Commonly related to EBV infection ○ Most are B lymphocyte in origin with polymorphous or monomorphous appearance ○ Usually random areas of pancreas allograft involved

DIFFERENTIAL DIAGNOSIS

Acute Infectious Pancreatitis

Posttransplant Ischemic Pancreatitis

• Patchy or extensive parenchymal inflammation ○ Septal and lobular inflammation ○ Acinar, ductal, and islet involvement • Predominantly neutrophil inflammatory infiltrate ○ Areas of necrosis with abscess formation ○ Bacterial or fungal colonies

• Increased or decreased serum amylase and lipase levels • Elevated serum glucose depending on severity • Coagulative necrosis of acinar tissue ○ Septal neutrophil infiltrate • Interstitial edema, hemorrhage, fat necrosis

Cytomegalovirus Infection • Acute pancreatitis suggesting tissue invasion ○ CMV infection affecting epithelial, endothelial, and stromal cells 506

○ Marked cell enlargement with dark intranuclear inclusions ○ Immunohistochemical stains positive for CMV ○ Mild, focal venous endotheliitis ○ Focal acinar damage ○ Secondary pancreatic abscess, infarction, and graft loss • Donor duodenal or gastrointestinal CMV infection ○ CMV intranuclear inclusions in endothelial and epithelial cells ○ Mucosal inflammation and ulceration

Pancreatic Duct Obstruction • • • •

Elevated serum amylase and lipase levels Normal serum glucose levels Elevated glucose in CP Ductal and focal inflammation ○ Lymphocytes, neutrophils

Intraabdominal and Opportunistic Infections

Clinical

Acute Rejection

Acute Pancreatitis

Elevation in serum amylase/lipase

(+)

(+)

Distribution

Septal, acinar

(+) peripancreatic septa, periphery of lobules

Intensity

Variable

Variable

Type of inflammation

Mononuclear cells

Neutrophils/mixed with mononuclear cells

Inflammatory cells

Activated predominantly T-cell subsets

Neutrophils

Sometimes eosinophil rich

Karyorrhexis

Macrophages

Necrosis

Pancreatic lobules

Activated T-cell infiltration

Neutrophils in lobules and septa

Ductitis

(+) by mononuclear cells

(+) neutrophils

Isletitis

(+) in severe cases

(+) in severe cases

Venulitis

(+)

(+)

Arteritis

(+) endotheliitis, vasculitis

(-)

Fat necrosis

(-)

(+)

• Mild acinar edema and inflammation

Peripancreatitis or Fluid Collection • Constitutional symptoms, peritonitis, or local pain from infection • Elevated serum amylase & lipase levels • Normal serum glucose levels • Polymorphous inflammation ○ Lymphocytes, neutrophils, plasma cells • Periphery of lobules; septal • Preserved acini

• Normal serum amylase and lipase levels • Serum autoantibodies to islet cells, such as ○ GAD65 ○ IA-2 ○ ICA ○ ZnT8 • Selective islet cell inflammation by lymphocytes and macrophages • Inflammation resolved on late stages • Loss of β cells

DIAGNOSTIC CHECKLIST

Chronic Rejection • Elevated serum amylase and lipase levels • Elevated serum glucose levels • Septal, acinar, and vascular inflammation ○ Mainly lymphocytes, plasma cells, macrophages • Acinar atrophy, septal fibrosis, loss of duct, transplant arteriopathy

Bacterial or Fungal Infection • • • • • •

Generalized or localized symptoms of infections Peritonitis, duodenal cuff perforation Elevated serum amylase and lipase Mainly neutrophils with purulent material, microabscesses Granulomatous inflammation ± central necrosis Septal and acinar locations of inflammation

Pancreas Transplantation

Differential Diagnosis of Rejection/Pancreatitis

Clinical and Pathologic Features • • • •

Systemic/constitutional symptoms Wound and intraabdominal symptoms Acute purulent pancreatitis CMV infection in tissue

SELECTED REFERENCES 1.

2.

3.

Cytomegalovirus Pancreatitis

4.

• Elevated serum amylase and lipase • Patchy, septal, or acinar inflammation

5.

Posttransplant Lymphoproliferative Disorder

6.

• Asymptomatic initially • Elevated serum amylase and lipase levels • Poly- to monomorphic lymphoblasts, plasma cells, occasional eosinophils • Low- to high-grade lymphoma

7.

8.

Schachtner T et al: Simultaneous pancreas/kidney transplant recipients are predisposed to tissue-invasive cytomegalovirus disease and concomitant infectious complications. Transpl Infect Dis. 19(5), 2017 Shah AP et al: Incidence and outcomes of cytomegalovirus in pancreas transplantation with steroid-free immunosuppression. Clin Transplant. 29(12):1221-9, 2015 Munivenkatappa RM et al: Pancreas. In Liapis H et al: Pathology of Solid Organ Transplantation. Heidelberg: Springer-Verlag Berlin Heidelberg. 37192, 2011 Drachenberg CB et al: The inflamed pancreas transplant: histological differential diagnosis. Semin Diagn Pathol. 21(4):255-9, 2004 Klassen DK et al: CMV allograft pancreatitis: diagnosis, treatment, and histological features. Transplantation. 69(9):1968-71, 2000 Knight RJ et al: Risk factors for intra-abdominal infection after pancreas transplantation. Am J Surg. 179(2):99-102, 2000 Drachenberg CB et al: Epstein-Barr virus-related posttransplantation lymphoproliferative disorder involving pancreas allografts: histological differential diagnosis from acute allograft rejection. Hum Pathol. 29(6):56977, 1998 Smets YF et al: Infectious disease complications of simultaneous pancreas kidney transplantation. Nephrol Dial Transplant. 12(4):764-71, 1997

Recurrent Autoimmune Isletitis/Diabetes Mellitus • Elevated serum glucose levels, insulin dependent

507

Pancreas Transplantation

Intraabdominal and Opportunistic Infections Pancreas Allograft With Infected Necrotic Tissue

CT of Peripancreatic Abscess in Allograft

Acute Infectious Pancreatitis

Acute Necrotizing Pancreatitis With Abscess

Chronic Fibrosing Pancreatitis

Chronic Fibrosing Pancreatitis

(Left) Axial contrast-enhanced CT in a patient with infected necrosis shows gas and fluid ſt replacing the pancreatic allograft in the right iliac fossa with a fistulous tract to the skin st confirmed at resection. (Right) Coronal CT reconstruction shows a fluid collection with an enhancing rim ſt representing an abscess surrounding the pancreatic allograft st in the right lower quadrant with communication ﬇ to the duodenum, consistent with dehiscence. Poorly enhancing renal allograft ﬊ due to rejection is also seen.

(Left) Pancreas transplant explant shows acute infectious pancreatitis with infiltrating neutrophils within the lobules and acini leading to significant disruption and epithelial injury. (Right) Pancreas transplant explant demonstrates acute necrotizing pancreatitis with fibrinous purulent exudate. This can be seen in the peripancreatic area and within the parenchymal tissue forming an abscess.

(Left) Pancreas transplant explant reveals chronic pancreatitis 1 year later with marked fibrosing ductal and septal inflammation. (Right) Pancreas transplant explant reveals chronic pancreatitis with inflammatory infiltrates in the septa. Pancreatic lobules also show mild to moderate atrophic changes. While a history of intraabdominal infection was given, there was no definite evidence of rejection.

508

Intraabdominal and Opportunistic Infections

CMV Immunohistochemistry (Left) Pancreas transplant biopsy shows a few acini ﬉ with chronic fibrosing parenchymal lymphocytic inflammation and scattered intranuclear CMV inclusions within a stromal cell ﬈. (Right) Pancreatic transplant biopsy with immunoperoxidase stain for CMV in a consecutive section stains positive in the intranuclear inclusion ﬊.

Fibrosing Pancreatic Infarct With CMV Infection

Pancreas Transplantation

Stromal Cell Intranuclear CMV Inclusion

Acinar Inflammation and CMV (Left) Low-magnification view of a pancreatic transplant biopsy with dysfunction and high CMV antibody titers demonstrates areas of parenchymal infarction ﬈, fibrous septa, and lobular inflammation. (Right) Highmagnification view of a pancreas transplant biopsy at the edge of infarction ﬊ reveals acinar inflammation in this patient with active CMV infection.

Intranuclear CMV Inclusion

CMV Immunohistochemistry (Left) Pancreas transplant biopsy in a patient with CMV infection shows an acinar cell ﬈ and a stromal cell ﬉ with dark-staining intranuclear CMV inclusions. (Right) Immunoperoxidase study for CMV demonstrates positive staining within inflamed acinar nuclei and cytoplasm ﬊ surrounded by inflamed fibroconnective tissue.

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SECTION 10

Vascularized Composite Allotransplantation

History of Vascularized Composite Allotransplantation

512

Allograft Rejection Acute T-Cell- and Antibody-Mediated Rejection Chronic Rejection

514 522

Vascularized Composite Allotransplantation

History of Vascularized Composite Allotransplantation

TERMINOLOGY Abbreviations

•

• Vascularized composite allotransplantation (VCA) • Composite tissue allotransplantation (CTA)

Definitions • Vascularized composite allografts ○ Transfer of peripheral tissues, including skin, muscle, nerve, tendon, and bone as functional unit (e.g., hand) to replace nonreconstructable tissue defects – Unlike organ transplants, most VCAs are □ Visible □ Contain highly immunogenic skin □ Require innervation to function ○ VCA field resulted from transplantation and microsurgery advances • In USA, VCA included within regulations covered by Organ Procurement and Transplantation Network in July 2013 ○ "Vascularized composite allograft" refers to body part that is – Vascularized and requires blood flow by surgical connection of blood vessels to function after transplantation □ Contain multiple tissue types – Recovered from human donor as anatomical/structural unit – Transplanted into human recipient as anatomical/structural unit – Minimally manipulated (i.e., processing that does not alter original relevant characteristics of organ relating to organ’s utility for reconstruction, repair, or replacement) – For homologous use (replacement or supplementation of recipient’s organ with donor organ that performs same basic function) – Not combined with another device – Susceptible to ischemia □ Only stored temporarily and not cryopreserved – Susceptible to allograft rejection □ Generally requires immunosuppression

CHRONOLOGY AND EVOLUTION

•

•

•

•

•

•

•

•

•

Timeline • AD 348 ○ 1st historical indication of VCA – Painting of Saints Cosmas and Damian transplanting Ethiopian Moor's limb in place of gangrenous limb • 1912 ○ Alexis Carrell – Awarded Nobel Prize in Physiology or Medicine "in recognition of his work on vascular suture and transplantation of blood vessels and organs" • 1915 ○ Harold Gilles – Joined Royal Medical Corps and became interested in plastic surgery of face, focusing on aesthetics, making patients similar to how they looked before injury • 1954 ○ Joseph E. Murray 512

•

•

•

– 1st successful kidney transplant in identical twins – Performed in USA 1960 ○ Peter Medawar and Sir Frank Macfarlane Burnet – Awarded Nobel Prize in Physiology or Medicine "for discovery of acquired immunological tolerance" 1964 ○ 1st hand transplant; performed in Ecuador – Graft lost within 3 weeks after surgery 1967 ○ Erle E. Peacock – 1st report of composite flexor tendon allograft 1990 ○ Joseph E. Murray and E. Donnall Thomas – Awarded Nobel Prize in Physiology or Medicine for "their discoveries concerning organ and cell transplantation in treatment of human disease" 1996 ○ Gunther O. Hofmann – 1st vascularized knee transplant – Performed in Germany 1998 ○ Jean-Michel Dubernard – 1st hand transplant after introduction of cyclosporine; performed in Lyon, France □ Graft lost in 2001 ○ Marshal Strome – 1st vascularized larynx transplant performed 1999 ○ Warren Breidenbach – 1st hand transplant in USA; performed in Louisville, KY □ Currently longest functional hand transplant 2000 ○ Wafa Fageeh – 1st transplant of human uterus, from living donor; performed in Saudi Arabia ○ Malaysia – Upper extremity transplant on 28-day-old neonate 2003 ○ University of Miami – 1st cases of abdominal wall transplant reported 2004 ○ The Hospital for Sick Children, Toronto, Canada – 1st lower extremity transplant, in 3-month-old ischiopagus twin 2005 ○ Jean-Michel Dubernard – 1st face transplant; performed in Lyon, France ○ Weilie Hu – 1st penis transplant; Guangzhou, China – Graft was removed 2 weeks later due to psychological trauma 2007 ○ 1st international consensus conference in VCA at 9th Banff Conference on Allograft Pathology in Spain – Banff CTA 2007 classification for skin allograft pathology was formalized – Now serves as standard for diagnosis of VCA rejection 2008 ○ Maria Siemionow

History of Vascularized Composite Allotransplantation

CLINICAL ISSUES Complications • Rejection ○ Most patients have at least 1 episode of rejection ○ Acute rejection often reversible with immunosuppressive modulation • Immunosuppressive complications ○ Infection ○ Posttransplant lymphoprolypherative disease ○ Graft-vs.-host disease (GVHD) • Metabolic disease ○ High levels of steroid use • Psychological factors ○ Lack of integration of graft with patient's self-identity • Death

○ Infections ○ Drug toxicity ○ Insect bites ○ GVHD ○ Allergic or irritant contact dermatitis ○ Eosinophilic cellulitis ○ Other causes • Classification of rejection ○ Banff CTA 2007 classification for skin allograft technology ○ Systemic pathologic component scores from skincontaining vascularized composite allografts

SUMMARY • VCA made significant progress in last 2 decades • Similar to other organ transplants, VCA faces challenges ahead, including ○ Problem of chronic rejection ○ Specific natural history of these transplants • 1st consensus conference in VCA gave rise to Banff CTA 2007 classification for skin allograft pathology ○ Serves as standard for diagnosis of VCA rejection ○ Component score approach from skin-containing VCA to complement Banff now developed • VCA is under definition of solid organ transplant • Unlike other transplants, most VCAs require innervation to function • As partner in transplantation, VCA should advance with focus on ensuring best outcomes for VCA recipients, both present and future

Vascularized Composite Allotransplantation

– 1st face transplant in USA (Cleveland, OH) • 2011 ○ Ömer Özkan and Munire Erman Akar – 1st reported clinical pregnancy after uterine transplantation; performed in Turkey ○ Pedro Cavadas – 1st reported adult lower extremity transplantation; performed in Valencia, Spain • 2012 ○ University of California, Davis – Larynx transplant performed ○ Ankara, Turkey – 1st quadruple extremity transplantation • 2014 ○ South Africa – 1st long-term phallus transplant ○ Mexico – 1st pediatric bilateral hand transplant • 2015 ○ Sweden – 1st live birth from uterine transplantation reported ○ USA – Bilateral hand transplant in 8-year-old boy • 2016 ○ Massachusetts General Hospital – 1st phallus transplant in USA ○ Cleveland Clinic – 1st deceased donor uterine transplant in USA ○ Baylor University Medical Center – 1st living donor uterine transplant in USA • Currently ○ > 200 patients worldwide have received VCA – > 75 upper extremity transplants – 6 knee transplants – 16 larynx transplants – 17 uterus transplants – 3 phallus transplants – > 38 abdominal wall transplants – > 30 face transplants ○ With increasing frequency of VCA, fewer cases reported

SELECTED REFERENCES 1.

Rosales I et al: Systematic pathological component scores for skin-containing vascularized composite allografts. Vascularized Composite Allotransplantation. epub, 2017 2. Shores JT et al: Outcomes after hand and upper extremity transplantation. J Mater Sci Mater Med. 28(5):72, 2017 3. Schneider M et al: Vascularized composite allotransplantation: a closer look at the banff working classification. Transpl Int. 29(6):663-71, 2016 4. Shanmugarajah K et al: The effect of MHC antigen matching between donors and recipients on skin tolerance of vascularized composite allografts. Am J Transplant. 17(7):1729-1741, 2016 5. Sicard A et al: An integrated view of immune monitoring in vascularized composite allotransplantation. Curr Opin Organ Transplant. 21(5):516-22, 2016 6. Madariaga ML et al: Immunomodulatory strategies directed toward tolerance of vascularized composite allografts. Transplantation. 99(8):15907, 2015 7. Lorenz RR et al: Total laryngeal transplant explanted: 14 years of lessons learned. Otolaryngol Head Neck Surg. 150(4):509-11, 2014 8. Nasir S et al: Lessons learned from the first quadruple extremity transplantation in the world. Ann Plast Surg. 73(3):336-40, 2014 9. Cendales L et al: Implementation of vascularized composite allografts in the United States: recommendations from the ASTS VCA Ad Hoc Committee and the Executive Committee. Am J Transplant. 11(1):13-7, 2011 10. Cendales LC et al: The Banff 2007 working classification of skin-containing composite tissue allograft pathology. Am J Transplant. 8(7):1396-400, 2008

Rejection • Differential diagnosis of skin rejection 513

Vascularized Composite Allotransplantation

Acute T-Cell- and Antibody-Mediated Rejection KEY FACTS

ETIOLOGY/PATHOGENESIS • Immunologic injury by cellular infiltrate composed of mostly T cells

CLINICAL ISSUES • Experience is still limited ○ Face, upper extremity, trachea, abdominal wall, uterus, penis • Clinical observation of skin important in early detection of rejection ○ Pink discoloration/rash initially ○ Erythematous macules progressing to red, scaly, infiltrated lichenoid papules

MICROSCOPIC • TCMR ○ Commonly occurs (> 80% of VCAs) ○ Inflammation extends to involve dermal stroma, epidermis, and adnexal structures

○ Perivascular inflammation becomes more dense and involves more vessels with increasing rejection ○ Keratinocyte apoptosis and eventually necrosis – Hyperkeratosis, hypergranulosis, and acanthosis eventually result as condition progresses to chronicity ○ Graded 0, I, II, III, or IV according to Banff criteria ○ Vasculitis can be seen as primary process (related to rejection) or secondary to ulceration • AMR ○ Rarely documented ○ Isolated cases of C4d(+), donor-specific antibody (+) acute rejection

TOP DIFFERENTIAL DIAGNOSES • • • • •

Chronic rejection Infection Insect bites Drug reactions/toxicity Allergic or irritant contact dermatitis

Grade I Acute T-Cell-Mediated Rejection

Grade II Acute TCMR

Grade III Acute TCMR

Grade IV Acute TCMR

(Left) In grade I acute T-cellmediated rejection (TCMR), there is mild perivascular inflammation ﬈, which can be recognized most often as mononuclear cells around small dermal vessels. (Right) At least grade II acute TCMR is present with severe perivascular inflammation ﬈. This could be considered higher grade if more than mild epidermal or adnexal involvement can be found.

(Left) Inflammation is present at the dermal-epidermal junction ﬈, leading to keratinolysis and detachment of the epidermis (i.e., bulla formation). This can be considered at least grade III TCMR due to this epidermal involvement. (Right) A nonhuman primate 140 days after transplantation has necrosis of the epidermis ﬉, which can be considered grade IV TCMR. Also present are findings seen in lower grades of TCMR such as perivascular inflammation ﬈.

514

Acute T-Cell- and Antibody-Mediated Rejection

Definitions • Sudden onset or short-term injury and morphologic changes in vascularized composite allografts (VCAs) due to alloimmune injury ○ VCA preferred over "composite tissue allograft" – Since "tissue" is regulated by FDA • T-cell-mediated rejection (TCMR) • Antibody-mediated rejection (AMR)

ETIOLOGY/PATHOGENESIS T-Cell-Mediated Rejection • Most common form of rejection (> 80% of hand and face VCAs) • Cellular infiltrate composed of mostly T cells ○ CD4 to CD8 (+) cell ratio ranges from 1.5:1 to 3:1 ○ T cells form perivascular cuffs – Result in epidermal and adnexal dyskeratosis ○ T cells of recipient origin, supported by expression of recipient HLA class 1 – 10-20% with T-regulatory cell phenotype [FOXP3(+)] ○ TIA1(+) cytotoxic T cells ○ Following may also be present – CD20(+) B cells – Plasma cells – Eosinophils – Mast cells – CD68(+) cells of macrophage/histiocyte lineage • HLA-DR and adhesion molecules may be upregulated in endothelial dermal vessel cells and epidermal keratinocytes ○ Changes not specific for rejection

Antibody-Mediated Rejection • Initially disputed, since C4d rarely detected • More recent evidence clearly shows potential of AMR, albeit rare ○ Shown in face VCA with C4d deposition and donorspecific antibodies (DSA) • May be more important as form of late rejection

CLINICAL ISSUES Presentation • Skin lesions ○ Pink discoloration/rash ○ Erythematous macules progressing to red, scaly, infiltrated lichenoid papules – Grade I rejection typically unremarkable clinically □ Can less often present with macules – Grade II rejection often presents with erythematous macules □ Sometimes slightly scaly ○ Vesicles ○ ± limb edema ○ Onychomadesis (shedding of nails) ○ Desquamation ○ Ulceration &/or necrosis • Clinical involvement should be assessed as ○ < 10%

○ 10-50% ○ > 50% of VCA

Treatment • Drugs ○ Calcineurin inhibitors ○ Steroids ○ Belatacept ○ Antithymocyte globulin (ATG), a.k.a. Thymoglobulin ○ Topical tacrolimus and clobetasol under study

Prognosis • Early mild rejection typically reversible with antirejection therapy ○ Large data sets are not readily available to establish longterm consequences

MACROSCOPIC

Vascularized Composite Allotransplantation

TERMINOLOGY

General Features • Careful clinical observation of skin important ○ Erythema, edema ○ Lichenoid, scaly maculopapules ○ Pallor ○ Ulceration, sloughing • Report of skin lesions may be present and known by pathologist at time of microscopic examination

MICROSCOPIC Histologic Features • TCMR ○ 1st well-developed criteria devised at 9th Banff Conference on Allograft Pathology (La Coruña, Spain, on June 26, 2007), termed Banff CTA-07 ○ Dermal and epidermal lymphocytic infiltration shows progression closely paralleling Banff grading system – Perivascular inflammation becomes more dense and involves more vessels with increasing rejection – Inflammation extends to involve dermal stroma, epidermis, and adnexal structures □ Adnexal involvement occurs at moderate to marked levels of rejection □ Lymphocyte exocytosis: Lymphocytes in epidermis rather than dermis □ Hypodermis can also be involved, most notably in more severe cases – Keratinocyte apoptosis and necrosis □ Marker of severe rejection ○ Lymphocytic inflammatory cell infiltrate – Inflammation can be mixed and include neutrophils – Eosinophils can be seen ○ Keratinocyte changes – Basal layer keratinocyte vacuolization – Hyperkeratosis □ Increase in thickness of anuclear layers of stratum corneum (most superficial layer of epidermis) □ Can be compact and orthokeratotic (with anuclear keratin layer) – Dyskeratosis is abnormal keratinization with shrunken, eosinophilic cells, and sometimes basophilic nuclear remnants 515

Vascularized Composite Allotransplantation

Acute T-Cell- and Antibody-Mediated Rejection – Hypergranulosis □ Hyperplasia of stratum granulosum (layer with coarse basophilic cytoplasmic granules, typically 1-5 cells thick) – Acanthosis □ Increased thickness of epidermis caused by hyperplasia or hypertrophy of spinous layer (eosinophilic layer just above basal layer that has prominent intercellular connections) – Saw-tooth appearance of dermal-epidermal junction – Apoptosis and necrosis □ Necrotic keratinocytes can be present as colloid or cytoid bodies within lower epidermal layers – Adnexal structure keratinocytes can have same changes □ Eccrine sweat glands most notably involved ○ Epithelial intracellular edema (referred to by term "spongiosis") – Both dermal and epidermal spongiosis can be present – May result in vesicle formation ○ Blisters in superficial layers with epidermal desquamation ○ Interface dermatitis in lichenoid pattern can be seen but may denote nonrejection etiology – Lichenoid (subepidermal) band hugs basal epidermal layer ○ Arteritis can be seen as primary process (related to rejection) or secondary to ulceration – Endarteritis (intima) mononuclear cells (analogous to Banff type II TCMR in kidney) – Transmural inflammation/necrosis (analogous to Banff type III TCMR in kidney) – Factors favoring rejection-induced vasculitis □ Absent history of trauma □ Vessels away from ulcer are also involved □ Multifocal involvement, including vessels of different sizes and depths within dermis ○ Thrombi in microvasculature correlated with adverse outcome and sometimes, but not always, with C4d and DSA ○ Underlying tissues such as muscle can have inflammation and resultant injury, but changes are less specific and less thoroughly studied ○ Mucosa-containing allografts can have similar changes to skin allografts, even though most of experience described above is from skin-containing allografts • AMR ○ Not defined in Banff CTA-07 and not yet clear ○ Limited published data indicates C4d deposition in dermal microvasculature can be indicative of AMR – C4d sometimes associated with microthrombi and graft loss ○ In analogy with renal transplant pathology, expect that some cases may have arterial inflammation and may lack C4d

ANCILLARY TESTS Immunohistochemistry • Stain for C4d to assess possibility of AMR

516

Immunofluorescence • Stain for C4d to assess possibility of AMR

Gene Expression • Possible role in diagnosis • Experimental studies suggest transcripts for CCL7, IL18, and IL1bβ may distinguish rejection from other types of inflammation

DIFFERENTIAL DIAGNOSIS Cellular Vs. Antibody-Mediated Rejection • Currently, most acute rejection considered to be primarily cell-mediated, but antibody-mediated rejection likely plays role • C4d used as marker of AMR in other organs ○ Does not have well-defined pattern of deposition that can be used to diagnose AMR • Antidonor HLA antibody positivity can raise level of suspicion for antibody-mediated component ○ History of sensitization can also be useful – Crossmatch results – Panel reactive antibody – Prior transfusions – Prior transplants

Chronic Rejection • Fibrosis can be caused by number of overlapping nonspecific causes, including both rejection and nonrejection causes • Histologic features of chronic rejection ○ Vascular narrowing – Myointimal proliferation ○ Adnexal loss ○ Skin and muscle atrophy ○ Deep tissue fibrosis ○ Nail changes

Infections • Cultures often needed • Special stains may be helpful • Fungal infections are particularly problematic ○ Dermatophyte infections can also have mild dermal perivascular inflammation ○ PAS stain useful to identify fungal organisms ○ Sabouraud medium useful to culture fungal organisms • Viral eruptions ○ Can have appearance of Banff grade I rejection both histologically and clinically – Can clinically present with macular (morbilliform) lesions – Can have mild lymphocytic perivascular infiltrates, which are also present in rejection ○ Can also mimic Banff grade II rejection – Can have dermal perivascular infiltrates that are more dense than grade I rejection • Viral infections can have red blood cell extravasation, feature less often seen in rejection

Acute T-Cell- and Antibody-Mediated Rejection

• Cells composing posttransplant lymphoproliferative disorder (PTLD) are typically overtly malignant on morphologic examination • Lymphocyte subset markers (e.g., CD3 for T cells, CD20 for B cells) can help this distinction since most PTLD due to Epstein-Barr virus (EBV)-driven B-cell proliferation • EBV can be detected by ancillary methods ○ EBV-encoded RNA (EBER) in situ hybridization ○ Immunohistochemistry for EBV latent membrane protein • Immunoglobulin gene rearrangement assay detects monoclonal rearrangement of immunoglobulin gene through molecular methods (e.g., PCR) • Experience thus far is limited in VCA patients

Insect Bites • Can contain high numbers of eosinophils in dermal infiltrate ○ Insects and other causes of eosinophilic dermatitis are considered to be important differential diagnostic considerations in acute rejection

Drug Reactions/Toxicity • Drug and viral eruptions are typically not limited to allografted skin • Drug reaction (or rash) with eosinophilia and systemic symptoms (DRESS) syndrome • Severe drug eruptions (e.g., toxic epidermal necrolysis) can sometimes lead to epidermal necrosis, mimicking grade 4 rejection ○ Dermal infiltrate is more superficial and sparse in these drug eruptions

Lichenoid Dermatoses • Lichen striatus, lichen planus, erythema multiforme, lichenoid lupus erythematosus, lichenoid drug eruptions, and lichenoid graft-vs.-host disease (GVHD) • Melanin incontinence can be seen in these disorders and not typically seen in VCA rejection

Allergic or Irritant Contact Dermatitis • Can be from topical application of medications or cosmetics • Contact dermatitis can be limited to VCA (e.g., hand or face), making resolution of differential diagnosis difficult • Dermal infiltrates and epidermal spongiosis ± vesiculation

Cutaneous Pseudolymphoma • Cutaneous pseudolymphomas have dense dermal lymphocytic infiltrate that occasionally contains eosinophils • Mixture of T cells and B cells present • Etiology often unclear

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Vasculitis (endarteritis and transmural inflammation): Important features of rejection but not yet included in Banff VCA schema • Recommend scoring individual components analogous to Banff schema for kidney allografts ○ Epidermal infiltrate, injury, dermal infiltrate, vascular inflammation, C4d deposition, chronic vascular lesions • Note specimen adequacy ○ ≥ 1 punch biopsy (4 mm) taken from most reddened &/or indurated but apparently viable area of involved skin ○ Sample should include adnexa, dermis, subcutaneous tissue, and vessels

Sentinel Flaps • Used for diagnostic biopsy when undesirable to biopsy clinical graft (e.g., face) • Good correlation with rejection in clinical graft

SELECTED REFERENCES 1.

