Atlas of Basic Liver Histology for Practicing Clinicians and Pathologists [1 ed.] 9789819957613, 9789819957620

Table of contents :
Preface
Contents
About the Authors
1: Basic Liver Histology
1.1 Liver Cell Plate
1.2 Zones of Liver
1.3 Basic Cell Types
1.3.1 Hepatocytes
1.3.2 Cholangiocytes
1.3.3 Kupffer Cells (KC)
1.4 Limiting Plate
1.5 Portal Tract
1.6 Central Vein
1.7 Sinusoids
1.8 Biliary Canaliculi
1.9 Special Stains
1.9.1 Masson Trichome (MT) Stain
1.9.2 Reticulin Stain
1.9.3 PAS (Periodic Acid Schiff) Stain and PAS-D (Periodic Acid Schiff with Diastase) Stain
1.9.4 Orcein Stain
1.9.5 Rhodanine Stain
1.9.6 Perl’s Stain
2: Basic Liver Pathology Terminologies
2.1 Cholestasis
2.1.1 Bland Canalicular Cholestasis
2.1.2 Hepatocellular Cholestasis and Canalicular Cholestasis
2.2 Pigment Deposits
2.2.1 Dubin-Johnson Pigments
2.2.2 Iron Pigments
2.2.2.1 Iron in Ductal Epithelial Cells (Perls Stain)
2.2.2.2 Hepatocellular and Kupffer Cell Iron Overload (Grade 3 Siderosis)
2.2.3 Copper and Copper Associated Proteins
2.2.3.1 Rhodanine Stain for Copper Deposits
2.3 Portal Inflammation
2.3.1 Mild Portal Inflammation
2.3.1.1 Portal Inflammation with Ductular Reaction
2.3.2 Moderate Portal Inflammation
2.3.3 Severe Portal Inflammation and Interface Hepatitis
2.3.4 Interface Hepatitis and Focal Emperipolesis
2.3.5 Lobular Inflammation
2.3.6 Portal Inflammation with Predominant Lymphocytes
2.3.7 Portal Inflammation with Predominant Neutrophils
2.3.8 Portal Inflammation with Predominant Eosinophils
2.3.9 Lymphoplasmacytic Infiltration at Portal Tract
2.4 Bile Duct Pathology
2.4.1 Ductopenia (Absence of Duct in the Portal Tract)
2.4.1.1 Ductopenia in PAS
2.4.2 Periductal Lamellar Fibrosis
2.4.2.1 Periductal Lamellar Fibrosis (Masson Trichrome Stain)
2.4.3 Neutrophilic Ductulitis
2.4.4 Lymphocytic Ductulitis
2.5 Fatty Change in Liver Disease
2.5.1 Macrovesicular Fatty Change
2.5.2 Micro and Macrovesicular Fatty Change
2.6 Central Perivenulitis
2.7 Necrosis
2.7.1 Bridging Necrosis
2.7.2 Confluent Necrosis
2.7.3 Lobular Inflammation with Spotty Necrosis
2.7.4 Submassive to Massive Necrosis in Explant Liver
2.7.5 Submassive to Massive Necrosis in Core Needle Biopsy
2.8 Extramedullary Hematopoesis
2.9 Multinucleated Giant Cell
2.10 Ballooning Degeneration of Hepatocyte
2.11 Mallory Denk Bodies with Cholestasis
2.11.1 Mallory Denk Body in HCC
2.12 Adequacy of Liver Biopsy
3: Pediatric Liver Pathology
3.1 Neonatal Hepatitis Syndrome
3.1.1 Histology
3.2 Extra Hepatic Biliary Atresia
3.2.1 Histology
3.2.1.1 Salient Histological Features
3.2.1.2 Bile Duct Pictures for EHBA
3.3 Paucity of Intrahepatic Bile Ducts: Alagille Syndrome
3.3.1 Histology
3.3.1.1 Salient Histopathological Features
3.4 Ductal Plate Malformation Disorders
3.4.1 Congenital Hepatic Fibrosis (CHF)
3.4.1.1 Histology
Salient Histological Features
3.4.2 CAROLI’s Disease
3.4.2.1 Microscopy
3.5 Disorders of Bile Canalicular Transporters
3.5.1 Progressive Familial Intrahepatic Cholestasis (PFIC)
3.5.1.1 Histology
Progressive Familial Intrahepatic Cholestasis-1
3.5.2 Progressive Familial Intrahepatic Cholestasis-2 (BSEP, Bile Salt Exporter Pump Deficiency)
3.5.2.1 Salient Histological Features
3.5.3 Progressive Familial Intrahepatic Cholestasis (PFIC-3)
3.5.3.1 Salient Histological Features
3.6 Primary Disorders of Bile Acid Synthesis
3.6.1 Salient Histological Features
3.7 Hereditary Disorders of Bilirubin Metabolism: Dubin-Johnson Syndrome
3.7.1 Salient Histological Features
3.8 Disorders of Lipoprotein and Lipid Metabolism
3.8.1 Neimann Pick Disease (NPD)
3.8.2 Histology
3.8.2.1 Salient Histological Features
3.8.3 Cholesteryl Ester Storage Disorder (CESD)
3.8.4 Salient Histological Features
3.9 Disorders of Carbohydrate Metabolism: Glycogen Storage Disorder (GSD)
3.9.1 Histology
3.9.1.1 Disorder of Carbohydrate Metabolism
3.10 Disorder of Copper Metabolism: Wilson Disease (WD)