2.

3.

GVHD • Limited experience with GVHD in VCA • GVHD has potential to occur in VCA due to presence of bone marrow in many VCAs, particularly hand VCAs ○ Forms vary from acute to chronic • NIH consensus document has laid forth features of chronic GVHD • Chronic forms include ○ Lichen planus-like skin changes ○ Sclerotic – Avascular collagen replaces papillary and reticular dermis – Hyperkeratosis – Flattening of rete ridges – Basal cell layer vacuolization – Lymphocytic infiltration – Epidermal melanin incontinence ○ Morpheic (localized scleroderma) ○ Fasciitis

Vascularized Composite Allotransplantation

Posttransplant Lymphoproliferative Disorder/Lymphoma

4.

5. 6.

7. 8.

9.

10. 11. 12. 13. 14.

Ng ZY et al: Graft vasculopathy in composite tissue allografts: literature review and clinicopathological study in a non-human primate model. In press, 2017 Kanitakis J et al: Capillary thrombosis in the skin: a pathologic hallmark of severe/chronic rejection of human vascularized composite tissue allografts? Transplantation. 100(4):954-7, 2016 Kueckelhaus M et al: Vascularized composite allotransplantation: current standards and novel approaches to prevent acute rejection and chronic allograft deterioration. Transpl Int. 29(6):655-62, 2016 Rosales I et al:. Systematic pathological component scores for skincontaining vascularized composite allografts. Am J Transplant. 16 (suppl 3), 2016 Schneider M et al: Vascularized composite allotransplantation: a closer look at the banff working classification. Transpl Int. 29(6):663-71, 2016 Weissenbacher A et al: Donor-specific antibodies and antibody-mediated rejection in vascularized composite allotransplantation. Curr Opin Organ Transplant. 21(5):510-5, 2016 Kueckelhaus M et al: Utility of sentinel flaps in assessing facial allograft rejection. Plast Reconstr Surg. 135(1):250-8, 2015 Wolfram D et al: Differentiation between acute skin rejection in allotransplantation and T-cell mediated skin inflammation based on gene expression analysis. Biomed Res Int. 2015:259160, 2015 Morelon E et al: Immunological issues in clinical composite tissue allotransplantation: where do we stand today? Transplantation. 93(9):855-9, 2012 Cendales LC et al: The Banff 2007 working classification of skin-containing composite tissue allograft pathology. Am J Transplant. 8(7):1396-400, 2008 Kanitakis J: The challenge of dermatopathological diagnosis of composite tissue allograft rejection: a review. J Cutan Pathol. 35(8):738-44, 2008 Swearingen B et al: Science of composite tissue allotransplantation. Transplantation. 86(5):627-35, 2008 Dubernard JM et al: Outcomes 18 months after the first human partial face transplantation. N Engl J Med. 357(24):2451-60, 2007 Cendales LC et al: Composite tissue allotransplantation: classification of clinical acute skin rejection. Transplantation. 81(3):418-22, 2006

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Acute T-Cell- and Antibody-Mediated Rejection 2007 Banff Working Classification of Skin-Containing Vascularized Composite Allograft Pathology Grade

Pathologic Features

Comments

0

No (or rare) inflammatory infiltrates

Corresponds to normal skin

I

Mild: Mild perivascular infiltration, no involvement of overlying epidermis

Lymphocytic infiltrate is typically only in upper dermis No involvement of adnexa Treatment not defined

II

III

Moderate: Moderate to severe perivascular inflammation ± mild epidermal &/or adnexal involvement (limited to spongiosis and exocytosis)

Infiltrate is expansive

No epidermal dyskeratosis or apoptosis

Cellular infiltrate can be mixed (e.g., including neutrophils) and not limited to lymphocyte

Severe: Dense inflammation and epidermal involvement with epithelial apoptosis, dyskeratosis, &/or keratinolysis

Inflammation forms nodules surrounding blood vessels and epidermal appendages (e.g., sweat glands in particular) Interface dermatitis occurs in lichenoid pattern Interface inflammation/dermatitis is important feature to identify, as it may relate to severity of rejection or may signal nonrejection etiology

IV

Necrotizing acute rejection: Frank necrosis of epidermis or other skin structures

Inflammation occasionally contains numerous eosinophils but this is not included in classification

Worldwide VCA experience is limited. Additional features, particularly of the vessels, need to be added (endarteritis, thrombi, C4d). Not all that occurs in a vascularized composite allograft is due to rejection. Specific differential diagnoses to consider include infections (particularly fungal), drug toxicity (e.g., topical steroids or other drugs), posttransplant lymphoproliferative diseases/lymphoma, insect bites, graft-vs.-host disease, allergic or irritant contact dermatitis, and eosinophilic cellulitis. Cendales LC et al: The Banff 2007 working classification of skin-containing composite tissue allograft pathology. Am J Transplant. 8(7):1396-400, 2008. 15. Shulman HM et al: Histopathologic diagnosis of chronic graft-versus-host disease: National Institutes of Health Consensus Development Project on Criteria for Clinical Trials in Chronic Graft-versus-Host Disease: II. Pathology Working Group Report. Biol Blood Marrow Transplant. 12(1):31-47, 2006 16. Kanitakis J et al: Pathological score for the evaluation of allograft rejection in human hand (composite tissue) allotransplantation. Eur J Dermatol. 15(4):235-8, 2005 17. Kanitakis J et al: Clinicopathologic features of graft rejection of the first human hand allograft. Transplantation. 76(4):688-93, 2003 18. Cendales LC et al: Hand transplantation. Hand Clin. 17(3):499-510, x, 2001 19. Dubernard JM et al: Human hand allograft: report on first 6 months. Lancet. 353(9161):1315-20, 1999

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Perivascular Inflammation (Left) Dense perivascular inflammation ﬇ indicates at least grade II acute TCMR. Grade II TCMR can have spongiosis or exocytosis but only mild epidermal involvement. Although not well represented in this section, if adnexal or epidermal dyskeratosis or keratinolysis could be established, the case would be considered grade III TCMR. (Right) An early sprinkling of perivascular inflammation ﬈ is present in this case of grade I TCMR.

Panniculitis

Vascularized Composite Allotransplantation

Grade II TCMR

Perivascular T Cells (Left) Penis vascularized composite allograft (VCA) shows focal mononuclear cells in the junction of fat ﬈ and the dermis. (Courtesy I. Rosales, MD.) (Right) Perivascular T cells are highlighted by CD3 immunohistochemistry. This density of perivascular inflammation ﬈ is considered at least grade II TCMR.

Perivascular T Cells

Macrophages in Perivascular Infiltrate (Left) CD3 immunohistochemistry illustrates numerous perivascular T cells. (Right) Early perivascular inflammation can be highlighted with immunohistochemistry. This is a CD68 stain, which illustrates that the inflammation actually consists of a mixture of CD3(+) T cells and CD68(+) monocyte/macrophages ﬈ in this case of grade I acute cellular rejection. A CD3 stain of the same case showed slightly more T cells in the same distribution.

519

Vascularized Composite Allotransplantation

Acute T-Cell- and Antibody-Mediated Rejection

Epidermal Infiltrate and Apoptosis

Epidermal Infiltrate

Extensive Epidermal Infiltrate

Inflammation in Sweat Ducts

Grade IV Epidermal Rejection

Epidermal Necrosis

(Left) Pig VCA with focal mononuclear infiltrates in the epidermis ſt accompanied by occasional apoptosis st and single cell necrosis ﬈ is shown. Pig skin is very similar to human skin and is useful for research studies. (Courtesy I. Rosales, MD.) (Right) A penis VCA shows very focal individual lymphocytes invading the epidermis ſt. This was considered Banff grade I TCMR. (Courtesy I. Rosales, MD.)

(Left) This biopsy of the skin from a pig VCA shows extensive mononuclear infiltrate in the epidermis ſt obscuring the normal interface at the epidermal basement membrane. (Courtesy I. Rosales, MD.) (Right) This biopsy from a penis VCA shows invasion of the sweat ducts by lymphocytes similar to, but more extensive than, those in the epidermis. (Courtesy I. Rosales, MD.)

(Left) This sample of pig VCA shows inflammation throughout the epidermis ﬈ with frank necrosis ﬇ and formation of microabscesses ſt. (Courtesy of I. Rosales, MD.) (Right) Necrosis of the epidermis is present ﬈. This case has a paucity of inflammatory cells, raising the possibility that this necrosis could be from a cause other than rejection (e.g., vascular compromise).

520

Acute T-Cell- and Antibody-Mediated Rejection

Endarteritis With Focal Necrosis of Artery (Left) Higher power view of the dense perivascular inflammation shows lymphocytes that appear somewhat "activated." In this case, some of the lymphocytes are also present in a vessel with a size intermediate between a small artery and arteriole ſt. (Right) Arteritis ﬉ is accompanied by perivascular inflammation ﬈ in this case of TCMR in a nonhuman primate 63 days after transplantation of a composite allograft. Arteritis is expected to be added to the Banff classification in the next revision.

Endarteritis

Vascularized Composite Allotransplantation

TCMR With Endarteritis

Endarteritis in TCMR (Left) A pig VCA shows endarteritis of a small subcutaneous artery. Endarteritis is not yet included in the Banff VCA schema, but is most likely a manifestation of TCMR. (Courtesy I. Rosales, MD.) (Right) This biopsy from a human penis VCA shows endarteritis with mononuclear cells in the intima of a small artery extending through the media ſt. C4d was negative. The rejection responded to Thymoglobulin. (Courtesy I. Rosales, MD.)

Fibrinoid Necrosis

C4d Deposition in Dermal Microvasculature (Left) Fibrinoid necrosis ﬉ is present in this case of TCMR that also had other evidence of TCMR. (Right) This VCA from a nonhuman primate underwent complete rejection at about 7 months. The sample shows widespread C4d deposition similar to that described in a patient with a face VCA. (Courtesy I. Rosales, MD.)

521

Vascularized Composite Allotransplantation

Chronic Rejection KEY FACTS

TERMINOLOGY

IMAGING

• Slowly developing late rejection in vascularized composite allografts (VCAs) due to alloimmune injury

• Useful to demonstrate arterial lesions

ETIOLOGY/PATHOGENESIS

• Atrophy of epidermis, adnexae • Dermal fibrosis • Chronic allograft vasculopathy (CAV); arterial intimal hyperplasia • Bones, joints, and bone marrow tend to be spared

• • • •

T cell mediated Antibody mediated: Limited evidence Allograft vasculopathy Thrombosis

MICROSCOPIC

CLINICAL ISSUES

ANCILLARY TESTS

• • • • • • • •

• C4d may be positive

Develops ~ 5-10 years post transplant insidiously Psoriasiform plaques, purpura, bruising Scleroderma-like changes, atrophy Nail, hair loss Hypo-/hyperpigmentation Necrotic ulceration Donor specific antibodies sometimes present Late graft loss may be inevitable with current therapy

TOP DIFFERENTIAL DIAGNOSES • Donor disease • Ischemia due to technical issues • Infection

DIAGNOSTIC CHECKLIST • Biopsies may not be deep enough to detect vascular lesions

Chronic Allograft Vasculopathy

Chronic Allograft Vasculopathy

Muscle Atrophy

Sparing of Nerve in Chronic Rejection

(Left) An artery has intimal fibrosis ſt compatible with chronic allograft vasculopathy. An adjacent vein is thrombosed ﬇. (Right) This nonhuman primate composite allograft artery has intimal thickening, likely from chronic allograft vasculopathy. Normally, the intima ﬉ is in close proximity to the media ﬇, but this case displays marked thickening of the intimal layer.

(Left) Atrophic skeletal muscle fibers ﬊ and chronic inflammation ﬈ consistent with chronic rejection are present in a longstanding nonhuman primate composite allograft. (Right) In a longstanding nonhuman primate vascularized composite allograft, chronic inflammation ﬈ is present in the soft tissue; however, a nerve fiber ﬊ is not inflamed. This allograft had evidence of rejection in other tissue compartments, demonstrating that nerves are typically preserved in composite allograft rejection.

522

Chronic Rejection

Definitions • Slowly developing late morphologic changes in vascularized composite allografts (VCAs) due to alloimmune injury • Chronic allograft vasculopathy (CAV) narrowing of lumen of artery due to neointimal proliferation

ETIOLOGY/PATHOGENESIS Alloimmune Reaction to Donor Antigens • T cell mediated • Antibody mediated: Limited evidence

Secondary Ischemia • Allograft vasculopathy • Thrombosis

CLINICAL ISSUES Presentation • Develops ~ 5-10 years post transplant ○ Insidious, may arise with few past acute rejection episodes • Associated with decreased immunosuppression ○ Nonadherence or reduction due to neoplasia • Appearance ○ Psoriasiform plaques ○ Purpura, bruising ○ Scleroderma-like changes, atrophy ○ Nail, hair loss ○ Hypo-/hyperpigmentation ○ Necrotic ulceration

Laboratory Tests

• Dermis ○ Fibrosis, thinning ○ Tertiary lymphoid organs, lymphoid follicles • Blood vessels ○ Intimal hyperplasia of graft arteries and stenosis (CAV) – Similar to solid organ allografts ○ Thrombosis of arteries • Muscles ○ Atrophy – Occurs particularly in intrinsic muscles – Fatty degeneration, possibly due to denervation • Bones ○ Decreased bone trabecular density • Joints ○ Often spared • Bone marrow ○ Often spared, replaced with recipient marrow

ANCILLARY TESTS Histochemistry • Trichrome stains useful to demonstrate fibrosis

Immunohistochemistry • C4d reported positive in dermal microvasculature in some animal and human VCA associated with DSA • T- and B-cell markers to assess infiltrates and vascular lesions

DIFFERENTIAL DIAGNOSIS Donor Vascular Disease • Arteriosclerosis typically has fibroelastosis and few or no T cells in intima (judging from solid organ transplants)

• Donor-specific antibodies (DSA)

Ischemia Due to Technical Issues

Treatment • Drugs ○ Limited experience ○ Restore or enhance immunosuppression if possible

• Anastomotic narrowing from surgical complication, trauma, or other processes • Imaging studies may demonstrate localized, circumscribed narrowing of vessels as opposed to CAV, which is typically uniform along length of vessel

Prognosis

Infection

• Limited data, but late graft loss reported and may be inevitable with current therapy

• Special stains and cultures may be needed to rule out infections since inflammatory changes of chronic rejection can be mimicked by infection

IMAGING Angiography • High-resolution peripheral CT for vascular stenosis and occlusion

High-Resolution Ultrasonography • Color Doppler to assess blood flow

Vascularized Composite Allotransplantation

TERMINOLOGY

DIAGNOSTIC CHECKLIST Clinically Relevant Pathologic Features • Can resemble scleroderma • Arteries are important target • Not yet incorporated into Banff VCA classification

Pathologic Interpretation Pearls

MICROSCOPIC Histologic Features • Epidermis ○ Atrophy (loss of rete pegs) • Adnexa ○ Atrophy or fibrosis of hair follicles, sweat, and sebaceous glands ○ Nail loss

• Punch biopsies may not be deep enough to demonstrate chronic vascular changes, muscular atrophy, and other soft tissue changes

SELECTED REFERENCES 1.

Kanitakis J et al: Chronic rejection in human vascularized composite allotransplantation (hand and face recipients): an update. Transplantation. 100(10):2053-61, 2016

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SECTION 11

Posttransplant Neoplastic Disorders

Posttransplant Lymphoproliferative Disease

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Posttransplant Neoplastic Disorders

Posttransplant Lymphoproliferative Disease KEY FACTS

TERMINOLOGY

MICROSCOPIC

• Posttransplant lymphoproliferative disorder (PTLD) • Frank lymphoma sometimes manifests in immunocompromised hosts in solid organ or bone marrow allograft recipients

• Mononuclear cells ("activated" lymphocytes) with enlarged nuclei, prominent nucleoli, & mitoses • Cells of PTLD may be monomorphic or polymorphic • Necrosis, often described as being in serpiginous pattern, is often present

CLINICAL ISSUES • Occurs in 1-5% of solid organ allograft recipients • Treatment ○ Reduction in immunosuppression, antiviral drugs (acyclovir, ganciclovir, α-interferon), chemotherapy, antiCD20 (rituximab) • Reported mortality of 40-60%

ANCILLARY TESTS

MACROSCOPIC

TOP DIFFERENTIAL DIAGNOSES

• Swollen kidneys • Blurring of corticomedullary junction & diffuse petechiae with vaguely nodular involvement

• Allograft rejection ○ CD3(+) T cells, granulocytes, & macrophages are more commonly seen in rejection

• Most express CD20 (~ 85-90%) • Immunohistochemical stains for EBV-associated antigens (such as LMP-1 & EBNA-2) • In situ hybridization for EBV-encoded RNA usually shows prominent staining in atypical lymphoid cells

Mitotically Active B-Cell PTLD

EBV ISH in EBV-Driven B-Cell PTLD

PTLD, Diffuse Large B-Cell Lymphoma Type

PTLD, Hodgkin Lymphoma Type

(Left) High-power view of a monomorphic EBV-driven Bcell posttransplant lymphoproliferative disease (PTLD) shows pleomorphic cells, many of which are enlarged & have irregular nuclear contours, prominent nucleoli, & mitoses ﬈. (Right) In situ hybridization (ISH) of Epstein-Barr virus-encoded RNA (EBER) shows that many cells in the lymphoid infiltrate are positive ﬉, indicating that this is a monomorphic EBVdriven B-cell PTLD, diffuse large B-cell lymphoma type.

(Left) At medium power, one can appreciate an arterial wall involved by monomorphic EBV-driven B-cell PTLD, diffuse large B-cell lymphoma type, with adjacent necrosis st & malignant lymphoid cells & admixed fibrin ﬇. (Right) This Hodgkin lymphoma-type PTLD contains many ReedSternberg variant cells ſt. (Courtesy N.L. Harris, MD.)

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Posttransplant Lymphoproliferative Disease

Abbreviations • Posttransplant lymphoproliferative disease (PTLD)

Synonyms • Posttransplant lymphoproliferative disorder

Definitions

Presentation

• Lymphoproliferative process can manifest as frank lymphoma arising in immunocompromised recipients of solid organ or bone marrow allografts

• • • •

ETIOLOGY/PATHOGENESIS Immunosuppression • Risk for PTLD increases with increasing immunosuppression ○ < 1% peripheral blood, stem cell, & bone marrow allograft recipients ○ 1% renal allograft recipients ○ 1-2% cardiac allograft recipients ○ ≥ 5% heart-lung or intestinal allograft recipients

Epstein-Barr Virus • Most important risk factor is Ebstein-Barr Virus (EBV) seronegativity at time of transplantation ○ 70% of PTLD are EBV(+) ○ Primary EBV infection increases risk for PTLD by 10-76x

B-Cell PTLD • Most PTLD are B-cell type • Usually driven by EBV ○ EBV(-) cases often have TP53 mutations • Monomorphic B-cell PTLD are monoclonal transformed B lymphocytic or plasmacytic proliferations fulfilling diffuse large B-cell lymphoma criteria (& less commonly Burkitt lymphoma or plasma cell neoplasms) • Can also be polyclonal

T-Cell & NK-Cell PTLD • T-/NK-cell PTLD account for 7-15% of PTLDs (larger reported range in 1 Japanese series of 2-45%) • Occur longer after transplant (median of 66 months) & are usually extranodal • ~ 1/3 are EBV(+) • Median survival is 6 months • EBV(+) cases survive longer • Types ○ Peripheral T-cell lymphoma, unspecified ○ Hepatosplenic T-cell lymphoma ○ T-cell large granular lymphocytic leukemia [EBV(-)]

CLINICAL ISSUES Epidemiology • Incidence ○ ~ 1% of renal allograft recipients (1.4% of 25,127 recipients from 1996-2000) – Allograft kidney affected in > 30% of PTLD patients ○ Kidney more often involved in kidney transplant (14%) than heart transplant patients (0.7%) – Conversely, heart (18%) is involved more than kidney (7%) transplant patients

•

• • •

Malaise Weight loss Lethargy Fever ○ Fever of unknown origin/unexplained fever Mononucleosis-type syndrome ○ Fever & malaise ○ Pharyngitis or tonsillitis – Sometimes recognized incidentally on tonsillectomy specimens ± lymphadenopathy Abdominal mass Hepatocellular or pancreatic dysfunction Central nervous system disease

Posttransplant Neoplastic Disorders

– Suggests immunologic reaction in allograft is pathogenic factor ○ 67% of PTLDs involving kidney allograft are donor origin – Donor origin PTLD more common in liver & lung allograft recipients & frequently involves allograft ○ PTLD account for 15% of tumors among adult transplant recipients (~ 51% in children)

TERMINOLOGY

Laboratory Tests • Quantitative EBV viral load testing with polymerase chain reaction ○ Serial assays more useful in individual patient than specific viral load measurements ○ Assays not standardized & cannot be compared between centers • Serological testing is not useful

Natural History • PTLD restricted to kidney transplant (~ 12% of cases) tend to occur early (~ 5 months) after surgery • EBV(-) PTLD & T-/NK-cell PTLD tend to present later (median time to occurrence 4-5 years & 6.5 years, respectively)

Treatment • Drugs ○ Reduction in immunosuppression ○ Antiviral drugs (acyclovir, ganciclovir, α-interferon) ○ Chemotherapy – Often: CHOP [cyclophosphamide, hydroxydaunomycin, vincristine (Oncovin), prednisone] ○ Anti-CD20 (rituximab) – Contributes to complete remission • Radiation ○ Localized radiation may be combined with chemotherapy • Graft nephrectomy ○ Permits discontinuation of immunosuppression • Cell immunotherapy (investigative) ○ EBV-specific cytotoxic T cells

Prognosis • Overall reported 5-year survival: 40-70% ○ 87% 5-year survival in children ○ Recent series shows improvement in outcome 527

Posttransplant Neoplastic Disorders

Posttransplant Lymphoproliferative Disease • Nondestructive PTLD tends to regress when ↓ immunosuppression • Polymorphic & less often monomorphic PTLD may also regress with reduction in immune suppression • Acute & chronic rejection may occur when immunosuppression reduced, leading to graft loss & mortality • Associated with adverse outcome ○ Multiple disease sites (not in pediatric patients) ○ Advanced stage ○ Older age at diagnosis ○ Late-onset disease ○ Higher international prognostic index ○ Elevated lactate dehydrogenase ○ Bone marrow vs. solid organ allograft recipients

•

IMAGING General Features • May show as mass on radiology studies

•

MACROSCOPIC General Features • • • •

Swollen kidneys Blurring of corticomedullary junction & diffuse petechiae Vaguely nodular involvement Localized mass