3.10.1 Histology
3.10.1.1 Salient Histological Features
3.11 Disorders of Iron Metabolism: Hereditary Hemochromatosis
3.11.1 Histology
3.11.1.1 Salient Histological Feature
Macroscopy
4: Autoimmune Diseases
4.1 Autoimmune Hepatitis (AIH)
4.1.1 Histology
4.1.1.1 Salient Histological Features
4.2 Primary Biliary Cholangitis
4.2.1 Histology
4.2.1.1 Salient Histological Features
4.2.1.2 Disorders Associated with Bile Duct Injury Other Than PBC
4.3 PSC and Overlap Syndrome
4.3.1 Histology
4.3.1.1 Salient Histological Features
4.3.1.2 p-ANCA (Perinuclear Antineutrophil Cytoplasmic Antibody)
4.3.1.3 Cytoplasmic (C) ANCA Positivity
4.3.1.4 Differences Between AIH and Viral Hepatitis
5: Fatty Liver Disease
5.1 Non-alcoholic Fatty Liver Disease (NAFLD)
5.1.1 Histology
5.1.1.1 Steatosis (Non-alcoholic Fatty Liver Disease, NAFLD)
5.1.1.2 Steatohepatitis (Non-alcoholic Steatohepatitis, NASH)
5.2 Alcoholic Liver Disease
5.2.1 Histology
5.2.1.1 Alcoholic Steatosis
5.2.1.2 Alcoholic Hepatitis
5.2.1.3 Severe Alcoholic Hepatitis and Alcoholic Cirrhosis
5.3 Alcoholic Cirrhosis
5.3.1 Gross Image
6: Viral Hepatitis
6.1 Acute Viral Hepatitis
6.1.1 Histology
6.2 Chronic Hepatitis
6.2.1 Hepatitis B Infection
6.2.2 Hepatitis C Infection
6.2.3 Chronic Hepatitis E
6.2.4 Histology of Chronic Hepatitis
6.2.5 Chronic Hepatitis Necroinflammatory and Fibrosis Scoring
7: Toxin Induced Hepatitis
7.1 Introduction
7.1.1 Patterns of Drug Induced Liver Injury
7.2 Steatohepatitis
7.2.1 Other Patterns of Drug Induced Liver Injury
7.3 Yellow Phosphorus Poisoning
7.3.1 Histology
7.3.1.1 Salient Histological Features
8: Vascular Liver Disease
8.1 Budd Chiari Syndrome (BCS)
8.1.1 Histology
9: Infiltrative Liver Disease
9.1 Sarcoidosis
9.1.1 Histology
9.2 Tuberculosis
9.2.1 Histology
10: Neoplasms of Liver
10.1 Regenerative Nodule
10.1.1 Histology
10.2 Focal Nodular Hyperplasia (FNH)
10.2.1 Histology
10.3 Benign Tumors
10.3.1 Hemangioma
10.3.1.1 Histology
10.3.2 Hepatic Adenoma
10.3.2.1 Histology
10.3.3 Bile Duct Adenoma
10.3.4 Biliary Cystadenoma
10.3.4.1 Histology
10.4 Malignant Tumors
10.4.1 Hepatocyte Origin
10.4.1.1 Hepatocellular Carcinoma (HCC)
Histology
Gross Image
10.4.1.2 Fibrolamellar Hepatocellular Carcinoma
Histology
10.4.1.3 Hepatoblastoma
Histology
10.4.2 Biliary Epithelial Origin
10.4.2.1 Cholangiocarcinoma
Histology
10.4.2.2 Mixed HCC/Cholangiocarcinoma
Histology
10.4.3 Mesenchymal Cell Origin
10.4.3.1 Hemangioendothelioma
Histology
10.4.3.2 Gastrointestinal Stromal Tumor
Histology
10.4.3.3 Metastatic Tumors: Colorectal, Breast
Histology
10.4.3.4 Neuroendocrine Tumors
Histology
11: Liver Transplant Pathology
11.1 Acute Cellular Rejection
11.1.1 Portal Inflammation
11.1.2 Ductulitis
11.1.3 Venous Endothelial Inflammation
11.1.3.1 Central Perivenulitis
11.1.3.2 Portal Venulitis: Inflammation of Portal Vein
11.2 Antibody Mediated Rejection
11.2.1 Portal Inflammation with Predominant Eosinophils
11.2.2 C4d Staining
11.3 Chronic Rejection (CR)
11.3.1 Histology
11.3.1.1 Salient Histological Features
11.4 Sepsis
11.4.1 Histology
11.4.1.1 Salient Histological Features
11.5 Small for Size Graft
11.5.1 Histology
11.5.1.1 Salient Histological Features
11.5.2 PRI (Preservation Reperfusion Injury)
11.5.2.1 Salient Histological Features
11.6 Complications Involving Other Organs in Liver Transplant Recipients
11.6.1 Cytomegalovirus (CMV) Ileitis
11.6.1.1 Salient Histological Features
11.6.2 Post-Transplant Lymphoproliferative Disorder
11.6.2.1 Salient Histological Features
12: Classical Histological Differences: Comparison Tables
12.1 Differentiation Between Neonatal Hepatitis and Biliary Atresia
12.2 Differentiation Between HAV and Autoimmune Hepatitis
12.3 Differentiating Features Between Granulomatous Hepatitis
12.4 Differences Between AIH and Viral Hepatitis
12.5 Chronic Hepatitis: Differential Diagnosis