MICROSCOPIC Histologic Features • WHO classification (2017) ○ 4 major categories – Nondestructive PTLD – Polymorphic PTLD – Monomorphic PTLD (T, B, NK) – Classic Hodgkin lymphoma – First 2 are specific for transplant recipients • Nondestructive PTLD ○ Plasmacytic hyperplasia & infectious mononucleosis-like PTLD ○ Involved tissue has architectural preservation ○ Nodal sinuses or tonsillar crypts still present ○ Follicles often floridly reactive or hyperplastic ○ In plasmacytic hyperplasia, plasma cells are prominently admixed with small lymphocytes ○ In infectious mononucleosis-like lesion, paracortical expansion with numerous immunoblasts admixed with T cells & plasma cells ○ Lesions may form masses ○ Occur at younger age than other PTLD types (children or adult solid organ recipients who have not had prior EBV infections) ○ Lymph nodes or tonsils are common sites ○ Often EBV(+) • Polymorphic PTLD ○ Immunoblasts, plasma cells, & small- to intermediatesized lymphoid cells effacing architecture of lymph nodes ○ Full range of B-cell maturation is present 528

•

•

○ Large, bizarre cells may resemble Reed-Sternberg cells [atypical immunoblasts (may be Hodgkin-like)] ○ Areas of geographic necrosis may be present ○ Distinction of polymorphic from monomorphic PTLD is not always clear cut ○ Frequency: 20-80%, depending on institution ○ Polymorphic variant is most common type in children & frequently follows primary EBV infection ○ Polymorphic variant is more common in PTLD involving kidney ○ Often EBV(+) Monomorphic PTLD ○ Fulfills criteria for 1 of B-cell or T-/NK-cell neoplasms recognized in immunocompetent hosts ○ Small B-cell lymphoid neoplasms, such as follicular lymphomas or MALT lymphomas, are not designated as PTLD – Even though some occur in posttransplant setting (e.g., MALT lymphoma) Monomorphic B-cell PTLD ○ Necrosis, often described as serpiginous ○ Monomorphic B-cell PTLD often fulfill criteria for diffuse large B-cell lymphoma ○ Burkitt lymphoma or plasma cell neoplasms occur less commonly ○ Cells often collected in vaguely nodular pattern in sheets ○ Mononuclear cells ("activated" lymphocytes) with enlarged nuclei, prominent nucleoli, & frequent mitoses ○ Cells may have blast-like features ○ Term "monomorphic" does not imply cellular monotony since cells may be bizarre, multinucleated, & ReedSternberg-like ○ May also be plasmacytic or plasmacytoid features ○ Burkitt lymphomas have monomorphic medium-sized transformed cells, often with multiple small nucleoli & dispersed chromatin, & may possess MYC gene translocations – e.g., characteristically t(8;14) but also t(8;22) or t(2;8) Monomorphic T-/NK-cell PTLD ○ Fulfill criteria for T-/NK-cell lymphomas ○ Most present at extranodal sites ○ Largest group consists of peripheral T-cell lymphoma, not otherwise specified (NOS) category ○ Peripheral T-cell lymphoma, NOS has wide range in morphology – Often accompanied by eosinophilia, pruritus, or hemophagocytic syndrome ○ Hepatosplenic T-cell lymphomas (up to 20%) arise in setting of chronic immunosuppression – Mostly long-term immunosuppression for solid organ transplantation in which it is regarded as late-onset PTLD of host origin – Thought to arise from cytotoxic T cells (usually of γ/δ variety) – Demonstrates medium-sized lymphoid cells infiltrating bone marrow, spleen, & liver Classic Hodgkin lymphoma-type PTLD ○ Least common form of PTLD ○ Occurs more commonly in renal transplant than other transplant recipients

Posttransplant Lymphoproliferative Disease

ANCILLARY TESTS Immunohistochemistry

• Cases of T-cell origin have clonal T-cell receptor gene rearrangements

Genetic Testing • Clonal cytogenetic abnormalities common, particularly in monomorphic PTLD

Array CGH • Demonstrate additions, gains, & losses

DIFFERENTIAL DIAGNOSIS

• Most are B-cell derived & express B-cell markers ○ CD20 (~ 85-90%) ○ CD30(+) in many B-cell PTLD cases (± anaplastic morphology) ○ CD138(+) in minority ○ EBV(+) cases usually have late germinal center/postgerminal center phenotype [CD10(-), Bcl6(+/-), IRF/MUM1(+)] ○ EBV(-) cases often have germinal center phenotype [CD10(+/-), Bcl-6(+), IRF/MUM1(-), CD138(-)] ○ EBV(-) monomorphic PTLD frequently lacks expression of cyclin-dependent kinase inhibitor [CDKN2A (p16INK4A)] ○ Monotypic immunoglobulin often with expression of γor α-heavy chain in ~ 50% of monomorphic B-cell PTLD ○ EBV replication in tumor [BZLF1(+) or BMRF1(+)] & plasma cell differentiation [XBP1(+)] predictive of poor prognosis (18% vs. 48% 1-year survival) • Classic Hodgkin lymphoma-type PTLD ○ Typically CD15(+), CD30(+), & EBV(+) ○ CD15(-) classic Hodgkin lymphoma case occur – Should be distinguished from other Hodgkin-like lesions ○ CD15, when present, often gives Golgi-type pattern of expression • T-/NK-cell PTLD have pan-T-cell & NK-cell antigens ○ CD4 or 8, CD30, ALK, & α/β or γ/δ T cell – Hepatosplenic T-cell lymphoma usually of γ/δ variety ○ ~ 1/3 EBV(+) • Immunohistochemical stains for EBV-associated antigens (such as LMP-1 & EBNA-2)

Allograft Rejection

In Situ Hybridization

Key Elements to Report

• Most EBV(+) (~ 85%) ○ In situ hybridization for EBV-encoded RNA usually shows prominent staining in atypical lymphoid cells • In situ hybridization for κ- & λ-light chains may demonstrate light chain restriction ○ ~ 50% of monomorphic PTLD & usually only focal in polymorphic PTLD

• Categories of PTLD according to 2017 WHO classification ○ Nondestructive PTLD (formerly "early lesions") – Plasmacytic hyperplasia – Infectious mononucleosis – Florid germinal center hyperplasia ○ Polymorphic PTLD ○ Monomorphic PTLD – B-cell neoplasms: Diffuse large B-cell lymphoma, Burkitt lymphoma, plasma cell myeloma, plasmacytoma-like lesion, other (e.g., indolent small Bcell lymphomas arising in transplant recipients) – T-cell neoplasms: Peripheral T-cell lymphoma, NOS; hepatosplenic T-cell lymphoma; other ○ Classic Hodgkin lymphoma-type PTLD

PCR • Gene rearrangement studies can demonstrate clonally rearranged immunoglobulin genes ○ More prominent in monomorphic B-cell PTLD ○ Can occur in polymorphic B-cell PTLD – Some report immunoglobulin gene variable regions without ongoing mutations in 75% of polymorphic PTLDs • Monomorphic B-cell PTLDs have oncogene abnormalities, such as RAS, TP53, & MYC rearrangements, BCL6 somatic hypermutation, & aberrant promoter methylation

• PTLD may be confused with allograft rejection because features mimicking rejection, such as tubulitis & endarteritis • Mixed infiltrate typical of rejection, including granulocytes & macrophages, rather than monotonous sheets of mononuclear cells of PTLD • Rejection predominantly of CD3(+) T-lymphocytes & CD68(+) macrophages, vs. CD20(+) B cells in PTLD • Concurrent rejection & PTLD may occur • Necrosis of infiltrate rare in rejection • Edema not typical of PTLD

Posttransplant Neoplastic Disorders

○ Reed-Sternberg-like cells may be seen in early, polymorphic, & some monomorphic PTLDs & cause diagnostic confusion ○ Cases should fulfill criteria for classic Hodgkin lymphoma ○ Usually both CD15 & CD30 are (+) ○ CD15(-) cases can occur but must be distinguished from Hodgkin-like lesions

Smooth Muscle Tumors • Spindle cell neoplasms may be seen in posttransplant setting & may be EBV(+) • Histology usually confirms prominent spindling ○ Not typical of PTLD • Immunohistochemical studies usually demonstrate smooth muscle differentiation (actin & desmin)

DIAGNOSTIC CHECKLIST Pathologic Interpretation Pearls • Detection of rare EBV(+) cells without lymphoid/plasmacytic proliferation is not diagnostic of PTLD • PTLDs do not include indolent small B-cell lymphomas arising in transplant setting, except for EBV(+) marginal zone lymphomas

REPORTING

SELECTED REFERENCES 1.

Swerdlow SH et al: Post-transplant lymphoproliferative disorders. In Swerdlow SH et al: WHO Classification. Tumours of Haematopoietic and Lymphoid Tissues. Revised 4th ed. Lyon: IARC, 2017

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Posttransplant Neoplastic Disorders

Posttransplant Lymphoproliferative Disease Classification & Features of Posttransplant Lymphoproliferative Disease Category

Morphology

EBV

Immunophenotype and Genetics

Plasmacytic hyperplasia

Plasma cells & lymphocytes

(+)

Usually polyclonal

Infectious mononucleosis

Mixture: Lymphocytes ± plasma cells, ± immunoblasts

(+)

Usually polyclonal; may have small oligoclonal TCR

Florid follicular hyperplasia Prominent hyperplastic germinal centers

Variable

Usually polyclonal; may have small oligoclonal B cells

Polymorphic PTLD

Most (+)

Monoclonal B cells; polyclonal T cells

Nondestructive PTLD

Mixture: Lymphocytes ± plasma cells ± immunoblasts (full spectrum of maturation)

Monomorphic PTLD B-cell neoplasms

Express B-cell markers

Diffuse large B-cell lymphoma

Large B cells

Variable

Clonal

Burkitt lymphoma

Medium-sized B cells with multiple nucleoli & finely dispersed chromatin

Variable

Clonal, may have translocations, e.g., t(8;14) but also t(8;22) or t(2;8)

Plasma cell myeloma

Diffuse plasma cell infiltrate with criteria sufficient to diagnose myeloma

Variable

Clonal

Plasmacytoma-like lesion

Localized site with diffuse plasma cell infiltrate

Usually (-)

Monotypic immunoglobulin

Peripheral T-cell lymphoma, NOS

May be polymorphous or polymorphous with medium- to large-sized cells & RS-type cells

Minority (+) Typically clonal with rearranged T-cell receptor genes

Hepatosplenic T-cell lymphoma

Liver, spleen, & bone marrow are diffusely involved with monotonous medium-sized cells with inconspicuous nucleoli

Minority (+) Typically clonal with rearranged T-cell receptor genes (usually of γ/δ type), may have +8 chromosomal modification

Classic Hodgkin lymphoma-type PTLD

Fulfills criteria for diagnosis of classic Hodgkin lymphoma & classically may have RS cells

Often (+)

T-cell neoplasms

Express T-cell (& less commonly NK-cell) markers

Clonality usually cannot be shown, e.g., with IgH rearrangement studies

Not otherwise specified = NOS; Posttransplant lymphoproliferative disease = PTLD; Reed-Sternberg = RS. Swerdlow SH et al: Post-transplant lymphoproliferative disorders. In Swerdlow SH et al: WHO Classification. Tumours of Haematopoietic and Lymphoid Tissues. Revised 4th ed. Lyon: IARC, 2017. 2.

Courville EL et al: EBV-negative monomorphic B-cell post-transplant lymphoproliferative disorders are pathologically distinct from EBV-positive cases and frequently contain TP53 mutations. Mod Pathol. 29(10):1200-11, 2016 3. Durrbach A et al: Long-term outcomes in belatacept- versus cyclosporinetreated recipients of extended criteria donor kidneys: final results from BENEFIT-EXT, a phase III randomized study. Am J Transplant. 16(11):31923201, 2016 4. Maksten EF et al: Post-transplant lymphoproliferative disorder following kidney transplantation: a population-based cohort study. Transpl Int. 29(4):483-93, 2016 5. Rosenberg AS et al: Hodgkin lymphoma post-transplant lymphoproliferative disorder: a comparative analysis of clinical characteristics, prognosis, and survival. Am J Hematol. 91(6):560-5, 2016 6. Haynes SE et al: Post-transplant lymphoproliferative disease and other malignancies after pediatric cardiac transplantation: an evolving landscape. Curr Opin Organ Transplant. 20(5):562-9, 2015 7. Morton M et al: Post-transplant lymphoproliferative disorder in adult renal transplant recipients: survival and prognosis. Leuk Lymphoma. 1-23, 2015 8. Singavi AK et al: Post-transplant lymphoproliferative disorders. Cancer Treat Res. 165:305-27, 2015 9. Gonzalez-Farre B et al: In vivo intratumoral Epstein-Barr virus replication is associated with XBP1 activation and early-onset post-transplant lymphoproliferative disorders with prognostic implications. Mod Pathol. 27(12):1599-611, 2014 10. Al-Mansour Z et al: Post-transplant lymphoproliferative disease (PTLD): risk factors, diagnosis, and current treatment strategies. Curr Hematol Malig Rep. 8(3):173-83, 2013 11. Bagg A et al: Immunosuppressive and immunomodulatory therapyassociated lymphoproliferative disorders. Semin Diagn Pathol. 30(2):102-12, 2013 12. Wistinghausen B et al: Post-transplant lymphoproliferative disease in pediatric solid organ transplant recipients. Pediatr Hematol Oncol. 30(6):520-31, 2013

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13. Bollard CM et al: T-cell therapy in the treatment of post-transplant lymphoproliferative disease. Nat Rev Clin Oncol. 9(9):510-9, 2012 14. Trappe R et al: Sequential treatment with rituximab followed by CHOP chemotherapy in adult B-cell post-transplant lymphoproliferative disorder (PTLD): the prospective international multicentre phase 2 PTLD-1 trial. Lancet Oncol. 13(2):196-206, 2012 15. Dharnidharka VR et al: Improved survival with recent post-transplant lymphoproliferative disorder (PTLD) in children with kidney transplants. Am J Transplant. 11(4):751-8, 2011 16. Ibrahim HA et al: Presence of monoclonal T-cell populations in B-cell posttransplant lymphoproliferative disorders. Mod Pathol. 24(2):232-40, 2011 17. Olagne J et al: Post-transplant lymphoproliferative disorders: determination of donor/recipient origin in a large cohort of kidney recipients. Am J Transplant. 11(6):1260-9, 2011 18. Picarsic J et al: Post-transplant Burkitt lymphoma is a more aggressive and distinct form of post-transplant lymphoproliferative disorder. Cancer. 117(19):4540-50, 2011 19. Khedmat H et al: Characteristics and prognosis of post-transplant lymphoproliferative disorders within renal allograft: Report from the PTLD.Int. Survey. Ann Transplant. 15(3):80-6, 2010 20. Parker A et al: Diagnosis of post-transplant lymphoproliferative disorder in solid organ transplant recipients - BCSH and BTS Guidelines. Br J Haematol. 149(5):675-92, 2010 21. Swerdlow SH: T-cell and NK-cell posttransplantation lymphoproliferative disorders. Am J Clin Pathol. 127(6):887-95, 2007

Posttransplant Lymphoproliferative Disease

PTLD, Immunoblastic Plasmacytoid Type (Left) Renal allograft protocol biopsy in a 3-year-old girl shows numerous plasma cells ﬈ amidst a hematolymphoid infiltrate, much of which was CD20(+), CD79a(+), & EBV ISH(+), compatible with the diagnosis of plasmacytic hyperplasia. (Right) Highpower view of the allograft from the autopsy of the recipient of a renal allograft shows involvement by PTLD with large atypical cells ſt among renal tubules. This PTLD was classified as an immunoblastic plasmacytoid PTLD. (Courtesy J.A. Ferry, MD.)

PTLD, Immunoblastic Plasmacytoid Type

Posttransplant Neoplastic Disorders

Plasmacytic Hyperplasia (Nondestructive PTLD)

PTLD, Infectious Mononucleosis Involving Tonsil (Left) High-power view of a renal allograft from the recipient's autopsy shows involvement by immunoblastic plasmacytoid PTLD with large atypical cells ﬈. (Courtesy J.A. Ferry, MD.) (Right) Lowpower view of a tonsil in a pediatric renal allograft recipient shows involvement by an infectious mononucleosis-like PTLD. The architecture is overall preserved, but it can be appreciated that lymphoid follicles show attenuated mantles. (Courtesy J.A. Ferry, MD.)

Mononucleosis PTLD

Immunoblasts in PTLD, Mononucleosis (Left) High-power view of the tonsil involved by mononucleosis-like PTLD shows that lymphoid follicles are floridly reactive with mostly small lymphocytes & scattered tingible body macrophages ﬈, immunoblasts ﬉, & apoptotic cells ﬊. (Courtesy J.A. Ferry, MD.) (Right) Higher power view of the interfollicular area of the tonsil with mononucleosis-like PTLD shows mostly small lymphocytes & scattered immunoblasts ﬈. (Courtesy J.A. Ferry, MD.)

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Posttransplant Neoplastic Disorders

Posttransplant Lymphoproliferative Disease

Nodular PTLD

PTLD With Necrosis

PTLD, Diffuse Large B-Cell Lymphoma Type With Necrosis

Infiltration of Fat by PTLD

Tubulitis in PTLD

EBV-Driven PTLD

(Left) A gross photograph shows nodular involvement of an allograft kidney by PTLD. (Courtesy P. Randhawa, MD.) (Right) A low-power view of the allograft from the autopsy of the recipient of a renal allograft shows involvement by PTLD with an area of necrosis ſt. (Courtesy J.A. Ferry, MD.)

(Left) PAS stain of a monomorphic EBV-driven Bcell PTLD, diffuse large B-cell lymphoma type, shows a relatively normal glomerulus ﬈ amidst a necrotic parenchyma ﬉ with a dense lymphoid infiltrate ﬊. (Right) This PTLD shows fatty infiltration in the surrounding soft tissues. (Courtesy P. Randhawa, MD.)

(Left) This B-cell PTLD has foci of tubulitis, illustrating that this pattern can be seen in PTLD as well as cellular rejection. (Courtesy P. Randhawa, MD.) (Right) EBER RNA stain shows tubulitis with EBV-infected cells, demonstrating that this PTLD is an EBV-driven process. (Courtesy P. Randhawa, MD.)

532

Posttransplant Lymphoproliferative Disease Vascular Involvement by PTLD Simulating Endarteritis (Left) High-power examination shows an arterial wall involved by monomorphic EBV-driven B-cell PTLD, diffuse large B-cell lymphoma type, with malignant lymphoid cells ſt & admixed fibrin ﬇. (Right) Hematoxylin & eosin stain shows vascular involvement by a B-cell PTLD, illustrating the fact that this can occur in PTLD, simulating endarteritis that can be seen in cellular rejection. (Courtesy P. Randhawa, MD.)

CD20(+) B-Cell PTLD

Posttransplant Neoplastic Disorders

PTLD, Diffuse Large B-Cell Lymphoma Type

Ki-67 Shows High Proliferation in PTLD (Left) CD20 immunohistochemistry of a monomorphic EBV-driven Bcell PTLD, diffuse large B-cell lymphoma type, shows that almost all of the atypical lymphoid infiltrate consists of CD20(+) B cells. (Right) A Ki-67 immunohistochemical stain is positive in most of the atypical lymphoid infiltrate in this monomorphic EBV-driven Bcell PTLD, diffuse large B-cell lymphoma type, indicating that it is highly proliferative.

Burkitt Lymphoma Type, Monomorphic

PTLD, Burkitt Lymphoma Type (Left) Low-power view of a submandibular gland in a patient with history of a renal allograft shows diffuse involvement by a lymphoid infiltrate in a Burkitt lymphoma-type monomorphic PTLD. (Courtesy J.A. Ferry, MD.) (Right) Giemsa stain of a Burkitt lymphoma-type PTLD shows that many of the cells have prominent nucleoli ſt. (Courtesy J.A. Ferry, MD.)

533

Posttransplant Neoplastic Disorders

Posttransplant Lymphoproliferative Disease

Pleomorphism in T-Cell PTLD

T-Cell PTLD

T-Cell PTLD

Hepatosplenic T-Cell Lymphoma

Hepatosplenic T-Cell Lymphoma

Hepatosplenic T-Cell Lymphoma

(Left) High-power view shows an atypical lymphoid infiltrate in a case of T-cell PTLD with a mixed pleomorphic cell population containing a mixture of small & large cells with a variety of shapes. (Right) High-power view of a T-cell PTLD shows large atypical cells ﬉, mitotic figures ﬈, & admixed eosinophils ſt.

(Left) Immunohistochemical stain of T-cell PTLD shows that most of the atypical infiltrate is composed of CD3(+) T cells with a variety of shapes & sizes. (Right) A liver biopsy in a renal allograft recipient with hepatosplenic Tcell lymphoma has diffuse portal & lobular involvement by a lymphoid infiltrate of small to medium lymphoid cells mimicking hepatitis. (Courtesy J.A. Ferry, MD.)

(Left) A portal tract has a number of small- to mediumsized lymphoid cells with only mild cytologic atypia that infiltrate into the lobules in a hepatosplenic T-cell lymphoma in a renal allograft recipient. (Courtesy J.A. Ferry, MD.) (Right) It can be appreciated in this image that the lymphoid cells primarily travel along the hepatic sinusoids ﬈ in the hepatic lobules in a hepatosplenic Tcell lymphoma in a renal allograft recipient. (Courtesy J.A. Ferry, MD.)

534

Posttransplant Lymphoproliferative Disease

Reed-Sternberg Variant Cells (Left) At low power, a Hodgkin lymphoma-type PTLD involving the tongue base can be appreciated with a heterogeneous cell population. (Courtesy N.L. Harris, MD.) (Right) Throughout this Hodgkin lymphoma-type PTLD, there are many Reed-Sternberg variant cells ſt. (Courtesy N.L. Harris, MD.)

Reed-Sternberg Variant Cells in Hodgkin Lymphoma Type

Posttransplant Neoplastic Disorders

PTLD, Hodgkin Lymphoma Type

Neural Involvement by PTLD (Left) This Hodgkin lymphomatype PTLD contains many Reed-Sternberg variant cells ſt. (Courtesy N.L. Harris, MD.) (Right) At medium-power examination, one can observe a nerve with involvement by Bcell PTLD. (Courtesy P. Randhawa, MD.)

Posttransplant Spindle Cell Tumor

EBER ISH in Posttransplant Spindle Cell Tumor (Left) Light microscopy shows a posttransplant spindle cell tumor. In this case, immunohistochemistry showed smooth muscle differentiation. (Courtesy P. Randhawa, MD.) (Right) In situ hybridization for EBER shows EBV-infected cells in a spindle cell tumor. Lymphoid markers would demonstrate that this is not a PTLD but rather a posttransplant spindle cell tumor caused by EBV. (Courtesy P. Randhawa, MD.)