Citation preview

Atlas of Basic Liver Histology for Practicing Clinicians and Pathologists K. S. Mouleeswaran Joy Varghese Mettu Srinivas Reddy

123

Atlas of Basic Liver Histology for Practicing Clinicians and Pathologists

K. S. Mouleeswaran • Joy Varghese Mettu Srinivas Reddy

Atlas of Basic Liver Histology for Practicing Clinicians and Pathologists

K. S. Mouleeswaran Department of Histopathology Gleneagles Global Health City Chennai, Tamil Nadu, India Mettu Srinivas Reddy Department of HPB and Liver Transplantation Gleneagles Global Health City Chennai, Tamil Nadu, India

Joy Varghese Department of Hepatology & Transplant Hepatology Gleneagles Global Health City Chennai, Tamil Nadu, India

ISBN 978-981-99-5761-3    ISBN 978-981-99-5762-0 (eBook) https://doi.org/10.1007/978-981-99-5762-0 © The Editor(s) (if applicable) and The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 This work is subject to copyright. All rights are solely and exclusively licensed by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. The publisher, the authors, and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, expressed or implied, with respect to the material contained herein or for any errors or omissions that may have been made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. This Springer imprint is published by the registered company Springer Nature Singapore Pte Ltd. The registered company address is: 152 Beach Road, #21-01/04 Gateway East, Singapore 189721, Singapore Paper in this product is recyclable.

Preface

The practice of liver transplantation has grown exponentially in India. Annually, more than 1500 liver transplants are being performed in over a hundred centers, and these numbers will continue to increase. There is increasing interest among liver clinicians and general pathologists to become conversant with liver pathology as a specialist field. While there are several excellent reference books concerning the subject, most of these are aimed at the experienced liver pathologist and are not appropriate for day-to-day usage. This present atlas is designed to fill that gap. It is arranged in a format which is logical, easily navigable, and concise in description yet with enough detail to guide the reader towards an appropriate diagnosis. The atlas is arranged in three sections to enable readers to gain incremental knowledge in this area. The first section explores normal liver histology and staining techniques commonly used in studying liver biopsies. The second section explores individual histological findings reported in liver biopsies with brief explanations regarding underlying pathogenesis. Finally, the third section looks at disease-specific findings in pediatric and adult liver diseases and in liver transplantation. The liver responds to injury in a limited number of ways leading to significant overlap in histological findings. Pathological diagnosis must consider not just the histological findings but also the clinical setting and relevant laboratory reports. This third section is organized such that the clinical presentation, laboratory workup, and histopathology findings of each clinical entity are presented together thus enabling the reader to appreciate the correlation between these three pillars of pathological diagnosis. Finally, the comprehensive index will help cross-referencing and serve as a ready reckoner for specific clinical diagnoses. All images in this atlas have been sourced from the Department of Histopathology at Gleneagles Global Health City, Chennai, from patients treated in the unit over the last 5 years. We would like to sincerely acknowledge the invaluable work done by Dr. Dinu Abirami Premkumar and Dr. Mohammed Fawaz N (Registrars in Hepatology), Dr. J.  John Vergilin (Consultant Histopathology), and our technical staff for their assistance in choosing cases, preparing the sections, and collating clinical and laboratory data to bring this work to fruition. Finally, we will forever owe our immense gratitude to our patients who teach us something new every single day.