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INDEX

A

ABH antigens, 40 ABO antibodies, 40 ABO blood group antigens, 40–41 - ABH antigens, 40 - ABO antibodies, 40 - ABO blood groups, 41 - apheresis for desensitizing ABO-incompatible transplantation, 42 - barrier to transplantation, 41 - current status, 41 - expression of ABH antigen, 40 - humoral immune response, 40 - rationale for crossing ABO barrier, 41 - transplantation and, 40–41 - transplantation of A2/A2B donor kidneys into blood group B candidates, 41 Abscesses - aspergillosis, 217 - candidiasis, 211 - infectious, pancreas transplantation associated, 484 - kidney, lymphocele vs., 107 Acanthosis, acute T-cell- and antibody-mediated rejection associated, 516 Accommodation, 94–95 - diagnostic checklist, 95 - differential diagnosis, 95 - prognosis, 95 - protocol biopsies, 91 Acid-fast stains, nocardiosis, 231 Acinar and interacinar inflammation, antibody-mediated rejection associated, 493 Acinar cell injury, antibody-mediated rejection associated, 493 Acini, chronic allograft rejection/graft sclerosis associated, 497 Acinitis, acute cellular rejection associated, 489 ACR. See Acute cellular rejection. Active AMR. See Antibody-mediated rejection, acute. Acute/active cellular rejection, recurrent diabetes mellitus vs., 500 Acute allergic tubulointerstitial nephritis, acute T-cellmediated rejection vs., 118 Acute allograft glomerulopathy, cytomegalovirus (CMV) infection vs., 206 Acute allograft ischemia, 100–103 - clinical issues, 101 - diagnostic checklist, 101 - differential diagnosis, 101

- prognosis, 101 Acute allograft rejection (AR) - early allograft loss, 80 - hepatic artery thrombosis vs., 251 - late allograft loss, 80 - portal vein thrombosis vs., 253 Acute AMR. See Antibody-mediated rejection, acute. Acute AMR Banff criteria, 268 Acute antibody-mediated rejection (AMR), 132–139, 234 - acute pyelonephritis vs., 190 - bile duct stricture vs., 255 - bile leak vs., 255 - bile sludge vs., 255 - biloma vs., 255 - clinical issues, 133 - diagnostic checklist, 134 - differential diagnosis, 134 - intestinal allograft diseases, 412 - intestine, 424–427 diagnostic checklist, 426 differential diagnosis, 426 prognosis, 425 - kidney, acute allograft ischemia vs., 101 - mTOR inhibitor toxicity vs., 187 - pancreas transplantation associated, 484 - prognosis, 133 - vascularized composite allotransplantation, 514–521 diagnostic checklist, 517 differential diagnosis, 516–517 gene expression, 516 prognosis, 515 Acute bacterial nephritis. See Acute pyelonephritis. Acute cell-mediated rejection. See T-cell-mediated rejection. Acute cellular rejection (ACR), 281, 308, 362. See also Recurrent hepatitis C virus; T-cell-mediated rejection. - acute antibody-mediated rejection, intestine vs., 426 - adenovirus vs., 458 - alloimmune response and, 362 - antibody-mediated rejection, lung vs., 389 - bacterial and fungal infections vs., 454 - cytomegalovirus vs., 462 - Epstein-Barr virus vs., 466 - grade A, 390–391 diagnostic checklist, 391 differential diagnosis, 391 grade B vs., 393 prognosis, 391 - grade B, 392–393 bacterial infections vs., 401 diagnostic checklist, 393 differential diagnosis, 393 prognosis, 393

i

INDEX - graft-vs.-host disease vs., 450 - heart, 342–345 diagnostic checklist, 343 differential diagnosis, 343 prognosis, 343 - herpes simplex virus vs., 462 - intestinal allograft diseases, 412 - intestine, 428–435 diagnostic checklist, 430–431 differential diagnosis, 430, 431 histologic grading, 431 prognosis, 429 - kidney, acute allograft ischemia vs., 101 - microaspiration vs., 399 - pancreas, 488–491 allograft diseases, 472 diagnostic checklist, 490 differential diagnosis, 490 prognosis, 489 - reperfusion injury vs., 419 - rotavirus vs., 462 - severe, chronic rejection, intestine vs., 437 - site of previous biopsy vs., 353 Acute erosive gastritis, stomach rejection vs., 441 Acute fatty liver of pregnancy, liver removal, 237 Acute glomerulonephritis, cytomegalovirus (CMV) infection vs., 206 Acute graft failure, alloimmune diseases and, 362 Acute humoral rejection. See Acute antibody-mediated rejection. Acute infectious pancreatitis, 506 Acute infectious peripancreatitis, 472 Acute interstitial nephritis - drug-associated, 61 - polyomavirus nephritis vs., 194 Acute ischemic injury, 101 Acute ischemic pancreatitis, 472 Acute liver failure - liver transplantation, 236 - native liver removal, 237 Acute lobar nephronia, 190 Acute mTOR inhibitor toxicity, 187 Acute obstruction, acute allograft ischemia vs., 101 Acute purulent peripancreatitis, 506 Acute pyelonephritis, 188–191 - diagnostic checklist, 190 - differential diagnosis, 190 - immunohistochemistry, 190 - prognosis, 189 Acute rejection. See Heart, acute cellular rejection; T-cellmediated rejection. Acute rheumatic fever, myocarditis vs., 357 Acute T-cell-mediated rejection, 116–127 - acute antibody-mediated rejection vs., 134 - acute pyelonephritis vs., 190 - adenovirus, kidney vs., 201 - Banff criteria, 119 - cytomegalovirus (CMV) infection vs., 206 - diagnostic checklist, 118 - differential diagnosis, 118 - engraftment syndrome vs., 171 ii

-

pancreas, antibody-mediated rejection vs., 494 pancreas transplantation associated, 484 renal artery or vein thrombosis vs., 109 vascularized composite allotransplantation, 514–521 Banff grading system, 515 criteria, 515 diagnostic checklist, 517 differential diagnosis, 516–517 gene expression, 516 prognosis, 515 Acute transient arteriopathy, acute T-cell-mediated rejection vs., 118 Acute transplant glomerulitis, recurrent disease in the allograft vs., 155 Acute tubular injury - adenovirus, kidney, 201 - polyomavirus nephritis vs., 194 Acute tubular necrosis, polyomavirus nephritis vs., 194 Acute tubulointerstitial nephritis, nocardiosis vs., 231 Acute tubulointerstitial (type I) rejection, polyomavirus nephritis vs., 194 Acute vascular rejection. See Acute antibody-mediated rejection. Acute viral myocarditis, myocarditis and, 356 Acyanotic heart disease, 327 Adalimumab, history of use, 55 Adams, William E., 366 Adenovirus, 235, 292–293 - acute cellular rejection, intestine vs., 430 - bacterial and fungal infections vs., 454 - diagnostic checklist, 293 - differential diagnosis, 293 - Epstein-Barr virus vs., 466 - herpes simplex virus vs., 291 - infections, 403 rotavirus vs., 462 - intestine, 456–459 diagnostic checklist, 458 differential diagnosis, 458 prognosis, 457 serotypes, 457 - kidney, 200–203 diagnostic checklist, 201 differential diagnosis, 201 prognosis, 201 - liver, 292–293 diagnostic checklist, 293 differential diagnosis, 293 prognosis, 293 - pancreas transplantation associated, 505 - prognosis, 293 Adenovirus (AdV) nephritis. See Adenovirus, kidney. Adenovirus tubulointerstitial nephritis, polyomavirus nephritis vs., 194 Adequacy, donor organ, 242 Adhesion molecules, acute T-cell-mediated rejection associated, 515 Adhesions, arterial, chronic rejection, intestine vs., 437 Adult indications, common, lung transplantation and, 363 AdV. See Adenovirus. AIH. See Autoimmune hepatitis.

INDEX Airway inflammation. See Acute cellular rejection, grade B. Akar, Munire Erman, 513 Alanine transaminase, 293 Alcoholic hepatitis, hepatic venous outflow obstruction vs., 258 Alcoholic liver disease, 287 Alcoholic steatohepatitis, 287 ALD. See Alcoholic liver disease. Alemtuzumab (anti-CD52), history of use, 55 Allergic contact dermatitis, acute T-cell- and antibodymediated rejection vs., 517 Alloantibody testing, 59 Allograft, vascularized composite, 512 Allograft gene expression profiling, 35 Allograft intestine, adenovirus associated, 457 Allograft ischemia, acute, 100–103 - clinical issues, 101 - diagnostic checklist, 101 - differential diagnosis, 101 - prognosis, 101 Allograft kidney evaluation, 68–73 - Banff classification, 70–71 background, 70 categories, 70–71 caveats, 71 - Banff scoring categories, 71–72 arterial fibrointimal thickening, 71 arteriolar hyalinosis, 72 C4d score in PTC, 72 glomerulitis, 71 inflammation in areas of interstitial fibrosis and tubular atrophy, 72 interstitial fibrosis, 71 interstitial inflammation, 71 mesangial matrix increase, 72 peritubular capillary basement membrane multilayering, 72 peritubular capillary inflammation, 72 total inflammation, 72 transplant glomerulopathy, 71–72 tubular atrophy, 71 tubulitis, 71 vascular inflammation, 71 - biopsy evaluation of, 69 processing, 69 protocol biopsies, 70 special considerations in, 69–70 - clinical implications, 69 current drug therapy, 69 risk factors, 69 - etiology/pathogenesis, 68 afferent phase, 68 consequences, 68 effects determined by many variables, 68 efferent phase, 68 - new approaches, 70 expected added value, 70 molecular tests, 70 - terminology, 68

Allograft rejection - immune responses to allografts, and HLA, 36–37 graft-vs.-host disease, 37 prevention and treatment of, 37 HLA antigens, 36 prevention and treatment of rejection, 36–37 rejection, 36 types of rejection, 36 - NK cells, 16 - NK-cell-mediated responses in rejection and tolerance, 6 - pancreas transplantation associated, 475 - posttransplant lymphoproliferative disease vs., 529 Allograft thrombosis, early allograft loss, 80 Allograft tolerance, NK cells, 16 Alloimmune reaction, to donor antigens, chronic rejection associated, 523 Alloimmune response, 362 - pancreas allograft diseases, 472 Alloreactivity, infections and, 5 Allotransplantation, 476 - of pancreas. See Pancreas transplantation. Alport posttransplant nephritis. See Antiglomerular basement membrane (anti-GBM) disease in Alport syndrome. Alport syndrome - antiglomerular basement membrane (anti-GBM) disease, 166–167 clinical issues, 167 diagnostic checklist, 167 differential diagnosis, 167 prognosis, 167 - hyperperfusion injury vs., 169 ALT. See Alanine transaminase. Alveolar damage, diffuse, organizing, organizing pneumonia vs., 397 Amiodarone, other causes of end-stage lung disease related to, 381 AMR. See Antibody-mediated rejection. Amyloidosis - end-stage heart disease related to, 337 - hypertrophic cardiomyopathy vs., 324 Anastomotic leakage, pancreas transplantation associated, 505 Anastomotic stenosis, as complication of lung transplantation, 385 Anastomotic stricture, hyperperfusion syndrome vs., 261 Anastomotic vascular insufficiency, as complication of lung transplantation, 385 Antibodies, donor-specific, antibody-mediated rejection associated, 493 Antibody-mediated injury, chronic allograft rejection/graft sclerosis associated, 497 Antibody-mediated rejection (AMR), 60, 234, 308, 362 - acute cellular rejection grade A vs., 391 intestine vs., 430 - alloimmune response and, 362 - bacterial and fungal infections vs., 454

iii

INDEX - chronic active, acute antibody-mediated rejection vs., 134 - heart, 346–347 diagnostic checklist, 347 differential diagnosis, 347 prognosis, 347 - hepatic artery thrombosis vs., 251 - hyperacute, renal artery or vein thrombosis vs., 109 - liver, 266–269 differential diagnosis, 267 prognosis, 267 - lung, 388–389 differential diagnosis, 389 prognosis, 389 - lung allograft rejection, 386 - pancreas, 492–495 acute, 493 acute cellular rejection vs., 490 chronic active, 493 diagnostic checklist, 493 differential diagnosis, 494 hyperacute, 493 prognosis, 493 - pancreas allograft diseases, 472 - preservation injury vs., 249 - reperfusion injury vs., 419 - site of previous biopsy vs., 353 - T-cell-mediated rejection vs., 264 Antibody-mediated rejection, acute, 132–139 - acute allograft ischemia vs., 101 - acute T-cell-mediated rejection vs., 118 - clinical issues, 133 - diagnostic checklist, 134 - differential diagnosis, 134 - engraftment syndrome vs., 171 - intestine, 424–427 diagnostic checklist, 426 differential diagnosis, 426 prognosis, 425 - pancreas transplantation associated, 484 - prognosis, 133 - renal artery or vein thrombosis vs., 109 Antibody-mediated rejection, chronic, 140–151 - accommodation vs., 95 - Banff 2017 criteria, 144 - chronic T-cell-mediated rejection vs., 130 - clinical issues, 141 - diagnostic checklist, 143 - differential diagnosis, 142–143 - prognosis, 141 - protocol biopsies, 91 - recurrent disease in the allograft vs., 155 - scoring, 144, 145 Antigen-specific assessment, of T-cell immune competence, 26–27 - BK virus, 27 - cytomegalovirus, 26 - Epstein-Barr virus (EBV), 26–27 Antiglomerular basement membrane (anti-GBM) disease in Alport syndrome, 166–167 - clinical issues, 167 iv

- diagnostic checklist, 167 - differential diagnosis, 167 - prognosis, 167 Antiglucagon, acute cellular rejection associated, 490 Anti-HLA antibodies, 38 - bone marrow transplant, 38 - characteristics, 33 - methods to detect, 39 - solid organ transplant, 38 - testing, 34–35 Antiinsulin, acute cellular rejection associated, 490 Antiinsulin antibody, recurrent diabetes mellitus associated, 499 Anti-interleukin-2 receptor antibodies, mechanism of action, 49 Antimetabolites, mechanism of action, 49 Antithymocyte globulin - history of use, 54 - mechanism of action, 49 Antiviral tubular toxicity, 61 Antivirals, direct-acting, 275 Apheresis and transplantation, 42–43 - indications for, 42–43 desensitization, ABO-incompatible transplantation, 42 desensitization, donor-specific anti-HLA antibodies, 42–43 ECP and transplantation, 42 posttransplant treatment of rejection, 43 recurrent focal segmental glomerulosclerosis, 43 transplant-associated thrombotic microangiopathy, 43 - therapeutic apheresis, 42 Arrhythmogenic cardiomyopathy. See Arrhythmogenic right ventricular cardiomyopathy (ARVC). Arrhythmogenic right ventricular cardiomyopathy (ARVC), 332–335 - differential diagnosis, 334 - dilated cardiomyopathy vs., 320 - genetics, 333 Arrhythmogenic right ventricular dysplasia (ARVD). See Arrhythmogenic right ventricular cardiomyopathy (ARVC). Arterial adhesions, chronic rejection, intestine vs., 437 Arterial thrombosis - chronic rejection, intestine vs., 437 - mucormycosis, 215 Arteries, chronic allograft rejection/graft sclerosis associated, 497 Arteriopathy - acute transient, acute T-cell-mediated rejection vs., 118 - transplant, chronic antibody-mediated rejection vs., 142 Arteriosclerosis - chronic rejection vs., 523 - donor biopsy, 75 - hypertensive, chronic T-cell-mediated rejection vs., 130 - protocol biopsies, 91 Arteritis - acute T-cell- and antibody-mediated rejection associated, 516

INDEX - intimal, acute cellular rejection associated, 489 Arthroconidia, 219 Artifact - from formalin perfusion overinflation of lung, emphysema vs., 369 - microaspiration vs., 399 ARVC. See Arrhythmogenic right ventricular cardiomyopathy. Ascending infection, acute pyelonephritis, 189, 190 Ascending pyelonephritis, 189 Aschoff nodules, myocarditis vs., 357 ASH. See Alcoholic steatohepatitis. Aspartate transaminase, 293 Aspergillosis, 216–217, 235, 301 - candidiasis vs., 211 - diagnostic checklist, 217 - differential diagnosis, 217 - mucormycosis vs., 215 - prognosis, 217 Aspergillus, 217 - bacterial and fungal infections, 453 - fungal infections, lung, 407 Aspiration pneumonia, microaspiration vs., 399 AST. See Aspartate transaminase. Asymptomatic infection, acute pyelonephritis, 189 Atheromatous embolism, acute T-cell-mediated rejection vs., 118 Atherosclerotic coronary artery disease, chronic allograft vasculopathy, heart vs., 349 Athletes heart, hypertrophic cardiomyopathy vs., 324 Atypical hemolytic uremic syndrome - complement in, 20–21 - eculizumab and, 22 - genetic defects in, 21 - laboratory testing for diagnosis of, 22–23 - and transplant, 21–22 Autoantibodies, putative, recurrent diabetes mellitus, 499 Autoimmune hepatitis, 235 - de novo. See Plasma cell-rich rejection. - liver transplantation, 236 - recurrent, 280–281 diagnostic checklist, 281 differential diagnosis, 281 plasma cell-rich rejection vs., 303 prognosis, 281 T-cell-mediated rejection vs., 264 Autotransplantation, 476 Auxiliary liver transplantation, 246 Azathioprine (AZA) - history of use, 54 - mechanism of action, 49 - therapeutic monitoring, 52

B

Bacille Calmette-Guérin granulomatous interstitial nephritis, tuberculosis vs., 225 Bacterial infections, 400–401 - acute cellular rejection, intestine vs., 430 - adenovirus vs., 458 - cytomegalovirus vs., 462 - diagnostic checklist, 401 - differential diagnosis, 401 - graft-vs.-host disease vs., 305 - herpes simplex virus vs., 462 - intestinal allograft diseases, 412 - intestinal transplantation, 452–455 diagnostic checklist, 454 differential diagnosis, 454 infectious agents, 453 prognosis, 453 - intraabdominal and opportunistic infections vs., 507 - nonalloimmune diseases and, 362 - pancreas allograft diseases, 472 - prognosis, 401 - recurrent diabetes mellitus vs., 500 - reperfusion injury vs., 419 - rotavirus vs., 462 Bacterial pneumonia, fungal infections, lung vs., 407 Bacterial pyelonephritis, mucormycosis vs., 215 Banff criteria, for acute T-cell-mediated rejection, 119 Basiliximab - history of use, 55 - therapeutic monitoring, 52 B-cell immune response, in solid organ transplantation, 6 B-cell posttransplant lymphoproliferative disease, 527, 530 - monomorphic, 528 Belatacept (Nulojix) - history of use, 55 - mechanism of action, 49 Benfield, John R., 366 Berylliosis, chronic, sarcoidosis of lung vs., 377 Bilateral sequential lung transplantation, 362 Bile duct biloma, 234 Bile duct leak, 234 Bile duct necrosis, hepatic artery thrombosis, 251 Bile duct obstruction, fibrosing cholestatic hepatitis B or C virus vs., 279 Bile duct sludge, 234 Bile duct stricture, 234, 254–255 - differential diagnosis, 255 Bile duct syndrome, vanishing. See Graft-vs.-host disease. Bile leak, 254–255 - differential diagnosis, 255 Bile sludge, 254–255 - differential diagnosis, 255 Biliary/cholestatic changes, preservation injury, 249 Biliary cirrhosis, recurrent primary, 282–283 - diagnostic checklist, 283 - differential diagnosis, 283 v

INDEX - prognosis, 283 Biliary complications - hepatic artery thrombosis vs., 251 - portal vein thrombosis vs., 253 - posttransplant, hyperperfusion syndrome vs., 261 - T-cell-mediated rejection vs., 264 Biliary obstruction - antibody-mediated rejection vs., 267 - cytomegalovirus vs., 289 - graft-vs.-host disease vs., 305 - preservation injury vs., 249 - recurrent hepatitis C virus vs., 276 Biliary stricture, postsurgical, recurrent primary sclerosing cholangitis vs., 285 Biloma, 254–255 - differential diagnosis, 255 Biomarkers - immune response, 25–26 immune activation markers, 25 immune tolerance markers, 25 - of transplant tolerance, 9 Biopsy - evaluation, 311 - heart allograft postbiopsy perforation, previous biopsy site vs., 351 posttransplant, 309 previous site, allograft rejection related to. See Heart allograft rejection, site of previous biopsy. - kidney allograft evaluation of, 69 processing, 69 protocol biopsies, 70 special considerations in, 69–70 - pancreas transplant diagnostic utility, 475 history, 480–481 work-up, 476 Biopsy site - changes. See Site of previous biopsy (SOPB). - previous, acute cellular rejection, heart vs., 343 - prior, myocarditis vs., 356 BK polyoma virus nephropathy, late stage, chronic T-cellmediated rejection vs., 130 BK polyomavirus interstitial nephritis, acute T-cellmediated rejection vs., 118 BK virus, 193 - antigen-specific assessment, of T-cell immune competence, 27 BK virus nephropathy. See Polyomavirus nephritis. Blastomycosis - coccidioidomycosis vs., 219 - cryptococcosis vs., 213 - histoplasmosis vs., 209 - paracoccidioidomycosis vs., 221 Bleeding, postoperative, pancreas transplantation associated, 483 Bleomycin, other causes of end-stage lung disease related to, 381 Blind loops, biopsy, bacterial and fungal infections, 453 Blood vessel disease, pancreas allograft diseases, 473 vi

Bloodstream infection, pancreas transplantation associated, 505 BO. See Bronchiolitis obliterans. Bone marrow transplant, anti-HLA antibodies, 38 Bortezomib, mechanism of action, 49 BOS. See Bronchiolitis obliterans syndrome. Bowel perforation, Epstein-Barr virus, 465 Brazilian blastomycosis. See Paracoccidioidomycosis. Breidenbach, Warren, 512 Broelsch, Christoph, 246 Bronchial anastomosis stenosis, alloimmune diseases and, 362 Bronchial-associated lymphoid tissue, acute cellular rejection, grade B vs., 393 Bronchiectasis, cystic fibrosis vs., 371 Bronchiolitis obliterans (BO), 362 - acute cellular rejection, grade B vs., 393 - with OP. See Organizing pneumonia. Bronchiolitis obliterans syndrome (BOS), 362, 395 Bronchopneumonia, with foreign material, microaspiration vs., 399 Budd-Chiari syndrome, hepatic venous outflow obstruction and, 257 Burkitt lymphomas, monomorphic B-cell PTLD, 528 Burnet, Frank Macfarlane, 512

C

C4d, 60 CAD. See Coronary artery disease. Calcineurin inhibitor toxicity, 61, 180–185 - acute allograft ischemia vs., 101 - chronic chronic T-cell-mediated rejection vs., 130 protocol biopsies, 91 - clinical issues, 181 - de novo focal segmental glomerulosclerosis vs., 161 - diagnostic checklist, 182 - differential diagnosis, 182 - prognosis, 181 - transplant renal artery stenosis vs., 111 Calcineurin inhibitors, mechanism of action, 48 Calicivirus infection, adenovirus vs., 458 Calne, Roy, 246 Candida albicans, 211 Candida glabrata, 211 Candida infections - bacterial and fungal infections, 453 - lung, 407 Candida krusei, 211 Candida parapsilosis, 211 Candida tropicalis, 211 Candidiasis, 210–211, 235, 301 - aspergillosis vs., 217 - cryptococcosis vs., 213 - diagnostic checklist, 211 - differential diagnosis, 211 - histoplasmosis vs., 209

INDEX - microsporidiosis vs., 223 - mucormycosis vs., 215 - prognosis, 211 Capillaritis, 60 - antibody-mediated rejection vs., 494 - peritubular, acute T-cell-mediated rejection vs., 118 Capillaropathy, peritubular, chronic antibody-mediated rejection vs., 142 Cardiac allograft, 312 Cardiac allograft vasculopathy (CAV), 308, 349 Cardiac xenograft, 312 Cardiomyopathy - arrhythmogenic. See Arrhythmogenic right ventricular cardiomyopathy. - dilated, 318–321 arrhythmogenic right ventricular cardiomyopathy vs., 334 differential diagnosis, 320 explanted heart, focal endocardial fibrosis in, endstage heart disease vs., 338 genetics, 319 hypertrophic cardiomyopathy vs., 324 prognosis, 319 - drug-induced, end-stage heart disease related to, 337 - hypertrophic, 322–325 differential diagnosis, 324 dilated cardiomyopathy vs., 320 genetic testing, 324 prognosis, 323 - ischemic, sarcoidosis vs., 331 - secondary forms, hypertrophic cardiomyopathy vs., 324 Carrell, Alexis, 366, 512 Caseating granulomatous inflammation, tuberculosis, 225 Catheter-related infection, pancreas transplantation associated, 505 CAV. See Cardiac allograft vasculopathy. Cavadas, Pedro, 513 CD56 (NCAM), 14 CD56(+++) NK cells, 14 Cell-mediated rejection - intestinal allograft diseases, 412 - pancreas allograft diseases, 472 Cellular-mediated rejection, antibody-mediated rejection vs., 516 Cellular mediators, fibrosis mechanism, 86 Cellular rejection, acute - acute antibody-mediated rejection, intestine vs., 426 - grade B, bacterial infections vs., 401 - graft-vs.-host disease vs., 450 - intestine, 428–435 diagnostic checklist, 430–431 differential diagnosis, 430, 431 histologic grading, 431 prognosis, 429 - kidney, acute allograft ischemia vs., 101 - microaspiration vs., 399 - reperfusion injury vs., 419 - severe, chronic rejection, intestine vs., 437 Cellular/T-cell-mediated rejection, 234 CF. See Cystic fibrosis. CHD. See Congenital heart disease.

Chemical gastropathy, stomach rejection vs., 441 Child-Pugh Score, 246–247 Chimerism studies, 35 Cholangitis, recurrent primary sclerosing, 284–285 - bile duct stricture vs., 255 - bile leak vs., 255 - bile sludge vs., 255 - biloma vs., 255 - differential diagnosis, 285 - hepatic artery thrombosis vs., 251 - prognosis, 285 - (ductopenic) rejection vs., 271 Cholangitis lenta, graft-vs.-host disease vs., 305 Chronic active AMR. See Antibody-mediated rejection, chronic. Chronic active gastritis ± Helicobacter pylori, stomach rejection vs., 441 Chronic airway rejection, microaspiration vs., 399 Chronic allograft rejection - hepatic artery thrombosis vs., 251 - pancreas transplantation, 496–497 Banff grading, 497 diagnostic checklist, 497 differential diagnosis, 497 prognosis, 497 - portal vein thrombosis vs., 253 Chronic allograft vasculopathy, heart, 348–349 - diagnostic checklist, 349 - differential diagnosis, 349 - prognosis, 349 Chronic antibody-mediated rejection, 140–151. See also Chronic allograft rejection, pancreas transplantation. - accommodation vs., 95 - Banff 2017 criteria, 144, 268 - clinical issues, 141 - diagnostic checklist, 143 - differential diagnosis, 142–143 - prognosis, 141 - protocol biopsies, 91 - recurrent disease in the allograft vs., 155 - scoring, 144, 145 Chronic berylliosis, sarcoidosis of lung vs., 377 Chronic calcineurin inhibitor toxicity, protocol biopsies, 91 Chronic cholestatic disorders, liver transplantation, 236 Chronic graft-vs.-host disease in HSCT patients resulting in bronchiolitis obliterans, 381 Chronic hepatitis - other forms of, recurrent hepatitis B virus vs., 272–273 - plasma cell-rich rejection vs., 303 Chronic humoral rejection. See Antibody-mediated rejection, chronic. Chronic hypersensitivity pneumonitis, idiopathic pulmonary fibrosis vs., 373 Chronic lung allograft dysfunction, 394–395 - diagnostic checklist, 395 - prognosis, 395 Chronic mTOR inhibitor toxicity, 187 Chronic obstructive pulmonary disease (COPD). See Emphysema.

vii

INDEX Chronic pancreatitis, 506 - chronic allograft rejection/graft sclerosis vs., 497 Chronic pyelonephritis, chronic T-cell-mediated rejection vs., 130 Chronic (ductopenic) rejection, 234, 270–271. See also Chronic allograft vasculopathy, heart; Chronic lung allograft dysfunction. - acute cellular rejection, grade B vs., 393 - acute T-cell- and antibody-mediated rejection vs., 516 - diagnostic checklist, 271 - differential diagnosis, 271 - intestinal allograft diseases, 412 - intestine, 436–439 diagnostic checklist, 437 differential diagnosis, 437 prognosis, 437 - intraabdominal and opportunistic infections vs., 507 - lung, alloimmune response and, 362 - prognosis, 271 - recurrent primary biliary cirrhosis vs., 283 - recurrent primary sclerosing cholangitis vs., 285 - vascularized composite allotransplantation, 522–523 diagnostic checklist, 523 differential diagnosis, 523 prognosis, 523 Chronic T-cell-mediated rejection, 128–131. See also Chronic allograft rejection, pancreas transplantation. - categories, 130 - clinical issues, 129 - diagnostic checklist, 130 - differential diagnosis, 130 - prognosis, 129 Chronic transplant arteriopathy, 111 Chronic transplant glomerulopathy, de novo focal segmental glomerulosclerosis vs., 161 Chronic valvular disease, end-stage heart disease related to, 338 Chronic viral hepatitis - liver transplantation, 236 - recurrent autoimmune hepatitis vs., 281 - T-cell-mediated rejection vs., 264 CLAD. See Chronic lung allograft dysfunction. Classic Hodgkin lymphoma-type posttransplant lymphoproliferative disease, 528–529, 530 Clostridium difficile, bacterial and fungal infections, 453 Clostridium difficile (pseudomembranous) colitis, colon rejection vs., 445 Clostridium difficile enterocolitis, graft-vs.-host disease vs., 450 Clostridium difficile with ulcers and exudate, acute cellular rejection, intestine vs., 430 CMV. See Cytomegalovirus. CMV glomerulopathy. See Cytomegalovirus (CMV) infection. CMV infection. See Cytomegalovirus infection. CMV nephropathy. See Cytomegalovirus (CMV) infection. CMV tubulointerstitial nephritis (TIN). See Cytomegalovirus (CMV) infection. CNIT. See Calcineurin inhibitor toxicity. Coccidioides immitis, 219 viii