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Preface

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We sincerely hope this atlas will help readers become more confident in understanding liver pathology reports and help stimulate an interest in the art of pathological diagnosis. Kind regards, Chennai, India Chennai, India  Chennai, India 

K. S. Mouleeswaran Joy Varghese Mettu Srinivas Reddy

Contents

1 Basic Liver Histology ����������������������������������������������������������������������   1 1.1 Liver Cell Plate ������������������������������������������������������������������������   1 1.2 Zones of Liver��������������������������������������������������������������������������   2 1.3 Basic Cell Types������������������������������������������������������������������������   3 1.3.1 Hepatocytes������������������������������������������������������������������   3 1.3.2 Cholangiocytes��������������������������������������������������������������   3 1.3.3 Kupffer Cells (KC)��������������������������������������������������������   4 1.4 Limiting Plate���������������������������������������������������������������������������   4 1.5 Portal Tract��������������������������������������������������������������������������������   4 1.6 Central Vein������������������������������������������������������������������������������   5 1.7 Sinusoids ����������������������������������������������������������������������������������   5 1.8 Biliary Canaliculi����������������������������������������������������������������������   5 1.9 Special Stains����������������������������������������������������������������������������   6 1.9.1 Masson Trichome (MT) Stain��������������������������������������   6 1.9.2 Reticulin Stain��������������������������������������������������������������   7 1.9.3 PAS (Periodic Acid Schiff) Stain and PAS-D (Periodic Acid Schiff with Diastase) Stain ������������������   8 1.9.4 Orcein Stain������������������������������������������������������������������   9 1.9.5 Rhodanine Stain������������������������������������������������������������   9 1.9.6 Perl’s Stain��������������������������������������������������������������������  10 2 Basic Liver Pathology Terminologies ��������������������������������������������  11 2.1 Cholestasis��������������������������������������������������������������������������������  11 2.1.1 Bland Canalicular Cholestasis��������������������������������������  11 2.1.2 Hepatocellular Cholestasis and Canalicular Cholestasis��������������������������������������������������������������������  11 2.2 Pigment Deposits����������������������������������������������������������������������  12 2.2.1 Dubin-Johnson Pigments����������������������������������������������  12 2.2.2 Iron Pigments����������������������������������������������������������������  12 2.2.3 Copper and Copper Associated Proteins����������������������  13 2.3 Portal Inflammation������������������������������������������������������������������  14 2.3.1 Mild Portal Inflammation����������������������������������������������  14 2.3.2 Moderate Portal Inflammation��������������������������������������  15 2.3.3 Severe Portal Inflammation and Interface Hepatitis������������������������������������������������������������������������  15 2.3.4 Interface Hepatitis and Focal Emperipolesis����������������  15 2.3.5 Lobular Inflammation ��������������������������������������������������  16 vii

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2.3.6 Portal Inflammation with Predominant Lymphocytes��  16 2.3.7 Portal Inflammation with Predominant Neutrophils ����  16 2.3.8 Portal Inflammation with Predominant Eosinophils����  17 2.3.9 Lymphoplasmacytic Infiltration at Portal Tract������������  17 2.4 Bile Duct Pathology������������������������������������������������������������������  18 2.4.1 Ductopenia (Absence of Duct in the Portal Tract)��������  18 2.4.2 Periductal Lamellar Fibrosis ����������������������������������������  18 2.4.3 Neutrophilic Ductulitis�������������������������������������������������  19 2.4.4 Lymphocytic Ductulitis������������������������������������������������  19 2.5 Fatty Change in Liver Disease��������������������������������������������������  19 2.5.1 Macrovesicular Fatty Change ��������������������������������������  20 2.5.2 Micro and Macrovesicular Fatty Change����������������������  20 2.6 Central Perivenulitis������������������������������������������������������������������  20 2.7 Necrosis������������������������������������������������������������������������������������  21 2.7.1 Bridging Necrosis ��������������������������������������������������������  21 2.7.2 Confluent Necrosis��������������������������������������������������������  21 2.7.3 Lobular Inflammation with Spotty Necrosis����������������  21 2.7.4 Submassive to Massive Necrosis in Explant Liver ������  21 2.7.5 Submassive to Massive Necrosis in Core Needle Biopsy��  22 2.8 Extramedullary Hematopoesis��������������������������������������������������  22 2.9 Multinucleated Giant Cell��������������������������������������������������������  22 2.10 Ballooning Degeneration of Hepatocyte����������������������������������  23 2.11 Mallory Denk Bodies with Cholestasis������������������������������������  23 2.11.1 Mallory Denk Body in HCC ����������������������������������������  23 2.12 Adequacy of Liver Biopsy��������������������������������������������������������  24 3 Pediatric Liver Pathology����������������������������������������������������������������  25 3.1 Neonatal Hepatitis Syndrome ��������������������������������������������������  25 3.1.1 Histology����������������������������������������������������������������������  25 3.2 Extra Hepatic Biliary Atresia����������������������������������������������������  26 3.2.1 Histology����������������������������������������������������������������������  27 3.3 Paucity of Intrahepatic Bile Ducts: Alagille Syndrome������������  28 3.3.1 Histology����������������������������������������������������������������������  29 3.4 Ductal Plate Malformation Disorders ��������������������������������������  30 3.4.1 Congenital Hepatic Fibrosis (CHF)������������������������������  30 3.4.2 CAROLI’s Disease��������������������������������������������������������  32 3.5 Disorders of Bile Canalicular Transporters������������������������������  33 3.5.1 Progressive Familial Intrahepatic Cholestasis (PFIC)��������������������������������������������������������  33 3.5.2 Progressive Familial Intrahepatic Cholestasis-2 (BSEP, Bile Salt Exporter Pump Deficiency) ��������������  34 3.5.3 Progressive Familial Intrahepatic Cholestasis (PFIC-3)����������������������������������������������������  35 3.6 Primary Disorders of Bile Acid Synthesis��������������������������������  36 3.6.1 Salient Histological Features����������������������������������������  36 3.7 Hereditary Disorders of Bilirubin Metabolism: Dubin-Johnson Syndrome��������������������������������������������������������  37

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3.7.1 Salient Histological Features����������������������������������������  38 3.8 Disorders of Lipoprotein and Lipid Metabolism����������������������  39 3.8.1 Neimann Pick Disease (NPD)��������������������������������������  39 3.8.2 Histology����������������������������������������������������������������������  39 3.8.3 Cholesteryl Ester Storage Disorder (CESD)����������������  40 3.9 Disorders of Carbohydrate Metabolism: Glycogen Storage Disorder (GSD)������������������������������������������������������������������������  41 3.9.1 Histology����������������������������������������������������������������������  41 3.10 Disorder of Copper Metabolism: Wilson Disease (WD)����������  42 3.10.1 Histology����������������������������������������������������������������������  43 3.11 Disorders of Iron Metabolism: Hereditary Hemochromatosis ��������������������������������������������������������������������  45 3.11.1 Histology����������������������������������������������������������������������  45 4 Autoimmune Diseases����������������������������������������������������������������������  47 4.1 Autoimmune Hepatitis (AIH) ��������������������������������������������������  47 4.1.1 Histology����������������������������������������������������������������������  47 4.2 Primary Biliary Cholangitis������������������������������������������������������  49 4.2.1 Histology����������������������������������������������������������������������  50 4.3 PSC and Overlap Syndrome ����������������������������������������������������  51 4.3.1 Histology����������������������������������������������������������������������  52 5 Fatty Liver Disease��������������������������������������������������������������������������  55 5.1 Non-alcoholic Fatty Liver Disease (NAFLD)��������������������������  55 5.1.1 Histology����������������������������������������������������������������������  56 5.2 Alcoholic Liver Disease������������������������������������������������������������  58 5.2.1 Histology����������������������������������������������������������������������  59 5.3 Alcoholic Cirrhosis ������������������������������������������������������������������  60 5.3.1 Gross Image������������������������������������������������������������������  60 6 Viral Hepatitis����������������������������������������������������������������������������������  63 6.1 Acute Viral Hepatitis����������������������������������������������������������������  63 6.1.1 Histology����������������������������������������������������������������������  63 6.2 Chronic Hepatitis����������������������������������������������������������������������  64 6.2.1 Hepatitis B Infection ����������������������������������������������������  64 6.2.2 Hepatitis C Infection ����������������������������������������������������  64 6.2.3 Chronic Hepatitis E������������������������������������������������������  65 6.2.4 Histology of Chronic Hepatitis ������������������������������������  65 6.2.5 Chronic Hepatitis Necroinflammatory and Fibrosis Scoring ����������������������������������������������������������  67 7 Toxin Induced Hepatitis������������������������������������������������������������������  69 7.1 Introduction������������������������������������������������������������������������������  69 7.1.1 Patterns of Drug Induced Liver Injury��������������������������  69 7.2 Steatohepatitis ��������������������������������������������������������������������������  70 7.2.1 Other Patterns of Drug Induced Liver Injury����������������  70 7.3 Yellow Phosphorus Poisoning��������������������������������������������������  70 7.3.1 Histology����������������������������������������������������������������������  71