Coccidioides posadasii, 219 Coccidioides spherules, coccidioidomycosis, 219 Coccidioidomycosis, 218–219 - cryptococcosis vs., 213 - diagnostic checklist, 219 - differential diagnosis, 219 - histoplasmosis vs., 209 - prognosis, 219 Colon, native, adenovirus associated, 457 Colon rejection, 444–447 - diagnostic checklist, 445 - differential diagnosis, 445 - intestinal allograft diseases, 413 - prognosis, 445 Combined small bowel and colon transplant ± liver, intestinal transplants, 412 Common variable immunodeficiency, sarcoidosis of lung vs., 377 Community-acquired respiratory viruses, viral infections, 403 Complement, 20–23 - in atypical hemolytic uremic syndrome, 20–21 - atypical hemolytic uremic syndrome and transplant, 21–22 - eculizumab and atypical hemolytic uremic syndrome, 22 - genetic defects in atypical hemolytic uremic syndrome/thrombotic microangiopathy, 21 - laboratory testing for diagnosis of atypical hemolytic uremic syndrome and monitoring of eculizumab, 22–23 - regulation and dysregulation of alternate complement pathway, 21 - in solid organ transplantation immune response, 5 Congenital heart disease, 326–329 - prognosis, 327 Congestive heart failure, hepatic venous outflow obstruction and, 257 Congestive hepatic venopathy. See Hepatic venous outflow obstruction. Congestive hepatopathy. See Hepatic venous outflow obstruction. Connective tissue disease (CTD), 375 Connective tissue disease-associated interstitial lung disease, idiopathic pulmonary fibrosis vs., 373 Connective tissue disease-associated lung disease, 374–375 - diagnostic checklist, 375 - differential diagnosis, 375 - prognosis, 375 Contact dermatitis - allergic, acute T-cell- and antibody-mediated rejection vs., 517 - irritant, acute T-cell- and antibody-mediated rejection vs., 517 Cooper, Joel, 366 Coronary artery disease. See also Ischemic heart disease. - atherosclerotic, chronic allograft vasculopathy, heart vs., 349 Coronary heart disease. See Ischemic heart disease. Cortical abscesses, candidiasis, 211

INDEX Corticosteroids, mechanism of action, 48 CR. See Chronic (ductopenic) rejection. Crescentic glomerulonephritis, cryptococcosis, 213 Crossmatch testing, 35 Crypt apoptosis - cytomegalovirus infection associated, 461 - Epstein-Barr virus, 466 Cryptococcosis, 212–213, 235, 301 - candidiasis vs., 211 - coccidioidomycosis vs., 219 - differential diagnosis, 213 - histoplasmosis vs., 209 - paracoccidioidomycosis vs., 221 - prognosis, 213 Cryptococcus gattii, 213 Cryptococcus infections - bacterial and fungal infections, 453 - lung, 407 Cryptococcus neoformans, 213 Cryptogenic fibrosing alveolitis. See Idiopathic pulmonary fibrosis (IPF). CTD. See Connective tissue disease. CTD-associated interstitial lung disease (CTD-ILD). See Connective tissue disease-associated lung disease. CTD-associated pulmonary arterial hypertension (CTDPAH). See Connective tissue disease-associated lung disease. Cutaneous pseudolymphoma, acute T-cell- and antibodymediated rejection vs., 517 Cyanotic heart disease, 327 Cyclosporine (CsA) - history of use, 54 - islet amyloid deposition and islet cell toxicity associated, 503 - mechanism of action, 48 - therapeutic monitoring, 50–51 - therapeutic ranges for trough concentrations, 53 Cystic fibrosis (CF), 370–371 - diagnostic checklist, 371 - differential diagnosis, 371 - prognosis, 371 Cytokine-producing CD56 (+++), 14 Cytomegalovirus (CMV), 205, 235, 288–289 - adenovirus vs., 293, 458 - antigen-specific assessment, of T-cell immune competence, 26 - bacterial and fungal infections vs., 454 - chronic rejection, intestine vs., 437 - diagnostic checklist, 289 - differential diagnosis, 289 - Epstein-Barr virus vs., 298, 466 - hepatitis E virus vs., 295 - herpes simplex virus vs., 291 - intestine, 460–463 diagnostic checklist, 462 differential diagnosis, 462 prognosis, 461 - pancreas transplantation associated, 505, 506 - pancreatitis acute cellular rejection vs., 490

intraabdominal and opportunistic infections vs., 507 - prognosis, 289 Cytomegalovirus (CMV) infection, 204–207, 403 - acute cellular rejection, intestine vs., 430 - diagnostic checklist, 206 - differential diagnosis, 206 - graft-vs.-host disease vs., 450 - on immune system, 205 - immunohistochemistry, 206 - prognosis, 205–206 Cytoplasmic vacuolization, 315 Cytotoxic NK cells, 14

D

DAAs. See Direct-acting antivirals. Daclizumab - history of use, 55 - islet amyloid deposition and islet cell toxicity associated with, 503 - therapeutic monitoring, 52 Danon disease, hypertrophic cardiomyopathy vs., 324 Darling disease. See Histoplasmosis. Date, Hiroshi, 366 DCM. See Dilated cardiomyopathy. De novo anti-GBM nephritis. See Antiglomerular basement membrane (anti-GBM) disease in Alport syndrome. De novo autoimmune hepatitis. See Plasma cell-rich rejection. De novo focal segmental glomerulosclerosis, 160–161 - clinical issues, 161 - diagnostic checklist, 161 - differential diagnosis, 161 - prognosis, 161 De novo glomerular disease, protocol biopsies, 91 De novo glomerulopathies, late allograft loss, 80 De novo membranous glomerulonephritis, 162–165 - clinical issues, 163 - diagnostic checklist, 163 - differential diagnosis, 163 - prognosis, 163 Death, definition, 246 Decompensated cirrhosis - liver transplantation, 236 - native liver removal, 237 Demetris, A. J., 247 Demikov, Vladimir, 366 Dermatoses, lichenoid, acute T-cell- and antibodymediated rejection vs., 517 Derom, F., 366 Diabetes mellitus - recurrent, 498–501 animal models, 499 autoimmune mechanisms, 499 chronic allograft rejection/graft sclerosis vs., 497 differential diagnosis, 500 prognosis, 499 - recurrent autoimmune, 472 ix

INDEX Diabetic mothers, infants of, hypertrophic cardiomyopathy vs., 324 Diabetic nephropathy - antiglomerular basement membrane (anti-GBM) disease in Alport syndrome vs., 167 - in end-stage kidney, 64 Diarrhea, rotavirus associated, 461 Diffuse alveolar damage, organizing, organizing pneumonia vs., 397 Dilated cardiomyopathy, 318–321 - arrhythmogenic right ventricular cardiomyopathy vs., 334 - differential diagnosis, 320 - explanted heart, focal endocardial fibrosis in, end-stage heart disease vs., 338 - genetics, 319 - hypertrophic cardiomyopathy vs., 324 - prognosis, 319 Direct-acting antivirals, 275 Diseases of native stomach, stomach rejection vs., 441 Distal nephron, polyomavirus nephritis, 193 Donor artery atherosclerosis, 111 Donor biopsy, 242 - performance, 242 - utility, 242 Donor-derived membranous glomerulonephritis, de novo membranous glomerulonephritis vs., 163 Donor frozen section findings, 243 Donor kidney evaluation, 74–79 - biopsy, 75 histologic features, 75 Maryland Aggregate Pathology Index (MAPI), 75 sample adequacy, 75 - donor biopsy reporting and scoring sheet, 76 - following findings, scored, 76 - pitfalls, 76 frozen section artifacts, 76 specimen handling, 76 superficial biopsy, 76 wedge biopsies, 76 - reporting, 75–76 - specimen evaluation, 74–75 frozen section, 74 gross, 74 reliability, 74–75 - surgical/clinical considerations, 74 change in patient management, 74 clinical setting, 74 goal of consultation, 74 kidney acceptance criteria, 74 Donor liver evaluation, 242–245 Donor organ evaluation, 242–243 Donor-specific antibodies, antibody-mediated rejection associated, 493 Donor thrombotic microangiopathy, hyperacute rejection vs., 113 Donor vascular disease, chronic rejection vs., 523 DRESS syndrome, acute T-cell- and antibody-mediated rejection vs., 517

x

Drug eruptions, lichenoid, acute T-cell- and antibodymediated rejection vs., 517 Drug-induced acute hepatitis with zone 3 necrosis, hepatic venous outflow obstruction vs., 258 Drug-induced acute interstitial nephritis - adenovirus, kidney vs., 201 - histoplasmosis vs., 209 Drug-induced cardiomyopathy, end-stage heart disease related to, 337 Drug-induced enterocolitis, graft-vs.-host disease vs., 450 Drug-induced hepatitis, adenovirus vs., 293 Drug-induced interstitial nephritis - acute pyelonephritis vs., 190 - tuberculosis vs., 225 Drug-induced liver injury, graft-vs.-host disease vs., 305 Drug-induced vanishing bile duct syndrome, 234 - chronic (ductopenic) rejection vs., 271 Drug reactions/toxicity - acute T-cell- and antibody-mediated rejection vs., 517 - intravenous drug abuse, sarcoidosis of lung vs., 377 - late allograft loss, 80 - pancreas allograft diseases, 472 Dubernard, Jean-Michel, 512 Duct obstruction pancreatitis, recurrent diabetes mellitus vs., 500 Ductopenic rejection. See Chronic (ductopenic) rejection. Ductulitis, acute cellular rejection associated, 489 Dysfunctional anastomosis, 105 Dyskeratosis, acute T-cell- and antibody-mediated rejection associated, 515

E Early chronic (ductopenic) rejection, 234 Early clinical era, liver transplantation, 246 Early graft failure, 240, 241 EBV. See Epstein-Barr virus. Eculizumab - atypical hemolytic uremic syndrome, 22 - laboratory testing for diagnosis of atypical hemolytic uremic syndrome and monitoring of, 22–23 - mechanism of action, 49 Electron microscopy - hypertrophic cardiomyopathy, 324 - polyomavirus nephritis, 194 Embolism, atheromatous, acute T-cell-mediated rejection vs., 118 Emphysema, 368–369 - diagnostic checklist, 369 - differential diagnosis, 369 - other causes of end-stage lung disease vs., 382 - prognosis, 369 Emphysematous pyelonephritis, 189–190 Encephalitozoon bieneusi, 223 Encephalitozoon cuniculi, 223 Encephalitozoon intestinalis, 223 Endarteritis, 60

INDEX Endocardial fibroelastosis - end-stage heart disease related to, 337 - focal endocardial fibrosis in, end-stage heart disease vs., 338 - primary, end-stage heart disease related to, 338 Endocrine portal venous drainage, 482 Endocrine systemic venous drainage, 482 Endogenous lipoid pneumonia/postobstructive, microaspiration vs., 399 Endomyocardial biopsy, indications, 310 Endothelial injury/activation, acute cellular rejection associated, 489 Endothelialitis, 60 - isolated, acute T-cell-mediated rejection vs., 118 End-stage kidney disease (ESKD). See also End-stage kidney. - other causes, 336–341 differential diagnosis, 338 End-stage kidney evaluation, 62–67 - epidemiology, 62 - etiology/pathogenesis, 62–63 in adults, 62 in children aged less than 18 years, 62–63 pathogenesis, 63 - macroscopic, 63 kidney size, 63 renal medulla, pathologic features of, 63 secondary changes in end-stage kidney disease, 63 - microscopic, 63–64 diabetic nephropathy, 64 general features, 63 glomerular diseases, 64 hypertensive nephrosclerosis, 64 secondary changes of end-stage kidney disease, 64 tubulointerstitial diseases, 64 vascular diseases, 64 - terminology, 62 End-stage lung disease, other causes, 380–383 - diagnostic checklist, 382 - differential diagnosis, 383 - prognosis, 381 End-stage renal disease (ESRD). See End-stage kidney. Engraftment syndrome (ES), 170–173 - clinical issues, 171 - differential diagnosis, 171 - prognosis, 171 Enteric infections, graft-vs.-host disease vs., 450 Enteritis - Epstein-Barr virus, 465 - infectious, 457 acute antibody-mediated rejection, intestine vs., 426 - intestinal allograft diseases, 412 Enterobacter infections, 453 Enterococcus faecalis, pancreas transplantation associated, 505 Enterococcus infections, 453 Enterocolitis, drug-induced, graft-vs.-host disease vs., 450 Eosinophilic intranuclear inclusions, adenovirus, 457 Eosinophilic pneumonia, organizing pneumonia vs., 397 Epithelial-to-mesenchymal phenotype, fibrosis mechanism, 86

Epstein-Barr virus (EBV), 235, 296–299 - antigen-specific assessment, of T-cell immune competence, 26–27 - diagnostic checklist, 298 - differential diagnosis, 298 - hepatitis E virus vs., 295 - intestine, 464–469 characteristics, 466 differential diagnosis, 466 prognosis, 465 - pancreas transplantation associated with, 505 - posttransplant lymphoproliferative disease associated with, 527 - prognosis, 297 - viral infections, 403 Epstein-Barr virus colitis, colon rejection vs., 445 Epstein-Barr virus enteritis, acute cellular rejection, intestine vs., 430 Epstein-Barr virus gastritis, stomach rejection vs., 441 Erosive gastritis, acute, stomach rejection vs., 441 Erythema multiforme, acute T-cell- and antibody-mediated rejection vs., 517 ES. See Engraftment syndrome. Escherichia coli, 227 - bacterial and fungal infections, 453 - pancreas transplantation associated, 505 Everolimus - history of use, 54 - mechanism of action, 49 - therapeutic monitoring, 51 - therapeutic ranges for trough concentrations, 53 Ex vivo perfusion, 59 Exfoliative rejection, 430 Exocrine bladder drainage, 482 Exocrine enteric drainage, 482 Exogenous lipoid pneumonia. See also Microaspiration. - fungal infections, lung vs., 407 Experimental drugs, 49 - bortezomib, 49 - eculizumab, 49 - IgG endopeptidase, 49 - intravenous immunoglobulin, 49 - rituximab, 49 Explant, indications and evaluation, 310, 414–417 Explanted heart, evaluation, 311 Explanted lung, 362 Extended donor criteria, 242 Extracorporeal photopheresis (ECP), and transplantation, 42

F Fabry disease, hypertrophic cardiomyopathy vs., 324 Fageeh, Wafa, 512 Failed liver allograft, evaluation of, 240–241 - causes, 240 - clinical presentation, 240 - specimen handling, 240–241 xi

INDEX Failed native and transplanted heart, evaluation of, 310–311 Failed native liver - approach, 236–237 - gross evaluation of, 236–239 Familial adenomatous polyposis, explant, 415 Fatty acid metabolic disorders, hypertrophic cardiomyopathy vs., 324 Fatty liver disease, 235 - recurrent, 286–287 differential diagnosis, 287 prognosis, 287 FCH. See Fibrosing cholestatic hepatitis. Fever, acute rheumatic, myocarditis vs., 357 Fibroelastosis, endocardial - end-stage heart disease related to, 337 - focal endocardial fibrosis in, end-stage heart disease vs., 338 - primary, end-stage heart disease related to, 338 Fibropolycystic disease, native liver removal, 237 Fibrosing cholestatic hepatitis, 279 Fibrosing cholestatic hepatitis B or C virus, 278–279 - diagnostic checklist, 279 - differential diagnosis, 279 - prognosis, 279 Fibrosing cytolytic liver failure. See Fibrosing cholestatic hepatitis B or C virus. Fibrosis - acute T-cell- and antibody-mediated rejection vs., 516 - cystic, 370–371 diagnostic checklist, 371 differential diagnosis, 371 prognosis, 371 - donor organ, 243 - focal endocardial in dilated cardiomyopathy, end-stage heart disease vs., 338 - interstitial, and tubular atrophy, not otherwise specified, protocol biopsies, 91 - stimulation of, chronic allograft rejection/graft sclerosis associated, 497 Fibrosis evaluation, 86–89 - mechanisms, 86 cellular mediators, 86 epithelial-to-mesenchymal phenotype, 86 molecular mediators, 86 - methods for assessment, 86–87 morphometric quantitative assessment, 87 qualitative visual assessment, 86–87 quantitative visual assessment, 87 - value, 86 Fibroviral hepatitis. See Fibrosing cholestatic hepatitis B or C virus. Fluid collection, intraabdominal and opportunistic infections vs., 507 Focal abscess, recurrent diabetes mellitus vs., 500 Focal endocardial fibrosis, in dilated cardiomyopathy, endstage heart disease vs., 338 Focal necrosis, recurrent diabetes mellitus vs., 500 Focal segmental glomerulosclerosis (FSGS) - calcineurin inhibitor toxicity vs., 182 xii

- de novo, 160–161 clinical issues, 161 diagnostic checklist, 161 differential diagnosis, 161 prognosis, 161 - lesions, 187 - mTOR inhibitor toxicity vs., 187 - recurrent, hyperperfusion injury vs., 169 Formalin perfusion, overinflation artifact from, emphysema vs., 369 Friedrich ataxia, hypertrophic cardiomyopathy vs., 324 FSGS. See Focal segmental glomerulosclerosis. Fulminant viral myocarditis, myocarditis and, 355 Fungal infections, 235, 300–301 - acute T-cell- and antibody-mediated rejection vs., 516 - cytomegalovirus vs., 462 - diagnosis, 301 - graft-vs.-host disease vs., 305 - herpes simplex virus vs., 462 - intestinal allograft diseases, 412 - intestinal transplantation, 452–455 diagnostic checklist, 454 differential diagnosis, 454 infectious agents, 453 prognosis, 453 - intraabdominal and opportunistic infections vs., 507 - lung, 407–409 differential diagnosis, 407 prognosis, 407 - microaspiration vs., 399 - nonalloimmune diseases and, 362 - pancreas allograft diseases, 473 - prognosis, 301 - recurrent diabetes mellitus vs., 500 - rotavirus vs., 462 - sarcoidosis of lung vs., 377 Fungal organisms - aspergillosis, 217 - candidiasis, 211 - cryptococcosis, 213 - paracoccidioidomycosis, 221 Fungal pancreatitis, 506 Fusariosis - aspergillosis vs., 217 - candidiasis vs., 211 - mucormycosis vs., 215

G Gago, Otto, 366 Garfein, Evan S., 367 Gastric rejection, intestinal allograft diseases, 413 Gastritis, Epstein-Barr virus, stomach rejection vs., 441 Gastroschisis, explant, 414 Genetic testing, hypertrophic cardiomyopathy, 324 Giant cell myocarditis, 356 - myocarditis vs., 356 - sarcoidosis vs., 331 Gilles, Harold, 512

INDEX Glomerular disease - de novo protocol biopsies, 91 recurrent disease in the allograft vs., 154 - donor-transmitted, recurrent disease in the allograft vs., 154 - in end-stage kidney, 64 - recurrent, protocol biopsies, 91 Glomeruli - adenovirus, kidney, 201 - microsporidiosis, 223 Glomerulitis, acute transplant, recurrent disease in the allograft vs., 155 Glomerulonephritis - de novo membranous, kidney transplantation, 162–165 clinical issues, 163 diagnostic checklist, 163 differential diagnosis, 163 prognosis, 163 - immune complex-mediated, de novo or recurrent, de novo focal segmental glomerulosclerosis vs., 161 - primary, membranous, graft-vs.-host glomerulopathies vs., 179 - tuberculosis, 225 Glomerulopathy - graft vs. host, 178–179 clinical issues, 179 differential diagnosis, 179 prognosis, 179 - transplant chronic, de novo focal segmental glomerulosclerosis vs., 161 chronic antibody-mediated rejection vs., 142 protocol biopsies, 91 Glomerulosclerosis - de novo focal segmental, 160–161 clinical issues, 161 diagnostic checklist, 161 differential diagnosis, 161 prognosis, 161 - donor biopsy, 75 - focal segmental, calcineurin inhibitor toxicity vs., 182 - recurrent focal segmental, de novo focal segmental glomerulosclerosis vs., 161 Glucocorticoids, islet amyloid deposition and islet cell toxicity associated, 503 Glutamic acid decarboxylase (GAD65), antibodies against, recurrent diabetes mellitus associated, 499 Glycogen storage disease, hypertrophic cardiomyopathy vs., 324 Graft dysfunction, primary, acute cellular rejection, grade A vs., 391 Graft failure - antibody-mediated rejection associated, 493 - causes, 415–416 - pancreas transplantation associated, 474 Graft ischemia, cytomegalovirus vs., 289 Graft loss, intestinal allograft diseases, 413 Graft pancreatitis, 482 - pancreas transplantation associated, 483

Graft rejection, cytomegalovirus vs., 289 Graft resection, Epstein-Barr virus associated, 465 Graft sclerosis - antibody-mediated rejection associated, 493 - pancreas transplantation, 496–497 Banff grading, 497 diagnostic checklist, 497 differential diagnosis, 497 prognosis, 497 Graft survival, following intestinal transplantation, 415 Graft tenderness, acute cellular rejection associated, 489 Graft thrombosis - pancreas, antibody-mediated rejection vs., 494 - pancreas transplantation associated, 482 Graft-vs.-host disease, 234 - colon rejection vs., 445 - fasciitis, 517 - HLA testing and, 37 - intestine, 448–451 diagnostic checklist, 450 differential diagnosis, 450 prognosis, 449 - lichenoid, acute T-cell- and antibody-mediated rejection vs., 517 - liver, 304–305 differential diagnosis, 305 prognosis, 305 - morpheic, 517 - prevention and treatment of, 37 - sclerotic, 517 - stomach rejection vs., 441 - T-cell- and antibody-mediated rejection vs., 517 - transfusion-associated, 45 Graft-vs.-host glomerulopathies, 178–179 - clinical issues, 179 - differential diagnosis, 179 - prognosis, 179 Granulomas - adenovirus, kidney, 201 - paracoccidioidomycosis, 221 Granulomatosis with polyangiitis - and other vasculitides, myocarditis vs., 356 - sarcoidosis of lung vs., 377 Granulomatous interstitial inflammation - coccidioidomycosis, 219 - cryptococcosis, 213 Granulomatous interstitial nephritis - candidiasis, 211 - histoplasmosis, 209 - mucormycosis, 215 - paracoccidioidomycosis, 221 Grocott-Gomori methenamine silver, nocardiosis, 231 Groth, Carl, 246 GVHD. See Graft-vs.-host disease.