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8 Vascular Liver Disease��������������������������������������������������������������������  73 8.1 Budd Chiari Syndrome (BCS)��������������������������������������������������  73 8.1.1 Histology����������������������������������������������������������������������  73 9 Infiltrative Liver Disease ����������������������������������������������������������������  75 9.1 Sarcoidosis��������������������������������������������������������������������������������  75 9.1.1 Histology����������������������������������������������������������������������  75 9.2 Tuberculosis������������������������������������������������������������������������������  76 9.2.1 Histology����������������������������������������������������������������������  76 10 Neoplasms of Liver��������������������������������������������������������������������������  79 10.1 Regenerative Nodule��������������������������������������������������������������  79 10.1.1 Histology��������������������������������������������������������������������  79 10.2 Focal Nodular Hyperplasia (FNH)������������������������������������������  80 10.2.1 Histology��������������������������������������������������������������������  80 10.3 Benign Tumors������������������������������������������������������������������������  82 10.3.1 Hemangioma ��������������������������������������������������������������  82 10.3.2 Hepatic Adenoma��������������������������������������������������������  82 10.3.3 Bile Duct Adenoma����������������������������������������������������  84 10.3.4 Biliary Cystadenoma��������������������������������������������������  85 10.4 Malignant Tumors������������������������������������������������������������������  86 10.4.1 Hepatocyte Origin������������������������������������������������������  86 10.4.2 Biliary Epithelial Origin ��������������������������������������������  91 10.4.3 Mesenchymal Cell Origin ������������������������������������������  94 11 Liver Transplant Pathology������������������������������������������������������������ 101 11.1 Acute Cellular Rejection�������������������������������������������������������� 101 11.1.1 Portal Inflammation���������������������������������������������������� 102 11.1.2 Ductulitis�������������������������������������������������������������������� 102 11.1.3 Venous Endothelial Inflammation������������������������������ 102 11.2 Antibody Mediated Rejection ������������������������������������������������ 103 11.2.1 Portal Inflammation with Predominant Eosinophils������������������������������������������������������������������ 103 11.2.2 C4d Staining �������������������������������������������������������������� 104 11.3 Chronic Rejection (CR)���������������������������������������������������������� 105 11.3.1 Histology�������������������������������������������������������������������� 105 11.4 Sepsis�������������������������������������������������������������������������������������� 107 11.4.1 Histology�������������������������������������������������������������������� 107 11.5 Small for Size Graft���������������������������������������������������������������� 108 11.5.1 Histology�������������������������������������������������������������������� 108 11.5.2 PRI (Preservation Reperfusion Injury) ���������������������� 108 11.6 Complications Involving Other Organs in Liver Transplant Recipients�������������������������������������������������������������� 109 11.6.1 Cytomegalovirus (CMV) Ileitis���������������������������������� 109 11.6.2 Post-Transplant Lymphoproliferative Disorder���������� 110 12 Classical  Histological Differences: Comparison Tables���������������� 113 12.1 Differentiation Between Neonatal Hepatitis and Biliary Atresia ������������������������������������������������������������������������ 113

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12.2 Differentiation Between HAV and Autoimmune Hepatitis���������������������������������������������������������������������������������� 114 12.3 Differentiating Features Between Granulomatous Hepatitis���������������������������������������������������������������������������������� 114 12.4 Differences Between AIH and Viral Hepatitis������������������������ 114 12.5 Chronic Hepatitis: Differential Diagnosis������������������������������ 115