H Hardy, James, 366 Harvesting injury. See Preservation injury. xiii

INDEX HAT. See Hepatic artery thrombosis. Haverich, Axel, 366 HBcAg. See Hepatitis B core antigen. HBcAG(+) organ donor, evaluation, 243 HBsAg. See Hepatitis B surface antigen. HBV. See Hepatitis B virus. HCM. See Hypertrophic cardiomyopathy. HCT. See Hematopoietic cell transplantation. HCV. See Hepatitis C virus. Heart - acute cellular rejection, 342–345 diagnostic checklist, 343 differential diagnosis, 343 prognosis, 343 Quilty effect, acute cellular rejection, heart vs., 343 - ischemic disease, 314–317 differential diagnosis, 315 dilated cardiomyopathy vs., 320 prognosis, 315 - normal, myocarditis vs., 356 Heart allograft - evaluation of failed native and transplanted heart, 310–311 - pathologic classification of cardiac allograft disease, 308–309 - vasculopathy, Quilty lesions vs., 351 Heart allograft rejection - acute cellular, 342–345 diagnostic checklist, 343 differential diagnosis, 343 prognosis, 343 Quilty lesions vs., 351 site of previous biopsy vs., 353 - antibody-mediated, 346–347 diagnostic checklist, 347 differential diagnosis, 347 prognosis, 347 - diffuse mild cellular, antibody-mediated, heart allograft rejection vs., 347 - Quilty lesions, 350–351 diagnostic checklist, 351 differential diagnosis, 351 prognosis, 351 site of previous biopsy vs., 353 - site of previous biopsy, 352–353. See also Heart allograft rejection, site of previous biopsy. acute cellular rejection, heart vs., 343 diagnostic checklist, 353 differential diagnosis, 353 prognosis, 353 Quilty lesions vs., 351 Heart allograft vasculopathy, chronic, 348–349 - diagnostic checklist, 349 - differential diagnosis, 349 - prognosis, 349 Heart disease - congenital, 326–329 prognosis, 327 - healed inflammation in, ischemic disease of explanted heart vs., 315 - ischemic, dilated cardiomyopathy vs., 320 xiv

- valvular, end-stage heart disease related to, 337 Heart ischemic injury, perioperative, acute cellular rejection, heart vs., 343 Heart transplantation - antibody-mediated rejection, heart, 346–347 - chronic allograft vasculopathy, heart, 348–349 diagnostic checklist, 349 differential diagnosis, 349 prognosis, 349 - congenital heart disease, 326–329 prognosis, 327 - dilated cardiomyopathy, 318–321 differential diagnosis, 320 genetics, 319 prognosis, 319 - evaluation of failed native and transplanted heart, 310–311 - explanted heart evaluation arrhythmogenic right ventricular cardiomyopathy, 332–335 end-stage heart disease, other causes, 336–341 sarcoidosis, myocarditis vs., 356 - history of, 312–313 milestones, 313 treatment issues, 313 - HLA matching, 37 - hypertrophic cardiomyopathy, 322–325 differential diagnosis, 324 genetic testing, 324 prognosis, 323 - infection, acute cellular rejection, heart vs., 343 - ischemic heart disease, 314–317 differential diagnosis, 315 prognosis, 315 - myocarditis, 354–359 differential diagnosis, 356–357 infectious causes of, 357 noninfectious causes of, 357 prognosis, 355–356 - pathologic classification of cardiac allograft disease, 308–309 - Quilty lesions, 350–351 - sarcoidosis, 330–331 differential diagnosis, 331 genetic background, 331 prognosis, 331 Heart-lung transplantation, 362 HELLP syndrome, liver removal, 237 Hematogenous infection, acute pyelonephritis, 189, 190 Hematoma, kidney, lymphocele vs., 107 Hematopoietic cell transplantation, 305 Hemochromatosis - dilated cardiomyopathy vs., 320 - end-stage heart disease related to, 337 Hemolytic uremic syndrome - atypical complement in, 20–21 eculizumab and, 22 genetic defects in, 21 laboratory testing for diagnosis of, 22–23 and transplant, 21–22

INDEX - early allograft loss, 80 - recurrent atypical acute antibody-mediated rejection vs., 134 hyperacute rejection vs., 113 renal artery or vein thrombosis vs., 109 Hemorrhagic infarction, pancreas transplantation associated, 484 Hepatic artery thrombosis, 234, 250–251 - diagnostic checklist, 251 - differential diagnosis, 251 - portal vein thrombosis vs., 253 - preservation injury vs., 249 - prognosis, 251 - recurrent primary sclerosing cholangitis vs., 285 Hepatic tumor - liver transplantation, 236 - native liver removal, 237 Hepatic vein stenosis, preservation injury vs., 249 Hepatic vein thrombosis, preservation injury vs., 249 Hepatic venous outflow obstruction, 234, 256–259 - differential diagnosis, 258 - prognosis, 257–258 Hepatitis - alcoholic, hepatic venous outflow obstruction vs., 258 - autoimmune, 235 de novo. See Plasma cell-rich rejection. liver transplantation, 236 recurrent, 280–281 - chronic other forms of, recurrent hepatitis B virus vs., 272–273 plasma cell-rich rejection vs., 303 - chronic viral (hepatitis B or C), T-cell-mediated rejection vs., 264 liver transplantation, 236 recurrent autoimmune hepatitis vs., 281 - de novo autoimmune. See Plasma cell-rich rejection. - drug-induced acute, with zone 3 necrosis, hepatic venous outflow obstruction vs., 258 adenovirus vs., 293 - fibrosing cholestatic, 279 - plasma cell (de novo AIH), recurrent autoimmune hepatitis vs., 281 - recurrent autoimmune, 280–281 plasma cell-rich rejection vs., 303 T-cell-mediated rejection vs., 264 - recurrent B virus, 272–273 - recurrent C virus, 274–277 - surgical, preservation injury vs., 249 Hepatitis A virus, hepatitis E virus vs., 295 Hepatitis B core antigen, 273 Hepatitis B surface antigen, 273 Hepatitis B virus, 235 - fibrosing cholestatic HBV or HCV hepatitis, 278–279 diagnostic checklist, 279 differential diagnosis, 279 prognosis, 279 - hepatitis E virus vs., 295 - infections, 403

- recurrent, 272–273 diagnostic checklist, 273 differential diagnosis, 273 prognosis, 273 Hepatitis C virus, 235 - fibrosing cholestatic HBV or HCV hepatitis, 278–279 diagnostic checklist, 279 differential diagnosis, 279 prognosis, 279 - hepatitis E virus vs., 295 - infections, 403 severe recurrent, fibrosing cholestatic hepatitis B or C virus vs., 279 - recurrent, 274–277 diagnostic checklist, 276 differential diagnosis, 276 Epstein-Barr virus, liver vs., 298 genotype, fatty liver disease vs., 287 prognosis, 275 Hepatitis E virus, 235, 294–295 - diagnostic checklist, 295 - differential diagnosis, 295 - prognosis, 295 Hepatocyte injury, preservation injury, 249 Hepatocyte necrosis, donor organ, 243 Hepatosplenic T-cell lymphomas, monomorphic T-/NK-cell PTLD, 528 Hereditary amyloidosis, 337 Herpes simplex virus (HSV), 235, 290–291 - adenovirus vs., 293 - diagnostic checklist, 291 - differential diagnosis, 291 - Epstein-Barr virus, liver vs., 298 - hepatitis E virus vs., 295 - infections, 403 - intestine, 460–463 diagnostic checklist, 462 differential diagnosis, 462 prognosis, 461 - pancreas transplantation associated, 505 - prognosis, 291 Herpesviridae, 205 HEV. See Hepatitis E virus. HHV-6. See Human herpesviruses 6. HHV-7. See Human herpesviruses 7. High-grade urothelial, polyomavirus nephritis, 193 Hirschsprung disease, explant, 414–415 Histocompatibility testing, 34–35 - additional testing, 35 allograft gene expression profiling, 35 chimerism studies, 35 immune function monitoring, 35 - anti-HLA antibody testing, 34–35 - crossmatch testing, 35 - HLA typing, 34 - major tests in, 34 Histoplasma capsulatum, 209 Histoplasmosis, 208–209 - cryptococcosis vs., 213 - diagnostic checklist, 209 - differential diagnosis, 209 xv

INDEX - fungal infections, lung, 407 - prognosis, 209 History of immunosuppression drugs, 54–55 - antibody therapies, 54–55 approved for other indications, 55 approved for use in transplantation, 54–55 - chronology of discovery, 55 - drugs in clinical use (1962-1994), 54 - modern era (1995-present), 54 - overview, 54 History of transplantation - heart, 312–313 milestones, 313 treatment issues, 313 - intestinal, 422–423 - kidney, 58–59 alloantibody testing, 59 allograft tolerance, 59 chronology and evolution, 58 ex vivo perfusion, 59 laboratory testing, 58–59 xenotransplantation, 59 - liver, 246–247 chronology and evolution, 246 liver allograft pathology, 246–247 organ allocation policy, 246–247 - lung, 366–367 landmark events in, 366 timeline of significant events in, 367 - pancreas, 480–481 chronology and evolution, 480 progress in treatment, 480 - vascularized composite allotransplantation, 512–513 HLA and transfusion, 44–45 - foreign (nonself) antigen exposure, 44 - human leukocyte antigen system, 44 - human leukocyte antigen-related immunological transfusion reactions, 44–45 donor human leukocyte antigen related, 44–45 febrile nonhemolytic transfusion reactions, 44 platelet refractoriness, 44 recipient human leukocyte antigen related, 44 transfusion-associated graft-vs.-host disease, 45 transfusion-related acute lung injury, 45 HLA molecules, 60 HLA testing - ABO blood group antigens, 40–41 ABH antigens, 40 ABO antibodies, 40 ABO blood groups, 41 barrier to transplantation, 41 current status, 41 expression of ABH antigen, 40 humoral immune response, 40 rationale for crossing ABO barrier, 41 transplantation and, 40–41 transplantation of A2/A2B donor kidneys into blood group B candidates, 41 - apheresis and transplantation, 42–43 desensitization, ABO-incompatible transplantation, 42 xvi

desensitization, donor-specific anti-HLA antibodies, 42–43 ECP and transplantation, 42 posttransplant treatment of rejection, 43 recurrent focal segmental glomerulosclerosis, 43 therapeutic apheresis, 42 transplant-associated thrombotic microangiopathy, 43 - histocompatibility testing, 34–35 additional testing, 35 anti-HLA antibody testing, 34–35 crossmatch testing, 35 HLA typing, 34 major tests in, 34 - HLA and transfusion, 44–45 foreign (nonself) antigen exposure, 44 human leukocyte antigen system, 44 human leukocyte antigen-related immunological transfusion reactions, 44–45 - human leukocyte antigen system, 32–33 antihuman leukocyte antigen antibodies, 33 genetics, 32 naming system for human leukocyte antigen genes and allele sequences, 33 properties, 32 structure, 32–33 - immune responses to allografts, 36–37 graft-vs.-host disease, 37 prevention and treatment of, 37 HLA antigens, 36 prevention and treatment of rejection, 36–37 rejection, 36 types of rejection, 36 - transplantation and HLA, 36–39 anti-HLA antibodies, 38 HLA antigens and matching resolution level, 39 HLA matching, 37–38 immune responses to allografts, 36–37 methods to detect anti-HLA antibodies, 39 types of transplants, 36 HLA-DR molecules, acute T-cell-mediated rejection associated, 515 Hofmann, Gunther O., 512 HSCR. See Hirschsprung disease. HSV. See Herpes simplex virus. Hu, Weilie, 512 Human herpesvirus 4. See Epstein-Barr virus. Human herpesviruses 6, viral infections, 403 Human herpesviruses 7, viral infections, 403 Human islet transplantation, 476 - advantages, 476 - definition, 481 - disadvantages, 476 - history, 481 - sources of islet cells, 476 - technique and location, 476 Human leukocyte antigen matching, pancreas transplantation, 475

INDEX Human leukocyte antigen system, 32–33. See also HLA testing. - antihuman leukocyte antigen antibodies, 33 - genetics, 32 classic human leukocyte antigens, 32 haplotype, 32 major histocompatibility complex, 32 - naming system for human leukocyte antigen genes and allele sequences, 33 - properties, 32 class I human leukocyte antigen molecules, 32 class II human leukocyte antigen molecules, 32 - structure, 32–33 class I, 32–33 class I and class II, 32 class II, 33 Human polyomavirus, 193 Humoral rejection. See Antibody-mediated rejection. Hurler disease, hypertrophic cardiomyopathy vs., 324 HVOO. See Hepatic venous outflow obstruction. Hyperacute antibody-mediated rejection, hyperperfusion syndrome vs., 261 Hyperacute rejection, 234. See also Antibody-mediated rejection. - intestinal, 425 - kidney transplantation, 112–115 clinical issues, 113 differential diagnosis, 113 prognosis, 113 Hyperfiltration injury. See Hyperperfusion injury. Hyperglycemia - acute cellular rejection associated, 489 - after pancreatic transplantation, causes, 499 - chronic allograft rejection/graft sclerosis associated, 497 Hypergranulosis, acute T-cell- and antibody-mediated rejection associated, 516 Hyperkeratosis, acute T-cell- and antibody-mediated rejection associated, 515 Hyperperfusion injury, 168–173 - clinical issues, 169 - diagnostic checklist, 169 - differential diagnosis, 169 - prognosis, 169 Hyperperfusion syndrome, 234 - liver, 260–261 differential diagnosis, 261 prognosis, 261 Hypersensitivity pneumonitis - chronic idiopathic pulmonary fibrosis vs., 373 sarcoidosis of lung vs., 377 - organizing pneumonia vs., 397 Hypertension - pancreas transplantation associated, 475 - pulmonary arterial, 378–379 diagnostic checklist, 379 differential diagnosis, 379 prognosis, 379

Hypertension-associated hypertrophy, hypertrophic cardiomyopathy vs., 324 Hypertensive arteriosclerosis, chronic T-cell-mediated rejection vs., 130 Hypertensive nephrosclerosis, in end-stage kidney, 64 Hypertrophic cardiomyopathy, 322–325 - differential diagnosis, 324 - dilated cardiomyopathy vs., 320 - genetic testing, 324 - prognosis, 323 Hypertrophy, hypertension-associated, hypertrophic cardiomyopathy vs., 324

I

Idiopathic DCM. See Dilated cardiomyopathy. Idiopathic hypertrophic subaortic stenosis. See Hypertrophic cardiomyopathy. Idiopathic pulmonary fibrosis (IPF), 372–373 - diagnostic checklist, 373 - differential diagnosis, 373 - prognosis, 373 IFI. See Invasive fungal infection. IgG endopeptidase, mechanism of action, 49 Immune complex-mediated glomerulonephritis, de novo or recurrent, de novo focal segmental glomerulosclerosis vs., 161 Immune function monitoring, 35 Immune monitoring, laboratory-based, 24–29 - approaches to immune monitoring, 24–25 global immune function analysis, 25 global immunophenotyping analysis by flow cytometry, 24 molecular methods, 25 - immune monitoring in organ transplantation, 24 - immune response biomarkers, 25–26 immune activation markers, 25 immune tolerance markers, 25 - infections in solid organ transplantation, 26–27 antigen-specific assessment of T-cell immune competence, 26–27 - relevance and application of immune monitoring, 27 Immune response biomarkers, 25–26 - immune activation markers, 25 - immune tolerance markers, 25 Immune responses - allografts, 36–37 graft-vs.-host disease, 37 HLA antigens, 36 prevention and treatment of rejection, 36–37 rejection, 36 types of rejection, 36 - innate, 4–5 complement, 5 dendritic cells, 5 innate immunity and complement in allograft rejection and tolerance, 4–5 macrophages, 5 xvii

INDEX neutrophils, 5 NK cells, 5 nucleotide-binding oligomerization domain (NOD)like receptors (NLRs), 4 PAMPs (pathogen-associated molecular patterns), 4 PRRs (pathogen-recognition receptors), 4 RIG-like helicases (RLHs), 4–5 Toll-like receptors (TLRs), 4 - organ transplantation, 4–7 B-cell immune responses, 6 defining, 4 immunosuppressive therapy in, 7 infection, tissue damage and immune response, 5–6 innate immune responses and, 4–5 manipulation of, 7 NF-kB and T cells in, 6–7 NK cells and, 6 Immunodeficiency, common variable, sarcoidosis of lung vs., 377 Immunodeficiency disorders, cystic fibrosis vs., 371 Immunofluorescence, polyomavirus nephritis, 194 Immunohistochemistry, polyomavirus nephritis, 194 Immunological tolerance, to allografts. See Regulatory immune cells. Immunology - complement, 20–23 in atypical hemolytic uremic syndrome, 20–21 atypical hemolytic uremic syndrome and transplant, 21–22 eculizumab and atypical hemolytic uremic syndrome, 22 genetic defects in atypical hemolytic uremic syndrome/thrombotic microangiopathy, 21 laboratory testing for diagnosis of atypical hemolytic uremic syndrome and monitoring of eculizumab, 22–23 regulation and dysregulation of alternate complement pathway, 21 - immune response, 4–7 B-cell immune responses, 6 defining, 4 immunosuppressive therapy in, 7 infection, tissue damage and immune response, 5–6 innate immune responses and, 4–5 manipulation of, 7 NF-kB and T cells in, 6–7 NK cells and, 6 - laboratory-based immune monitoring, 24–29 approaches to immune monitoring, 24–25 immune monitoring in organ transplantation, 24 immune response biomarkers, 25–26 infections in solid organ transplantation, 26–27 relevance and application of immune monitoring, 27 - NK cells, 14–19 biology, 14–16 education (tolerance) and regulation, 15–16 expression profile of mature populations, 17 memory, 15 NKT cells (natural killer T cells) in solid organ transplantation, 16 xviii

in solid organ transplantation, 16 - regulatory immune cells and transplant tolerance, 8–13 Immunosuppression - adverse effects, pancreas transplantation associated, 474 - complications, vascularized composite allotransplantation associated, 513 - decreased, chronic rejection associated, 523 - NK cells, 16 - posttransplant lymphoproliferative disease associated, 527 Immunosuppressive drugs, 48–49 - antibody therapies, 54–55 approved for other indications, 55 approved for use in transplantation, 54–55 - anti-interleukin-2 receptor antibodies, 49 - antimetabolites, 49. See also Azathioprine (Aza); Mycophenolate mofetil (MMF); Mycophenolic acid (MPA). - antithymocyte globulin, 49 - calcineurin inhibitors, 48. See also Cyclosporine (CsA); Tacrolimus (FK506). - chronology of discovery, 55 - corticosteroids, 48 - drugs in clinical use (1962-1994), 54 - experimental drugs, 49 bortezomib, 49 eculizumab, 49 IgG endopeptidase, 49 intravenous immunoglobulin, 49 rituximab, 49 - history of, 54–55 - mammalian target of rapamycin inhibitors, 48–49. See also Everolimus; Sirolimus (rapamycin). - mechanism of action of, 48–49 - modern era (1995-present), 54 - modulation of immune response, 7 - overview, 54 - prevention and treatment of rejection, 36–37 - T-cell costimulation blockade, 49 - therapeutic drug monitoring, 50–53 azathioprine, 52 basiliximab, 52 clinical issues, 50 cyclosporine (CsA), 50–51 daclizumab, 52 etiology/pathogenesis, 50 everolimus, 51 monoclonal antibodies, 52 mycophenolate mofetil (MMF, RS-61443), 51–52 sirolimus (rapamycin), 51 tacrolimus (FK506), 51 terminology, 50 therapeutic ranges for trough concentrations, 53 Immunosuppressive protocols, pancreas transplantation, 480 Immunotactoid glomerulopathy, cytomegalovirus (CMV) infection vs., 206 In situ hybridization, cytomegalovirus (CMV) infection, 206

INDEX Inborn errors of metabolism, end-stage heart disease related to, 337 Inclusions, herpes simplex virus associated, 462 Infants of diabetic mothers, hypertrophic cardiomyopathy vs., 324 Infarction - hemorrhagic, pancreas transplantation associated, 484 - ischemic, pancreas transplantation associated, 484 - myocardial, myocarditis vs., 356 - right ventricle transmural, arrhythmogenic right ventricular cardiomyopathy vs., 334 Infections - acute cellular rejection grade B vs., 393 heart vs., 343 - acute T-cell- and antibody-mediated rejection vs., 516 - antibody-mediated rejection, lung vs., 389 - antibody-mediated rejection vs., 494 - chronic rejection vs., 523 - cytomegalovirus vs., 289 - infection, tissue damage and immune response, 5–6 - involving acinar tissue, recurrent diabetes mellitus vs., 500 - late allograft loss, 80 - microaspiration vs., 399 - pancreas transplantation associated, 475 - plasma cell-rich rejection vs., 303 - portal vein thrombosis vs., 253 - reperfusion injury vs., 419 - solid organ transplantation, 26–27 Infectious enteritis, 457 - acute antibody-mediated rejection, intestine vs., 426 Infectious granulomatous pyelonephritis, nocardiosis vs., 231 Infectious mononucleosis, 465 Infectious myocarditis, sarcoidosis vs., 331 Inflammation, donor organ, 243 Infliximab, history of use, 55 Insect bites, acute T-cell- and antibody-mediated rejection vs., 517 Insulin resistance, new-onset type 2 diabetes, pancreas transplantation associated, 474 Interacinar capillaries, inflammation, antibody-mediated rejection associated, 493 Interface dermatitis, acute T-cell- and antibody-mediated rejection associated, 516 Interstitial fibrosis - donor biopsy, 75 - and tubular atrophy, not otherwise specified, protocol biopsies, 91 Interstitial mononuclear inflammation, polyomavirus nephritis, 193 Interstitial nephritis - adenovirus, kidney, 201 - BK polyomavirus, acute T-cell-mediated rejection vs., 118 Interstitial pneumonia, usual, organizing pneumonia vs., 397 Interstitium - adenovirus, kidney, 201

- microsporidiosis, 223 Intestinal allograft diseases, pathologic classification, 412–413 - alloimmune response, 412 - multivisceral transplant issues, 413 - nonalloimmune diseases, 412–413 - recurrent intestinal diseases, 413 - retransplantation, 413 Intestinal atresia, explant, 414 Intestinal transplantation - acute antibody-mediated rejection, 424–427 diagnostic checklist, 426 differential diagnosis, 426 prognosis, 425 - acute cellular rejection, 428–435 diagnostic checklist, 430–431 differential diagnosis, 430, 431 histologic grading, 431 prognosis, 429 - adenovirus, 456–459 diagnostic checklist, 458 differential diagnosis, 458 prognosis, 457 serotypes, 457 - bacterial and fungal infections, 452–455 diagnostic checklist, 454 differential diagnosis, 454 infectious agents, 453 prognosis, 453 - chronic rejection, 436–439 diagnostic checklist, 437 differential diagnosis, 437 prognosis, 437 - colon rejection, 444–447 diagnostic checklist, 445 differential diagnosis, 445 prognosis, 445 - contraindications, 415 - Epstein-Barr virus, 464–469 characteristics, 466 differential diagnosis, 466 prognosis, 465 - graft survival following, 415 - graft-vs.-host disease, 448–451 - history of, 422–423 - indications and evaluation of explant, 414–417 - pathologic classification of intestinal allograft diseases, 412–413 alloimmune response, 412 multivisceral transplant issues, 413 nonalloimmune diseases, 412–413 recurrent intestinal diseases, 413 retransplantation, 413 - reperfusion injury, 418–421 diagnostic checklist, 419 differential diagnosis, 419 prognosis, 419 - rotavirus, cytomegalovirus, and herpes simplex virus, 460–463 diagnostic checklist, 462 differential diagnosis, 462 xix

INDEX prognosis, 461 - significant events, 423 - stomach rejection, 440–443 diagnostic checklist, 441 differential diagnosis, 441 grading, 441 - timeline, 422 - transplant landmarks, 422–423 - types, 412, 422 Intimal arteritis, acute cellular rejection associated, 489 Intraabdominal and opportunistic infections, pancreas, 504–509 - diagnostic checklist, 506–507 - differential diagnosis, 506–507 - modes, 505 - prognosis, 506 - sources, 505 Intranuclear inclusions, in tubular epithelium, polyomavirus nephritis, 193 Intravenous immunoglobulin, mechanism of action, 49 Invasive fungal infection, 301 Invasive Quilty type B (QB), 351 Invasive zygomycosis. See Mucormycosis. IPF. See Idiopathic pulmonary fibrosis. IRI. See Reperfusion injury. Irritant contact dermatitis, acute T-cell- and antibodymediated rejection vs., 517 Ischemia - acute allograft, 100–103 clinical issues, 101 diagnostic checklist, 101 differential diagnosis, 101 prognosis, 101 - due to technical issues, chronic rejection vs., 523 - herpes simplex virus vs., 291 - secondary, chronic rejection associated, 523 Ischemia/reperfusion injury. See Reperfusion injury. Ischemic cardiomyopathy. See also Ischemic heart disease. - sarcoidosis vs., 331 Ischemic cholangiopathy, chronic (ductopenic) rejection vs., 271 Ischemic heart disease, 314–317 - differential diagnosis, 315 - dilated cardiomyopathy vs., 320 - prognosis, 315 Ischemic hepatitis, hepatic artery thrombosis vs., 251 Ischemic infarction, pancreas transplantation associated, 484 Ischemic injury, 249. See also Preservation injury. - acute antibody-mediated rejection, intestine vs., 426 - adenovirus vs., 458 - hepatic artery thrombosis, 251 - perioperative, acute cellular rejection, heart vs., 343 Ischemic tubular injury, acute pyelonephritis vs., 190 ISHLT guidelines, pathologic classification of rejection, 386 Islet amyloid, 473 Islet amyloid deposition, 502–503 - pathogenic mechanisms, 503 - potential immunosuppressive agents, 503 - prognosis, 503 xx

Islet cell antibodies (ICAs), recurrent diabetes mellitus associated, 499 Islet cell damage, immune-mediated, pancreas transplantation associated, 474 Islet cell toxicity, 472, 502–503 - pathogenic mechanisms, 503 - potential immunosuppressive agents, 503 - prognosis, 503 Islet cell transplantation, 473 Islet cells, sources, 476 Islets, chronic allograft rejection/graft sclerosis associated, 497 Isolated atrial amyloidosis, 337 Isolated central perivenulitis, hepatic venous outflow obstruction vs., 258 Isolated endothelialitis, acute T-cell-mediated rejection vs., 118 Isolated small bowel transplant, intestinal transplants, 412 ITx. See Intestinal transplantation.