About the Authors

K. S. Mouleeswaran  completed his medical training in Stanley Medical college, Chennai, and training in pathology at the Post-graduate Institute of Medical Education and Research, Chandigarh, India. He is currently the head of histopathology at Gleneagles Global Health City, Chennai. He has 13 years of experience in Histopathology and has a special interest in pathology of the liver. He has been an invited speaker at national meetings in pathology and liver diseases. Joy  Varghese  is currently the Director of Hepatology & Transplant Hepatology at Gleneagles Global Health City, Chennai, India. He is one of the most senior transplant hepatologists in India, having served as the head of transplant hepatology for more than 15  years and having managed more than 2000 liver transplants, including 400 pediatric liver transplants. He completed his DM (Gastro) at Stanley Medical College hospital, Chennai. After gaining Transplant Hepatology experience from Essen University Hospital, Germany, he joined as Asst. Professor in the Department of Medical Gastroenterology at Amrita Institute of Medical Science & Research Institute. He has vast experience in the field of general hepatology and has delivered more than 1000 awareness talks on liver disease/liver transplantation in India and abroad. He has published more than 100 peer-­ reviewed papers in the field of liver diseases. He received a PhD degree from the Tamil Nadu Dr. MGR Medical University. He is an adjunct professor in the Department of Hepatology at SRMC, Chennai. Mettu Srinivas Reddy  completed his medical training and general surgical training at JIPMER, Pondicherry, and Post-graduate Institute of Medical Education and Research, Chandigarh, in India. He obtained his PhD by research and completed his training in hepatobiliary surgery and liver transplantation in the UK. He has been involved in liver transplantation and liver surgery for over 15  years and is currently the director of liver transplantation and hepatobiliary surgery at Gleneagles Global Hospital, Chennai. He has over 100 peer-reviewed publications and is a regular invited speaker at national and international meetings.

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1

Basic Liver Histology

1.1

Liver Cell Plate

This image shows hepatic parenchyma with portal tract (PT) on the left side (marked by box and enlarged) and central vein (CV) on the right side

of image. Enlarged view of portal tract showing portal venule (blue arrow), hepatic arteriole (red arrow), and bile ductule (green arrow).

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 K. S. Mouleeswaran et al., Atlas of Basic Liver Histology for Practicing Clinicians and Pathologists, https://doi.org/10.1007/978-981-99-5762-0_1

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1.2 Zones of Liver

1  Basic Liver Histology

Zone 1: Periportal area surrounding the portal tract (marked by green circle). Zone 3: Perivenular area surrounding the central vein (marked by red circle). Zone 2: It is the area which comes between zone 1 and zone 3 (marked by arrow). Clinical importance: Direction of blood flow is usually from zone 1 to zone 3 (zone 3 more prone for ischemic injury), whereas biliary drainage is from zone 3 to zone 1.

1.3  Basic Cell Types

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1.3 Basic Cell Types

1.3.2 Cholangiocytes

1.3.1 Hepatocytes

• Cholangiocytes constitute 3–5% of the endogenous liver cell population, beginning with the canals of Hering, bile ductules, and then lining the intrahepatic and extrahepatic bile duct system. • Cholangiocytes modify the composition of bile during its transit through bile ducts by the secretion and absorption of water, electrolytes, and other organic solutes and almost 40% of basal bile flow is from ductal epithelium.

• Hepatocytes comprise 65% of the cells in the liver and 80% of hepatic volume. • They are polyhedral cells approximately 20–30 μm in diameter. • Hepatocytes are arranged as cords or sheets of cells radiating from the central venules and interlaced by sinusoidal spaces. • They are important for plasma protein synthesis, bile salt synthesis, bilirubin metabolism, drug metabolism and glucose metabolism. The following image shows hepatocytes (few are marked by arrow) arranged in sheets with sinusoidal spaces in between (few are marked by arrowhead).

The following image show hepatocytes (marked by arrow) arranged in sheets in between portal tract (PT) and central vein (CV).

This image shows hilar portal tract with biliary duct lined by cholangiocytes (marked by arrow). PV—Portal vein, BD—bile duct, Ar— hepatic artery, NB—nerve bundle.

This image shows bile ductules (BD) lined by cholangiocytes (marked by arrows). Cholangiocytes are low cuboidal epithelial cells with basally located nuclei (marked by arrowheads).

1  Basic Liver Histology

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1.3.3 Kupffer Cells (KC) • Kupffer cells are the resident macrophages in the liver. They comprise more than 80% of tissue macrophages in the body and 15% of cells in the liver. • Kupffer cells reside in the sinusoids anchored to endothelial cells or occasionally hepatocytes. • They are important in host response to various injuries including that induced by toxins and infectious agents. • PAS-Diastase resistant hypertrophied Kupffer cells will usually be seen in inherited metabolic/storage disorders/active hepatitis. This image shows hepatic lobule with single cell plate thick hepatocytes and two hypertrophied Kupffer cells (marked by arrowhead) with dot like basophilic nuclei and abundant dark eosinophilic cytoplasm. This bright eosinophilic cytoplasm contains partially PAS-Diastase resistant partially digested/undigested cellular material. Arrow highlights normal flat Kupffer cells lining the sinusoids.

1.4 Limiting Plate The area where the portal tract meets the hepatic parenchyma is the limiting plate, usually it is regular and circumscribed (marked by dotted line).

This image shows hilar portal tract—part of portal venule (PV), arteriole (Ar), bile ducts (BD), nerve bundle with adjoining liver parenchyma. Limiting plate marked by dotted line.