J Jejunum, native, adenovirus associated, 457 John Cunningham (JC) virus, 193 Jones methenamine silver, acute pyelonephritis, 190

K Keenan, Robert J., 367 Keshavjee, Shaf, 367 Kidney allograft, protocol biopsies, evaluation of, 70 Kidney diseases, in nonrenal transplant recipient, 174–177 - clinical issues, 175 - diagnostic checklist, 176 Kidney explant. See Transplant nephrectomy. Kidney transplantation - accommodation, 94–95 diagnostic checklist, 95 differential diagnosis, 95 prognosis, 95 - acute allograft ischemia, 100–103 clinical issues, 101 diagnostic checklist, 101 differential diagnosis, 101 prognosis, 101 - acute antibody-mediated rejection, 132–139 clinical issues, 133 diagnostic checklist, 134 differential diagnosis, 134 prognosis, 133 - acute pyelonephritis, 188–191 - acute T-cell-mediated rejection, 116–127 - adenovirus, 200–203

INDEX - antiglomerular basement membrane (anti-GBM) disease in Alport syndrome, 166–167 clinical issues, 167 diagnostic checklist, 167 differential diagnosis, 167 prognosis, 167 - aspergillosis, 216–217 - calcineurin inhibitor toxicity, 180–185 clinical issues, 181 diagnostic checklist, 182 differential diagnosis, 182 prognosis, 181 - candidiasis, 210–211 - chronic antibody-mediated rejection, 140–151 Banff 2017 criteria, 144 clinical issues, 141 diagnostic checklist, 143 differential diagnosis, 142–143 prognosis, 141 scoring, 144, 145 - chronic T-cell-mediated rejection, 128–131 categories, 130 clinical issues, 129 diagnostic checklist, 130 differential diagnosis, 130 prognosis, 129 - coccidioidomycosis, 218–219 - cryptococcosis, 212–213 - cytomegalovirus infection, 204–207 - de novo membranous glomerulonephritis, 162–165 clinical issues, 163 diagnostic checklist, 163 differential diagnosis, 163 prognosis, 163 - engraftment syndrome (ES), 170–173 clinical issues, 171 differential diagnosis, 171 prognosis, 171 - focal segmental glomerulosclerosis, de novo, 160–161 clinical issues, 161 diagnostic checklist, 161 differential diagnosis, 161 prognosis, 161 - graft-vs.-host glomerulopathies, 178–179 clinical issues, 179 differential diagnosis, 179 prognosis, 179 - histoplasmosis, 208–209 - history of, 58–59 alloantibody testing, 59 allograft tolerance, 59 chronology and evolution, 58 ex vivo perfusion, 59 laboratory testing, 58–59 xenotransplantation, 59 - HLA matching, 37 - hyperacute rejection, 112–115 clinical issues, 113 differential diagnosis, 113 prognosis, 113

- hyperperfusion injury, 168–173 clinical issues, 169 diagnostic checklist, 169 differential diagnosis, 169 prognosis, 169 - kidney diseases in nonrenal transplant recipient, 174–177 clinical issues, 175 diagnostic checklist, 176 - lymphocele, 106–107 clinical issues, 107 diagnostic checklist, 107 differential diagnosis, 107 prognosis, 107 - lymphocele, urine leak vs., 105 - malakoplakia, 226–229 - microsporidiosis, 222–223 - mTOR inhibitor toxicity, 186–187 - mucormycosis, 214–215 - nocardiosis, 230–231 - paracoccidioidomycosis, 220–221 - pathologic classification of renal allograft diseases. See Renal allograft diseases, pathologic classification of. - polyomavirus nephritis, 192–199 - protocol biopsies, 90–93 - recurrent diseases in the allograft, 152–159 clinical issues, 153–154 diagnostic checklist, 155 differential diagnosis, 154–155 prognosis, 154 - renal artery or vein thrombosis, 108–109 clinical issues, 109 diagnostic checklist, 109 differential diagnosis, 109 prognosis, 109 - tolerance, 96–99 diagnostic checklist, 98 prognosis, 97 - transplant renal artery stenosis, 110–111 clinical issues, 111 diagnostic checklist, 111 differential diagnosis, 111 prognosis, 111 - tuberculosis, 224–225 - urine leak, 104–105 clinical issues, 105 differential diagnosis, 105 prognosis, 105 - urine leak, lymphocele vs., 107 Kidney transplantation evaluation - allograft kidney, 68–73 - donor kidney, 74–79 - end-stage kidney, 62–67 - fibrosis, 86–89 mechanisms, 86 methods for assessment, 86–87 value, 86 - transplant nephrectomy, 80–85 Klebsiella infections, 453

xxi

INDEX

L

Laboratory-based immune monitoring, 24–29 - approaches to immune monitoring, 24–25 global immune function analysis, 25 global immunophenotyping analysis by flow cytometry, 24 molecular methods, 25 - immune monitoring in organ transplantation, 24 - immune response biomarkers, 25–26 immune activation markers, 25 immune tolerance markers, 25 - infections in solid organ transplantation, 26–27 antigen-specific assessment of T-cell immune competence, 26–27 - kidney transplantation, history of, 58–59 - relevance and application of immune monitoring, 27 Lamina propria - cytomegalovirus infection associated, 461 - rotavirus infection associated, 461 LAMP-2 deficiency, hypertrophic cardiomyopathy vs., 324 Langerhans cell histiocytosis, other causes of end-stage lung disease related to, 381 Large bile duct obstruction. See Bile duct stricture. Large eosinophilic intranuclear inclusions, cytomegalovirus (CMV) infection, 206 Large intranuclear inclusions, cytomegalovirus (CMV) infection, 206 Late chronic (ductopenic) rejection, 234 Late graft failure, 240, 241 LB (Lymphocytic bronchiolitis). See Acute cellular rejection, grade B. Leopard syndrome, hypertrophic cardiomyopathy vs., 324 Lichen planus, acute T-cell- and antibody-mediated rejection vs., 517 Lichen striatus, acute T-cell- and antibody-mediated rejection vs., 517 Lichenoid dermatoses, acute T-cell- and antibodymediated rejection vs., 517 Lichenoid lupus erythematosus, acute T-cell- and antibodymediated rejection vs., 517 Light chain (primary systemic) amyloidosis, 337 Light chain (myeloma) cast nephropathy, mTOR inhibitor toxicity vs., 187 Lipoid pneumonia, other causes of end-stage lung disease vs., 382 L-ITx. See Liver and intestinal transplantation. Liver allograft diseases, pathologic classification of, 234–235 - alloimmune response, 234 - infections, 235 - nonalloimmune diseases, 234 - recurrent diseases, 235 Liver and intestinal transplantation, 414 Liver injury, drug-induced, graft-vs.-host disease vs., 305 Liver transplantation - adenovirus, 292–293 diagnostic checklist, 293 xxii

-

-

-

-

-

-

-

-

-

-

-

-

-

-

differential diagnosis, 293 prognosis, 293 antibody-mediated rejection, 266–269 differential diagnosis, 267 prognosis, 267 bile duct stricture, leak, sludge, biloma, 254–255 chronic (ductopenic) rejection, 270–271 diagnostic checklist, 271 differential diagnosis, 271 prognosis, 271 cytomegalovirus, 288–289 diagnostic checklist, 289 differential diagnosis, 289 prognosis, 289 Epstein-Barr virus, 296–299 diagnostic checklist, 298 differential diagnosis, 298 prognosis, 297 evaluation of the donor liver, 242–245 of failed liver allograft, 240–241 fibrosing cholestatic hepatitis B or C virus, 278–279 diagnostic checklist, 279 differential diagnosis, 279 prognosis, 279 fungal infections, 300–301 diagnosis, 301 prognosis, 301 graft-vs.-host disease, 304–305 differential diagnosis, 305 prognosis, 305 gross evaluation of failed native liver, 236–239 hepatic artery thrombosis, 250–251 hepatic venous outflow obstruction, 256–259 differential diagnosis, 258 prognosis, 257–258 hepatitis E virus, 294–295 diagnostic checklist, 295 differential diagnosis, 295 prognosis, 295 herpes simplex virus, 290–291 diagnostic checklist, 291 differential diagnosis, 291 prognosis, 291 history, 246–247 hyperperfusion syndrome, 260–261 differential diagnosis, 261 prognosis, 261 indications, 236 major milestone in, 247 pathologic classification of liver allograft diseases, 234–235 plasma cell-rich rejection, 302–303 diagnostic checklist, 303 differential diagnosis, 303 prognosis, 303 portal vein thrombosis, 252–253 preservation injury, 248–249 hyperperfusion syndrome vs., 261 recurrent autoimmune hepatitis, 280–281 diagnostic checklist, 281

INDEX differential diagnosis, 281 prognosis, 281 - recurrent fatty liver disease, 286–287 differential diagnosis, 287 prognosis, 287 - recurrent hepatitis B virus, 272–273 diagnostic checklist, 273 differential diagnosis, 273 prognosis, 273 - recurrent hepatitis C virus, 274–277 diagnostic checklist, 276 differential diagnosis, 276 prognosis, 275 - recurrent primary biliary cirrhosis, 282–283 diagnostic checklist, 283 differential diagnosis, 283 prognosis, 283 - recurrent primary sclerosing cholangitis, 284–285 differential diagnosis, 285 prognosis, 285 - T-cell-mediated rejection, 262–265 differential diagnosis, 264 prognosis, 263 Lobar lung transplantation, 362 Lobular necrosis, pancreas transplantation associated, 484 Lung allograft - examination of native and transplanted lungs, 362, 364–365 diagnostic checklist, 365 prognosis, 365 - pathologic classification of lung allograft disease, 362–363 alloimmune response, 362 indications for lung transplantation, 363 nonalloimmune diseases, 362 - pathologic classification of lung allograft rejection, 386–387 Lung allograft rejection - antibody-mediated, 386 - pathologic classification of, 386–387 prognosis, 386 Lung disease - connective tissue disease-associated, 374–375 diagnostic checklist, 375 differential diagnosis, 375 idiopathic pulmonary fibrosis vs., 373 prognosis, 375 - end-stage, other causes, 380–383 diagnostic checklist, 382 differential diagnosis, 383 prognosis, 381 Lung failure, native lung, evaluation of - connective tissue disease-associated lung disease, 374–375 diagnostic checklist, 375 differential diagnosis, 375 idiopathic pulmonary fibrosis vs., 373 prognosis, 375 - cystic fibrosis, 370–371 diagnostic checklist, 371 differential diagnosis, 371

prognosis, 371 - emphysema, 368–369 diagnostic checklist, 369 differential diagnosis, 369 other causes of end-stage lung disease vs., 382 prognosis, 369 - idiopathic pulmonary fibrosis, 372–373 connective tissue disease-associated lung disease vs., 375 diagnostic checklist, 373 differential diagnosis, 373 prognosis, 373 - other causes of end-stage lung disease, 380–383 diagnostic checklist, 382 differential diagnosis, 383 prognosis, 381 - pulmonary arterial hypertension, 378–379 connective tissue disease-associated lung disease vs., 375 diagnostic checklist, 379 differential diagnosis, 379 prognosis, 379 - sarcoidosis, lung, 376–377 differential diagnosis, 377 prognosis, 377 Lung infections, graft-vs.-host disease vs., 450 Lung injury, acute, transfusion-related, 45 Lung transplantation - antibody-mediated rejection, 388–389 differential diagnosis, 389 prognosis, 389 - bacterial infections, 400–401 diagnostic checklist, 401 differential diagnosis, 401 prognosis, 401 - chronic allograft dysfunction, 394–395 diagnostic checklist, 395 prognosis, 395 - cystic fibrosis, 370–371 diagnostic checklist, 371 differential diagnosis, 371 prognosis, 371 - examination of native and transplanted lungs, 364–365 diagnostic checklist, 365 prognosis, 365 - fungal infections, 407–409 differential diagnosis, 407 prognosis, 407 - grade A acute cellular rejection, 390–391 diagnostic checklist, 391 differential diagnosis, 391 prognosis, 391 - grade B acute cellular rejection, 392–393 diagnostic checklist, 393 differential diagnosis, 393 prognosis, 393 - history of, 366–367 - HLA matching, 37 - idiopathic pulmonary fibrosis, 372–373 diagnostic checklist, 373 differential diagnosis, 373 xxiii

INDEX prognosis, 373 indications for, 363 landmark events in, 366 microaspiration, 398–399 organizing pneumonia, 396–397 other causes of end-stage lung disease, 380–383 diagnostic checklist, 382 differential diagnosis, 383 prognosis, 381 - pathologic classification of lung allograft diseases, 362–363 - pathologic classification of rejection, 386–387 prognosis, 386 - pulmonary arterial hypertension, 378–379 diagnostic checklist, 379 differential diagnosis, 379 prognosis, 379 - sarcoidosis, lung, 376–377 differential diagnosis, 377 prognosis, 377 - surgical aspects and complications, 384–385 acute graft failure, 385 anastomotic site, vascular insufficiency to, 385 anastomotic stenosis, 385 primary graft dysfunction, 385 prognosis, 385 thromboembolism, 385 - timeline of significant events in, 367 - viral infections, 402–405 diagnostic checklist, 404 genetic testing, 404 prognosis, 403 Lupus erythematosus, lichenoid, acute T-cell- and antibody-mediated rejection vs., 517 Lymphangioleiomyomatosis, other causes of end-stage lung disease related to, 381 Lymphocele, 106–107 - clinical issues, 107 - diagnostic checklist, 107 - differential diagnosis, 107 - prognosis, 107 - urine leak vs., 105 Lymphocytic bronchiolitis. See Acute cellular rejection, grade B. Lymphoma - acute T-cell- and antibody-mediated rejection vs., 517 - recurrent, graft-vs.-host glomerulopathies vs., 179 Lymphoproliferative disorder, posttransplant - acute T-cell-mediated rejection vs., 118 - colon rejection vs., 445 - T-cell-mediated rejection vs., 264 Lysosomal storage diseases, hypertrophic cardiomyopathy vs., 324 -

xxiv

M

Macroaspiration, microaspiration vs., 399 Macrophage markers, acute cellular rejection associated, 490 Macrovesicular steatosis, donor organ, 242–243 Major histocompatibility complex, genetics, 32 Major vascular thrombosis (renal artery or vein), hyperacute rejection vs., 113 Mal, Hervé, 366 Malacoplakia. See Malakoplakia. Malakoplakia, 226–229 - diagnostic checklist, 227 - differential diagnosis, 227 - prognosis, 227 Mammalian target of rapamycin (mTOR) drugs, 186 Mammalian target of rapamycin (mTOR) inhibitors - everolimus. See Everolimus. - mechanism of action, 48–49 - sirolimus. See Sirolimus (rapamycin). Maryland Aggregate Pathology Index (MAPI), donor kidney biopsy, 75 Medawar, Peter, 512 Megalocytic interstitial nephritis, malakoplakia vs., 227 Melanin incontinence, acute T-cell- and antibody-mediated rejection vs., 517 Membranous glomerulonephritis - de novo, 162–165 clinical issues, 163 diagnostic checklist, 163 differential diagnosis, 163 prognosis, 163 - donor-derived, de novo membranous glomerulonephritis vs., 163 - primary, graft-vs.-host glomerulopathies vs., 179 Merkel cell polyomavirus, 193 Mesangial proliferative glomerulonephritis, histoplasmosis, 209 Metabolic disorder - liver transplantation, 236 - native liver removal, 237 - vascularized composite allotransplantation associated, 513 Metabolic testing, hypertrophic cardiomyopathy, 324 Metabolism, inborn errors of, end-stage heart disease related to, 338 MGN. See Membranous glomerulonephritis, de novo. Microangiopathy - donor thrombotic, hyperacute rejection vs., 113 - thrombotic complement in, 20–23 genetic defects in, 21 graft-vs.-host glomerulopathies vs., 179 recurrent diseases in the allograft vs., 155 Microaspiration, 398–399 - bacterial infections, 401 - differential diagnosis, 399 - nonalloimmune diseases and, 362

INDEX - prognosis, 399 Microsporidia, 223 Microsporidiosis, 222–223 - differential diagnosis, 223 - prognosis, 223 Microvillus inclusion disease, explant, 415 Mild acute cellular rejection, 429–430 Minor histocompatibility antigens, 60 Mitochondrial cytopathies, hypertrophic cardiomyopathy vs., 324 MMVT. See Modified multivisceral transplantation. Model for End-Stage Liver Disease Score, 247 Moderate acute cellular rejection, 430 Modern era, liver transplantation, 246 Modified multivisceral transplantation, 414 Molecular mediators, fibrosis mechanism, 86 Monoclonal antibodies - mechanism of action, 49 - therapeutic monitoring, 52 Monomorphic posttransplant lymphoproliferative disease, 528, 530 - B-cell, 528 - T-cell and NK-cell, 528 Mononuclear cell infiltrate, acute cellular rejection associated, 489 Mononucleosis-type syndrome, posttransplant lymphoproliferative disease associated, 527 mTOR inhibitor toxicity, 61, 186–187 - differential diagnosis of, 187 Mucormycosis, 214–215 - aspergillosis vs., 217 - candidiasis vs., 211 - diagnostic checklist, 215 - differential diagnosis, 215 - prognosis, 215 Mucoviscidosis. See Cystic fibrosis. Multilamination, 60 Multiple necrotizing microabscesses with neutrophils, nocardiosis, 231 Multivisceral transplant, intestinal transplants, 412 Multivisceral transplantation, 414 - history of, 422–423 - significant event, 423 - timeline, 422 - transplant landmarks, 422–423 - types, 422–423 Muromonab-CD3 (Orthoclone OKT3), history of use, 54 Murray, Joseph E., 512 MVT. See Multivisceral transplantation. Mycobacterial infection, 453 - sarcoidosis of lung vs., 377 Mycobacterium avium-intracellulare, 225 - infection, tuberculosis vs., 225 Mycobacterium bovis, 225 Mycobacterium tuberculosis, 225 Mycophenolate, islet amyloid deposition and islet cell toxicity associated with, 503 Mycophenolate mofetil (MMF) - history of use, 54 - mechanism of action, 49 - therapeutic monitoring, 51–52

Mycophenolate mofetil colitis, colon rejection vs., 445 Mycophenolic acid (MPA) - history of use, 54 - mechanism of action, 49 - therapeutic ranges for trough concentrations, 53 Mycotic pseudoaneurysm, candidiasis, 211 Myocardial infarction, myocarditis vs., 356 Myocarditis, 354–359 - acute viral, myocarditis and, 356 - differential diagnosis, 356–357 - dilated cardiomyopathy vs., 320 - giant cell myocarditis vs., 356 sarcoidosis vs., 331 - infectious, sarcoidosis vs., 331 - infectious causes, 357 - noninfectious causes, 357 - prognosis, 355–356 Myocytolysis, 315

N

NAFLD. See Nonalcoholic fatty liver disease. NAMF. See Non-Aspergillus mycelial fungi. NASH. See Nonalcoholic steatohepatitis. Native lungs, examination of, 364–365 - diagnostic checklist, 365 - prognosis, 365 Native stomach, diseases of, stomach rejection vs., 441 Necrosis, drug-induced acute hepatitis with zone 3, hepatic venous outflow obstruction vs., 258 Necrotic nodules, coccidioidomycosis, 219 Necrotizing enterocolitis, explant, 414 Necrotizing glomerulonephritis, cryptococcosis, 213 Neoplasia - late allograft loss, 80 - nonalloimmune diseases and, 362 - pancreas allograft diseases, 473 Nephritis - acute allergic tubulointerstitial, acute T-cell-mediated rejection vs., 118 - BK polyomavirus interstitial, acute T-cell-mediated rejection vs., 118 Nephropathy - diabetic antiglomerular basement membrane (anti-GBM) disease in Alport syndrome vs., 167 in end-stage kidney, 64 - perfusion, hyperacute rejection vs., 113 - radiation, chronic T-cell-mediated rejection vs., 130 - subclinical BK polyomavirus, protocol biopsies, 91 Nephrosclerosis, hypertensive, in end-stage kidney, 64 Neutrophilic infiltrates, preservation injury, 249 Neutrophils, bacterial and fungal infections, 453 NF-kB, immune response in solid organ transplantation, 6–7 Nippostrongylus brasiliensis, 231

xxv

INDEX NK cells, 14–19 - biology, 14–16 - education (tolerance) and regulation, 15–16 - expression profile of mature populations, 17 - immune response, in solid organ transplantation, 6 - memory, 15 - NK-cell-mediated responses in rejection and tolerance, 6 - NKT cells (natural killer T cells) in solid organ transplantation, 16 - in solid organ transplantation, 16 NK-cell posttransplant lymphoproliferative disease, 527, 530 - monomorphic, 528 Nocardia, 231 Nocardia asteroides, 231 Nocardiosis, 230–231 - differential diagnosis, 231 - prognosis, 231 Nodules, Aschoff, myocarditis vs., 357 Nonalcoholic fatty liver disease, 287 Nonalcoholic steatohepatitis, 287 Nonalloimmune diseases, 362 - pancreas allograft diseases, 472–473 Non-Aspergillus mycelial fungi, lung infections, 407 Nondestructive posttransplant lymphoproliferative disease, 528, 530 Nonfungal pneumonia, fungal infections, lung vs., 407 Noninvasive Quilty type A (QA), 351 Nonischemic cardiomyopathy. See Dilated cardiomyopathy. Nonnocardial bacterial pyelonephritis, nocardiosis vs., 231 Nonrenal transplant recipient, kidney diseases in, 174–177 - clinical issues, 175 - diagnostic checklist, 176 Nonseptate fungal hyphae, mucormycosis, 215 Noonan syndrome, hypertrophic cardiomyopathy vs., 324

O

Obstruction - acute, acute allograft ischemia vs., 101 - acute T-cell-mediated rejection vs., 118 - transplant renal artery stenosis vs., 111 Ohio Valley disease. See Histoplasmosis. OP. See Organizing pneumonia. OP pattern. See Organizing pneumonia. Opportunistic infections, pancreas transplantationassociated, 475 Organ Procurement and Transplantation Network, 512 Organ transplantation, immune response, 4–7 - B-cell immune responses, 6 - defining, 4 - immunosuppressive therapy, 7 - infection, tissue damage and immune response, 5–6 - innate immune responses, 4–5 - manipulation, 7 - NF-kB and T cells, 6–7 xxvi

- NK cells, 6 Organizing diffuse alveolar damage, organizing pneumonia vs., 397 Organizing pneumonia, 396–397 - differential diagnosis, 397 - nonalloimmune diseases, 362 - stages, 397 Orthotopic liver transplantation, 246 Overinflation artifact from formalin perfusion, emphysema vs., 369 Özkan, Ömer, 513

P

Pancreas after kidney (PAK) transplantation, 474 - outcomes, 475 Pancreas allograft diseases - alloimmune responses, 472 - drug toxicity, 472 - nonalloimmune, 472–473 - pathologic classification, 472–473 Pancreas transplant alone (PTA), 474 - outcomes, 475 Pancreas transplantation - acute cellular rejection, 488–491 diagnostic checklist, 490 differential diagnosis, 490 prognosis, 489 - antibody-mediated rejection, 492–495 acute, 493 chronic active, 493 diagnostic checklist, 493 differential diagnosis, 494 hyperacute, 493 prognosis, 493 - benefits, 474 - biopsy, 480–481 diagnostic utility, 475 work-up, 476 - causes of chronic hyperglycemia after, 499 - challenges, 474–475 - chronic allograft rejection/graft sclerosis, 496–497 Banff grading, 497 diagnostic checklist, 497 differential diagnosis, 497 prognosis, 497 - clinical considerations, 474–479 - clinical implications of, 474 - complications, 482–487 clinical implications, 483 diagnostic checklist, 484 differential diagnosis, 484 preventive strategies and treatment, 483 prognosis, 483 risk factors, 483 - comprehensive histologic examination, 476 - definition, 480 - donor selection, 475

INDEX - dysfunction, 475 - history, 480–481 chronology and evolution, 480 progress in treatment, 480 - human leukocyte antigen matching, 475 - immunosuppressive protocols, 480 - impediments to, 480 - indications, 474 - intraabdominal and opportunistic infection, 504–509 diagnostic checklist, 506–507 differential diagnosis, 506–507 modes, 505 prognosis, 506 sources, 505 - islet cell toxicity and islet amyloid deposition, 502–503 pathogenic mechanisms, 503 potential immunosuppressive agents, 503 prognosis, 503 - outcome, 475 - recipient selection, 475 - recurrent diabetes mellitus animal models, 499 autoimmune mechanisms, 499 differential diagnosis, 500 prognosis, 499 - salvage techniques, 483 - surgical aspects, 482–487 - surgical techniques, evolution, 482 - types, 474 Pancreatic duct obstruction, intraabdominal and opportunistic infections vs., 506 Pancreatic dysfunction, pancreas transplantation associated, 475 Pancreatic homotransplantation. See Pancreas transplantation. Pancreatic transplantation. See Pancreas transplantation. Pancreaticoduodenal allograft. See Pancreas transplantation. Pancreatitis - acute infectious, 506 - acute ischemic, 472 - chronic, 506 allograft rejection/graft sclerosis vs., 497 - cytomegalovirus acute cellular rejection vs., 490 intraabdominal and opportunistic infections vs., 507 - differential diagnosis, 506 - duct obstruction, recurrent diabetes mellitus vs., 500 - fungal, 506 - graft, 482 mild ischemic, pancreas transplantation associated, 484 pancreas transplantation associated, 482 - infectious, pancreas transplantation associated, 484 - posttransplant ischemic, intraabdominal and opportunistic infections vs., 506 - reflux (aseptic), 473 Papillary necrosis - candidiasis, 211 - cryptococcosis, 213 Paracoccidioides brasiliensis, 221

Paracoccidioidomycosis, 220–221 - cryptococcosis vs., 213 - diagnostic checklist, 221 - differential diagnosis, 221 - prognosis, 221 Parvovirus B19, viral infections, 403 Pauci-immune (ANCA-associated) crescentic glomerulonephritis, antiglomerular basement membrane (anti-GBM) disease in Alport syndrome vs., 167 Pauci-immune glomerulonephritis, nocardiosis vs., 231 PBC. See Primary biliary cirrhosis. PCM. See Paracoccidioidomycosis. Peacock, Erle E., 512 Pediatric End-Stage Liver Disease Score, 247 Pediatric indications, common, lung transplantation and, 363 Pediatric transplant patients, adenovirus associated, 457 Perfusion nephropathy, hyperacute rejection vs., 113 Periodic acid-Schiff stains, nocardiosis, 231 Perioperative ischemic injury, acute cellular rejection, heart vs., 343 Peripancreatitis - acute cellular rejection vs., 490 - acute infectious, 472 - acute purulent, 506 - intraabdominal and opportunistic infections vs., 507 - pancreas transplantation associated, 484 Peripheral T-cell lymphoma, monomorphic T-/NK-cell PTLD, 528 Peritubular capillaritis, acute T-cell-mediated rejection vs., 118 Peritubular capillaropathy, chronic antibody-mediated rejection vs., 142 Peritubular capillary margination, chronic antibodymediated rejection vs., 142 Perivascular rejection. See Acute cellular rejection. PH. See Pulmonary hypertension. Pharyngitis, posttransplant lymphoproliferative disease associated, 527 Plasma cell hepatitis (de novo AIH), recurrent autoimmune hepatitis vs., 281 Plasma cell-rich rejection, 302–303 - diagnostic checklist, 303 - differential diagnosis, 303 - prognosis, 303 Pneumocystis infections, 453 Pneumocystis pneumonia, fungal infections, lung vs., 407 Pneumonia - aspiration, microaspiration vs., 399 - bacterial, fungal infections, lung vs., 407 - endogenous lipoid, postobstructive, microaspiration vs., 399 - eosinophilic, organizing pneumonia vs., 397 - exogenous lipoid, fungal infections, lung vs., 407 - lipoid, other causes of end-stage lung disease vs., 382 - nonfungal, fungal infections, lung vs., 407 - organizing, 396–397 - pneumocystis, fungal infections, lung vs., 407 - usual interstitial, organizing pneumonia vs., 397 xxvii

INDEX Pneumonitis - hypersensitivity, chronic idiopathic pulmonary fibrosis vs., 373 sarcoidosis of lung vs., 377 - microaspiration vs., 399 Podocyte, mTOR inhibitor effect, 187 Polymorphic posttransplant lymphoproliferative disease, 528, 530 Polyomavirus nephritis, 192–199 - adenovirus, kidney vs., 201 - classification, 194 - cytomegalovirus (CMV) infection vs., 206 - diagnostic checklist, 194 - differential diagnosis, 194 - prognosis, 193 Polyomavirus nephropathy. See also Polyomavirus nephritis. - subclinical BK, protocol biopsies, 91 Portal vein thrombosis, 234, 252–253 - diagnostic checklist, 253 - differential diagnosis, 253 - pancreas transplantation associated, 483 - prognosis, 253 Porter, K. A., 247 Postbiopsy perforation, site of previous biopsy vs., 353 Postoperative bleeding, pancreas transplantation associated, 483 Postsurgical biliary stricture, recurrent primary sclerosing cholangitis vs., 285 Posttransplant ischemic pancreatitis, intraabdominal and opportunistic infections vs., 506 Posttransplant lymphoproliferative disease (PTLD), 234–235, 506, 526–535 - acute cellular rejection, heart vs., 343 - acute cellular rejection, intestine vs., 430 - acute cellular rejection vs., 490 - acute T-cell- and antibody-mediated rejection vs., 517 - acute T-cell-mediated rejection vs., 118 - B-cell, 527, 530 monomorphic, 528 - classic Hodgkin lymphoma-type, 528–529, 530 - classification & features, 530 - colon rejection vs., 445 - diagnostic checklist, 529 - differential diagnosis, 529 - Epstein-Barr virus, 465, 527 - graft-vs.-host disease vs., 305 - immunosuppression, 527 - intraabdominal and opportunistic infections vs., 507 - monomorphic, 528, 530 - natural history, 527 - nondestructive, 528, 530 - polymorphic, 528, 530 - prognosis, 527–528 - reporting, 529 - T-cell and NK-cell, 527 monomorphic, 528 - T-cell-mediated rejection vs., 264 - WHO classification, 528

xxviii

Posttransplant lymphoproliferative disorder. See Posttransplant lymphoproliferative disease. Posttransplant neoplastic disorders, posttransplant lymphoproliferative disease, 526–535 - B-cell, 527, 530 monomorphic, 528 - classic Hodgkin lymphoma-type, 528–529, 530 - classification & features, 530 - diagnostic checklist, 529 - differential diagnosis, 529 - Epstein-Barr virus, 527 - immunosuppression, 527 - monomorphic, 528, 530 - natural history, 527 - nondestructive, 528, 530 - polymorphic, 528, 530 - prognosis, 527–528 - reporting, 529 - T-cell and NK-cell, 527 monomorphic, 528 - WHO classification, 528 Posttransplant plasma cell hepatitis. See Plasma cell-rich rejection. Posttransplant smooth muscle (spindle cell) tumor, Epstein-Barr virus, 465 Posttransplant tumors, intestinal allograft diseases, 413 Preservation injury, 234, 248–249. See also Reperfusion injury. - differential diagnosis, 249 - liver, hyperperfusion syndrome vs., 261 - prognosis, 249 Primary biliary cholangitis, recurrent, chronic (ductopenic) rejection vs., 271 Primary biliary cirrhosis, 235 - recurrent, 282–283 diagnostic checklist, 283 differential diagnosis, 283 prognosis, 283 Primary endocardial fibroelastosis, end-stage heart disease related to, 338 Primary graft dysfunction - acute cellular rejection, grade A vs., 391 - antibody-mediated rejection, lung vs., 389 - nonalloimmune diseases and, 362 Primary liver disease, recurrence of, failed liver allograft, 240 Primary nonfunction - early allograft loss, 80 - liver, hyperperfusion syndrome vs., 261 Primary oxalosis, early allograft loss, 80 Primary sclerosing cholangitis, 235 - liver removal, 237 - recurrent, 284–285 chronic (ductopenic) rejection vs., 271 differential diagnosis, 285 prognosis, 285 Prominent acute inflammation, paracoccidioidomycosis, 221 Protein kinase AMP-activated γ2 (PRKAG2) deficiency, hypertrophic cardiomyopathy vs., 324

INDEX Protocol biopsies, 90–93 - allograft kidney, 70 Proximal nephron, polyomavirus nephritis, 193 PSC. See Primary sclerosing cholangitis. Pseudallescheriasis - aspergillosis vs., 217 - candidiasis vs., 211 - mucormycosis vs., 215 Pseudolymphoma, cutaneous, acute T-cell- and antibodymediated rejection vs., 517 Pseudomembranous colitis, colon rejection vs., 445 Pseudomonas aeruginosa, pancreas transplantation associated, 505 Pseudoobstruction, explant, 415 Psychological factors, vascularized composite allotransplantation associated, 513 PTLD. See Posttransplant lymphoproliferative disease. Pulmonary alveolar proteinosis, other causes of end-stage lung disease related to, 381 Pulmonary arterial hypertension (PAH), 378–379 - connective tissue disease-associated lung disease vs., 375 - diagnostic checklist, 379 - differential diagnosis, 379 - prognosis, 379 Pulmonary fibrosis, idiopathic, 372–373 - connective tissue disease-associated lung disease vs., 375 - diagnostic checklist, 373 - differential diagnosis, 373 - prognosis, 373 Pulmonary hypertension (PH), 379 - secondary, pulmonary arterial hypertension vs., 379 Putative autoantibodies, recurrent diabetes mellitus, 499 PVN. See Polyomavirus nephritis. PVT. See Portal vein thrombosis. Pyelitis, candidiasis, 211 Pyelonephritis - acute, 188–191 acute antibody-mediated rejection vs., 134 diagnostic checklist, 190 differential diagnosis, 190 immunohistochemistry, 190 prognosis, 189 - acute T-cell-mediated rejection vs., 118 - chronic, chronic T-cell-mediated rejection vs., 130

Q QA (noninvasive Quilty type A), 351 QB (invasive Quilty type B), 351 Quilty effect, acute cellular rejection, heart vs., 343 Quilty lesions, 350–351 - diagnostic checklist, 351 - differential diagnosis, 351 - prognosis, 351 - site of previous biopsy vs., 353

R

Radiation nephropathy, chronic T-cell-mediated rejection vs., 130 Radiation pneumonitis, other causes of end-stage lung disease related to, 381 Rapamycin, 187 RAS. See Restrictive allograft syndrome. RAT. See Renal artery thrombosis. Reactive gastropathy, stomach rejection vs., 441 Reactive surface enterocytes, adenovirus, 457 Recurrent atypical hemolytic uremic syndrome - acute antibody-mediated rejection vs., 134 - hyperacute rejection vs., 113 Recurrent autoimmune diabetes mellitus, 473. See also Recurrent diabetes mellitus. Recurrent autoimmune hepatitis, 280–281 - diagnostic checklist, 281 - differential diagnosis, 281 - plasma cell-rich rejection vs., 303 - prognosis, 281 - T-cell-mediated rejection vs., 264 Recurrent autoimmune isletitis. See also Recurrent diabetes mellitus. - intraabdominal and opportunistic infections vs., 507 Recurrent diabetes mellitus, 498–501 - animal models, 499 - autoimmune mechanisms, 499 - differential diagnosis, 500 - intraabdominal and opportunistic infections vs., 507 - prognosis, 499 Recurrent diabetic insulitis. See Recurrent diabetes mellitus. Recurrent disease - early allograft loss, 80 - late allograft loss, 80 Recurrent fatty liver disease, 286–287 - differential diagnosis, 287 - prognosis, 287 Recurrent focal segmental glomerulosclerosis - de novo focal segmental glomerulosclerosis vs., 161 - hyperperfusion injury vs., 169 Recurrent glomerular disease, protocol biopsies, 91 Recurrent hepatitis B virus, 272–273 - diagnostic checklist, 273 - differential diagnosis, 273 - prognosis, 273 Recurrent hepatitis C virus, 274–277 - diagnostic checklist, 276 - differential diagnosis, 276 - Epstein-Barr virus, liver vs., 298 - prognosis, 275 Recurrent intestinal diseases, intestinal allograft diseases, 413 Recurrent lymphoma, graft-vs.-host glomerulopathies vs., 179 Recurrent membranous glomerulonephritis, de novo membranous glomerulonephritis vs., 163 xxix

INDEX Recurrent primary biliary cholangitis, chronic (ductopenic) rejection vs., 271 Recurrent primary biliary cirrhosis, 282–283 - diagnostic checklist, 283 - differential diagnosis, 283 - prognosis, 283 Recurrent primary disease, nonalloimmune diseases and, 362 Recurrent primary sclerosing cholangitis, 284–285 - bile duct stricture vs., 255 - bile leak vs., 255 - bile sludge vs., 255 - biloma vs., 255 - differential diagnosis, 285 - hepatic artery thrombosis vs., 251 - prognosis, 285 - (ductopenic) rejection vs., 271 Reflux (aseptic) pancreatitis, 473 Regulatory immune cells - adaptive immune components, 8 - innate immune components, 8 - mechanisms of tolerance in normal immunity, 8 - organ transplantation and immunity, 8 - transplant tolerance, 8–13 assessment of transplant tolerance, 9 biomarkers of, 9 induction of transplant tolerance in clinical practice, 10 operational, 8–9 regulatory immune cells, 9 rejection vs. graft acceptance/tolerance, 9 use of regulatory cell therapies, 10 Reitz, Bruce, 366 Rejection - differential diagnosis, 506 - vascularized composite allotransplantation-associated, 513 classification, 513 Renal allograft diseases, pathologic classification of, 60–61 - alloimmune responses, 60 antibody-mediated rejection, 60 T-cell-mediated rejection, 60 - anatomic complications, 61 major vessel disease, 61 pelvis/ureter, 61 - definitions, 60 - donor disease, 61 - drug toxicity and hypersensitivity, 61 antiviral tubular toxicity, 61 calcineurin inhibitor toxicity, 61 drug-associated acute interstitial nephritis, 61 mTOR inhibitor toxicity, 61 - infection, 61 bacterial and fungal infection, 61 viral infection, 61 - other diseases, 61 idiopathic, 61 neoplasia, 61 - recurrent and de novo diseases, 61 donor-specific de novo diseases, 61 xxx

recipient-specific de novo diseases, 61 recurrent primary disease, 61 - terminology, 60 Renal artery thrombosis, 108–109 - clinical issues, 109 - diagnostic checklist, 109 - differential diagnosis, 109 - prognosis, 109 Renal cell carcinoma - malakoplakia vs., 227 - polyomavirus nephritis, 193 Renal medulla, pathologic features of, 63 Renal vein thrombosis, 108–109 - clinical issues, 109 - diagnostic checklist, 109 - differential diagnosis, 109 - prognosis, 109 Reperfusion injury, 418–421. See also Preservation injury. - acute cellular rejection, intestine vs., 430 - adenovirus vs., 458 - colon rejection vs., 445 - diagnostic checklist, 419 - differential diagnosis, 419 - intestinal allograft diseases, 412 - prognosis, 419 - severe, acute antibody-mediated rejection, intestine vs., 426 Research era, liver transplantation, 246 Respiratory syncytial virus, viral infections, 403 Resting CD56, 14 Restrictive allograft syndrome, 395 Retransplantation - explant, 415 - intestinal allograft diseases, 413 Rhabdomyolysis, mTOR inhibitor toxicity vs., 187 Rheumatic fever, acute, myocarditis vs., 357 Right ventricle transmural infarction, arrhythmogenic right ventricular cardiomyopathy vs., 334 Right ventricular fat, normal, arrhythmogenic right ventricular cardiomyopathy vs., 334 Rituximab - history of use, 55 - mechanism of action, 49 Rotavirus - acute cellular rejection, intestine vs., 430 - adenovirus vs., 458 - bacterial and fungal infections vs., 454 - Epstein-Barr virus vs., 466 - intestine, 460–463 diagnostic checklist, 462 differential diagnosis, 462 prognosis, 461 RSV. See Respiratory syncytial virus. RVT. See Renal vein thrombosis.

INDEX

S

Sarcoidosis - heart, 330–331 differential diagnosis, 331 genetic background, 331 myocarditis vs., 357 prognosis, 331 - histoplasmosis vs., 209 - lung, 376–377 differential diagnosis, 377 prognosis, 377 - mucormycosis vs., 215 - tuberculosis vs., 225 Secondary amyloidosis, 337 Secondary ischemia, chronic rejection associated, 523 Segmental glomerulosclerosis - de novo focal, 160–161 clinical issues, 161 diagnostic checklist, 161 differential diagnosis, 161 prognosis, 161 - focal, calcineurin inhibitor toxicity vs., 182 - recurrent focal de novo focal segmental glomerulosclerosis vs., 161 hyperperfusion injury vs., 169 Senile systemic amyloidosis. See Wild-type transthyretin amyloidosis. Septa, chronic allograft rejection/graft sclerosis associated, 497 Septal edema, pancreas transplantation associated, 484 Serum HCV RNA organ donor, evaluation, 243 Severe acute cellular rejection, 430 - chronic rejection, intestine vs., 437 Severe acute tubular necrosis, mTOR inhibitor toxicity vs., 187 Severe recurrent hepatitis C virus infection, fibrosing cholestatic hepatitis B or C virus vs., 279 Severe reperfusion injury, acute antibody-mediated rejection, intestine vs., 426 Siemionow, Maria, 512–513 Simian virus 40 (SV40), 193 Simultaneous pancreas-kidney (SPK) transplantation, 474 - outcomes, 475 Single lung transplantation, 362 Sinusoidal dilatation, causes of, hepatic venous outflow obstruction vs., 258 Sinusoidal obstruction syndrome, hepatic venous outflow obstruction and, 257 Sirolimus (rapamycin), 187 - history of use, 54 - islet amyloid deposition and islet cell toxicity associated, 503 - mechanism of action, 48–49 - therapeutic monitoring, 51 - therapeutic ranges for trough concentrations, 53 Site of previous biopsy (SOPB), 352–353

Size syndrome, small for, recurrent primary sclerosing cholangitis vs., 285 Skin lesions - acute T-cell- and antibody-mediated rejection associated, 515 - graft-vs.-host disease vs., 450 Skin rash, herpes simplex virus associated, 461 Small bowel, adenovirus associated, 457 Small-for-size syndrome. See Hyperperfusion syndrome. Smooth muscle tumors, posttransplant lymphoproliferative disease vs., 529 Snover, D., 247 Solid organ transplantation (SOT) - NK cells in, 16 - NKT cells (natural killer T cells) in, 16 SOPB. See Site of previous biopsy. South American blastomycosis. See Paracoccidioidomycosis. Spindle cell neoplasms, posttransplant lymphoproliferative disease vs., 529 Spindle cell tumor, Epstein-Barr virus, 465 Spongiosis, acute T-cell- and antibody-mediated rejection associated, 516 Staphylococcus, bacterial and fungal infections, 453 Starnes, Vaughn, 366 Starzl, Thomas, 246 Steatohepatitis - de novo, fatty liver disease vs., 287 - liver transplantation, 236 Stem cell transplant, HLA matching, 38 Stomach rejection, 440–443 - diagnostic checklist, 441 - differential diagnosis, 441 - grading, 441 Stomal output, adenovirus, 457 Storage disease, hypertrophic cardiomyopathy vs., 324 Strome, Marshal, 512 Strong, Russell, 246 Stuart Welch, C., 246 Stump leak, anastomotic and duodenal, pancreas transplantation associated, 483 Subclinical acute T-cell-mediated rejection, protocol biopsies, 91 Subclinical BK polyomavirus nephropathy, protocol biopsies, 91 Surfactant dysfunction disorders, other causes of endstage lung disease related to, 381 Surgical aspects and complications, lung, 384–385 - acute graft failure, 385 - anastomotic site, vascular insufficiency to, 385 - anastomotic stenosis, 385 - primary graft dysfunction, 385 - prognosis, 385 - thromboembolism, 385 Surgical complications, pancreas allograft diseases, 473 Surgical hepatitis, preservation injury vs., 249 Surveillance biopsies. See Protocol biopsies. Sustained virologic response, 275 SVR. See Sustained virologic response.

xxxi

INDEX

T

Tacrolimus (FK506) - history of use, 54 - islet amyloid deposition and islet cell toxicity associated, 503 - mechanism of action, 48 - therapeutic monitoring, 51 - therapeutic ranges for trough concentrations, 53 TB. See Tuberculosis. T-cell immune competence, antigen-specific assessment of, 26–27 - BK virus, 27 - cytomegalovirus, 26 - Epstein-Barr virus (EBV), 26–27 T-cell marker (CD3), acute cellular rejection-associated, 490 T-cell posttransplant lymphoproliferative disease, 527, 530 - monomorphic, 528 T-cell-mediated injury, chronic allograft rejection/graft sclerosis-associated, 497 T-cell-mediated mechanisms, recurrent diabetes mellitus, 499 T-cell-mediated rejection, 60. See also Acute cellular rejection. - acute, 116–127 acute antibody-mediated rejection vs., 134 acute pyelonephritis vs., 190 adenovirus, kidney vs., 201 Banff criteria, 119 cytomegalovirus (CMV) infection vs., 206 diagnostic checklist, 118 differential diagnosis, 118 engraftment syndrome vs., 171 pancreas, antibody-mediated rejection vs., 494 pancreas transplantation-associated, 484 renal artery or vein thrombosis vs., 109 vascularized composite allotransplantation, 514–521 - antibody-mediated rejection vs., 267 - borderline/suspicious, acute T-cell-mediated rejection vs., 118 - chronic, 128–131 categories, 130 clinical issues, 129 diagnostic checklist, 130 differential diagnosis, 130 prognosis, 129 - Epstein-Barr virus, liver vs., 298 - liver, 262–265 differential diagnosis, 264 prognosis, 263 - pancreas, 489 - plasma cell-rich rejection vs., 303 - recurrent autoimmune hepatitis vs., 281 - recurrent hepatitis B virus vs., 272–273 - recurrent hepatitis C virus vs., 276 - recurrent primary biliary cirrhosis vs., 283 xxxii

- resolving/partially treated, acute T-cell-mediated rejection vs., 118 - subclinical acute, protocol biopsies, 91 T cells - immune response in solid organ transplantation, 6–7 - regulatory, 8 TCMR. See T-cell-mediated rejection; T-cell-mediated rejection, acute. Technical surgical complications, failed liver allograft, 240 Thomas, E. Donnall, 512 Thrombi - in glomeruli and arteries, donor biopsy, 75 - in microvasculature, acute T-cell- and antibodymediated rejection associated, 516 Thromboembolic disease, pulmonary arterial hypertension vs., 379 Thromboembolism, as complication of lung transplantation, 385 Thrombosis - arterial, chronic rejection, intestine vs., 437 - aspergillosis, 217 - graft, pancreas, antibody-mediated rejection vs., 494 - major vascular(renal artery or vein), hyperacute rejection vs., 113 - vascular antibody-mediated rejection vs., 267 hyperperfusion syndrome vs., 261 Thrombotic microangiopathy - acute T-cell-mediated rejection vs., 118 - complement in, 20–23 - donor, hyperacute rejection vs., 113 - early allograft loss, 80 - engraftment syndrome vs., 171 - genetic defects in, 21 - graft-vs.-host glomerulopathies vs., 179 - histoplasmosis, 209 - recurrent diseases in the allograft vs., 155 "Time-zero" donor biopsy, 242 - evaluation, 243 Tissue damage, in immune response, in organ transplantation, 5–6 Tissue Gram stain (Brown-Brenn), acute pyelonephritis, 190 Tolerance, kidney transplantation, 96–99 - diagnostic checklist, 98 - prognosis, 97 Tonsillitis, posttransplant lymphoproliferative disease associated, 527 Toxic tubular injury, acute pyelonephritis vs., 190 Toxoplasmosis, microsporidiosis vs., 223 Tracheobronchitis, microaspiration vs., 399 Transfusion reactions, human leukocyte antigen-related immunological, 44–45 - donor human leukocyte antigen related, 44–45 - febrile nonhemolytic transfusion reactions, 44 - platelet refractoriness, 44 - recipient human leukocyte antigen related, 44 - transfusion-associated graft-vs.-host disease, 45 - transfusion-related acute lung injury, 45

INDEX Transient arteriopathy, acute, acute T-cell-mediated rejection vs., 118 Transmural inflammation, acute cellular rejection associated, 489 Transplant arteriopathy, chronic antibody-mediated rejection vs., 142 Transplant glomerulitis, 60 - acute, recurrent disease in the allograft vs., 155 Transplant glomerulopathy, 60 - chronic, de novo focal segmental glomerulosclerosis vs., 161 - chronic antibody-mediated rejection vs., 142 - protocol biopsies, 91 Transplant nephrectomy, evaluation of, 80–85 - clinical implications, 81 clinical presentation, 81 clinicopathologic correlation, 81 - etiology/pathogenesis, 80 causes of early allograft loss, 80 causes of late allograft loss, 80 - macroscopic, 81 anatomic features, 81 general features, 81 specimen handling, 81 - microscopic, 81 ancillary studies, 81 general features, 81 - terminology, 80–85 Transplant renal artery stenosis, 110–111 - clinical issues, 111 - diagnostic checklist, 111 - differential diagnosis, 111 - prognosis, 111 Transplant tolerance, 8–13 - assessment of transplant tolerance, 9 - biomarkers of, 9 - induction of transplant tolerance in clinical practice, 10 - operational, 8–9 - regulatory immune cells, 9 - rejection vs. graft acceptance/tolerance, 9 - use of regulatory cell therapies, 10 Transplantation, common indications, 310 Transplantation and HLA, 36–39 - anti-HLA antibodies, 38 - HLA antigens and matching resolution level, 39 - HLA matching, 37–38 - immune responses to allografts, 36–37 - methods to detect anti-HLA antibodies, 39 - types of transplants, 36 Transplanted hearts, explant, 311 Transplanted lungs, 362 - examination of, 364–365 diagnostic checklist, 365 prognosis, 365 Trulock, Elbert, 366 Tuberculosis, 224–225 - aspergillosis vs., 217 - cryptococcosis vs., 213 - differential diagnosis, 225 - histoplasmosis vs., 209 - mucormycosis vs., 215

- paracoccidioidomycosis vs., 221 - prognosis, 225 Tubular atrophy, donor biopsy, 75 Tubular injury, polyomavirus nephritis, 193 Tubular necrosis/injury, acute, acute antibody-mediated rejection vs., 134 Tubules - adenovirus, kidney, 201 - microsporidiosis, 223 Tubulitis, 60 - polyomavirus nephritis, 193 Tubulointerstitial diseases, in end-stage kidney, 64 Tubulointerstitial nephritis, acute allergic, acute T-cellmediated rejection vs., 118 Tubulopathy, calcineurin inhibitor toxicity vs., 182 Tufting enteropathy, explant, 415 Tumors, explant, 415 Tyrosine phosphatase-like molecule (IA-2), antibodies against, recurrent diabetes mellitus associated, 499

U Upper urinary tract infection. See Acute pyelonephritis. Ureteral ischemic injury, 105 Urinary obstruction, late allograft loss, 80 Urinary tract infection, postoperative, pancreas transplantation associated, 505 Urine leak - kidney transplantation, 104–105 clinical issues, 105 differential diagnosis, 105 prognosis, 105 - lymphocele vs., 107 Usual interstitial pneumonia (UIP). See also Idiopathic pulmonary fibrosis (IPF). - organizing pneumonia vs., 397

V Valley fever. See Coccidioidomycosis. Valvular disease, chronic, end-stage heart disease related to, 338 Valvular heart disease, end-stage heart disease related to, 337 Vanishing bile duct syndrome. See also Graft-vs.-host disease. - drug-induced, chronic (ductopenic) rejection vs., 271 Varicella-zoster virus - adenovirus vs., 293 - Epstein-Barr virus, liver vs., 298 - hepatitis E virus vs., 295 - viral infections, 403 Vascular disease - donor, chronic rejection vs., 523 - in end-stage kidney, 64 - late allograft loss, 80 xxxiii

INDEX - liver transplantation, 236 - native liver removal, 237 Vascular thrombosis - antibody-mediated rejection vs., 267 - hyperperfusion syndrome vs., 261 Vascularized composite allografts, 512 Vascularized composite allotransplantation - acute antibody-mediated rejection, 514–521 diagnostic checklist, 517 differential diagnosis, 516–517 gene expression, 516 prognosis, 515 - acute T-cell-mediated rejection, 514–521 Banff grading system, 515 criteria, 515 diagnostic checklist, 517 differential diagnosis, 516–517 gene expression, 516 prognosis, 515 - chronic rejection, 522–523 diagnostic checklist, 523 differential diagnosis, 523 prognosis, 523 - complications, 513 - history, 512–513 Vasculitis, nocardiosis vs., 231 Vasculopathy - calcineurin inhibitor toxicity vs., 182 - chronic allograft, heart, 348–349 diagnostic checklist, 349 differential diagnosis, 349 prognosis, 349 Veith, Frank J., 366 Venulitis, acute cellular rejection associated, 489 Vessels, microsporidiosis, 223 Villous blunting, rotavirus infection associated, 461 Viral colitis, non-EBV, colon rejection vs., 445 Viral cytopathic effect, adenovirus, kidney, 201 Viral eruptions, acute T-cell- and antibody-mediated rejection vs., 516 Viral hepatitis, chronic - (hepatitis B or C), T-cell-mediated rejection vs., 264 - recurrent autoimmune hepatitis vs., 281 Viral infections, 402–405 - acute cellular rejection, intestine vs., 430 - adenovirus vs., 458 - bacterial and fungal infections vs., 454 - chronic rejection, intestine vs., 437 - diagnostic checklist, 404 - Epstein-Barr virus vs., 466 - genetic testing, 404 - graft-vs.-host disease vs., 305 - intestinal allograft diseases, 413 - microaspiration vs., 399 - nonalloimmune diseases and, 362 - pancreas, opportunistic, 505 - pancreas allograft diseases, 472 - prognosis, 403 - recurrent diabetes mellitus vs., 500 - reperfusion injury vs., 419 Volvulus, explant, 414 xxxiv

VZV. See Varicella-zoster virus.

W

Wegener granulomatosis, myocarditis vs., 356 Whole-organ pancreas transplantation, 482 Wild-type transthyretin amyloidosis, 337 Wound dehiscence, bacterial and fungal infections, 453 Wound infection, pancreas transplantation, 505

X Xanthogranulomatous pyelonephritis, malakoplakia vs., 227 Xenotransplantation, kidney transplantation, history of, 59

Z Zone 3 ischemic injury, hepatic venous outflow obstruction vs., 258