1.5 Portal Tract The following images show interlobular portal tract (PT in the center) containing • Portal venule (PV) with dilated lumen and single layer of flat endothelial lining. • Arteriole (Ar): Thick fibromuscular wall with round, regular small lumen. • Bile duct (BD): Single layer of cuboidal epithelium with basally located nuclei.

1.8  Biliary Canaliculi

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1.7 Sinusoids

1.6 Central Vein It is a terminal hepatic venule where the sinusoids drain. All the terminal hepatic venules drain into the branches of hepatic vein and finally into IVC through right and left hepatic veins. The following images show hepatic parenchyma (zone 3) with central vein (CV). The hepatocytes surrounding the central vein is called centrizonal/zone 3 hepatocytes.

• Sinusoids are blood filled vascular spaces interlaced in between the hepatocytes, lined by a single layer of endothelial cells. • The endothelial cells contain fenestra, which allow diffusion of macromolecules from the sinusoidal spaces into the hepatocytes. • Blood from the portal venules and hepatic arterioles flow into the sinusoids (zone 1–3) and drain into the central/terminal hepatics venules (refer schematic diagram). • Kupffer cells and pit cells (cytotoxic lymphoid cells) are located on the vascular surface of the sinusoids. This image shows part of central vein with adjoining hepatic parenchyma (zone 3)—hepatocytes in single cell plate thickness separated by sinusoidal spaces (marked by arrowhead). Few of sinusoidal spaces contain spindle shaped Kupffer cells, in close attachment to the hepatocytes (marked by arrow).

PAS - D stain

1.8 Biliary Canaliculi • Biliary canaliculi are formed by modification of the contact surface of the hepatocytes covered by microvilli. • The intralobular canalicular network drains into the canals of Herring, lined by single layer of low cuboidal epithelium (refer schematic diagram). • Bile flows from the hepatocyte into the adjoining biliary canaliculi, which in turn drains via the

1  Basic Liver Histology

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canals of Herring into short bile ductules to interlobular bile ducts of the adjacent portal tract. • The biliary canaliculi cannot be distinctly seen in routine staining techniques, because they are collapsed and juxtaposed in between the hepatocytes.

Features Collagen composition H&E stain

MT stain

This image shows hepatic parenchyma with biliary canaliculi highlighted as brown linear structures (uptake for MDR3 protein by the canaliculi) in between the hepatocytes.

Orcein stain

Reticulin stain

Pathological Fibroconnective tissue fibrosis Type V Type I and type III Not visualized in Visualized as routine staining eosinophilic material Does not highlight Highlights as blue or bluish green Does not highlight Elastin fibers of collagen are highlighted as black/brown Seen as golden Shows sinusoidal brown collagen connective tissue framework as black, fibers continuous, linear corrugated structures

• Fibrosis staging scoring: Phase F0 F1 F2 F3 F4

1.9 Special Stains 1.9.1 Masson Trichome (MT) Stain MT stain is tricolor stain where the hepatocytes appear red, whereas the fibrotic area appears blue and blue green. MT stain always highlights pathological fibrosis which could be either pericellular, perivenular, porto-portal, or porto-central fibrosis. So, MT stain is mainly used to assess the fibrosis location and staging of fibrosis. Differences between normal fibroconnective tissue and pathological fibrosis Features Constituents

Fibroconnective tissue Normal supporting network of hepatocytes including sinusoidal framework between the hepatocytes, in the portal tracts and bile ductules

Pathological characteristics No fibrosis Portal fibrosis without septa Portal fibrosis with few septa Numerous septa. Incomplete nodules Complete nodules: cirrhosis

The Batts–Ludwig scale is similar Metavir scoring. This scale runs from 0 to 4: 0: a lack of fibrosis, 1: portal fibrosis, 2: rare portal-to-portal septa, 3: fibrous septa, 4: definite or likely cirrhosis. Portal fibrosis with septal fibrous expansion of portal tract marked by arrow (Metavir or Batts Ludwig Score 2).

Pathological fibrosis Secondary to any chronic inflammation MT Stain

1.9  Special Stains

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MT Stain

MT Stain

Porto-portal bridging fibrosis images (Metavir F3, H&E, and MT stain)

In addition pericellular fibrosis (marked by arrowhead) around degenerating hepatocytes (marked by arrow) seen in alcoholic/non-­ alcoholic steatohepatitis and viral hepatitis. In non-alcoholic steatohepatitis, different scoring system is used which is NAS CRN Scoring system (discussed in Chap. 5).

MT Stain

MT Stain

1.9.2 Reticulin Stain

MT Stain

Metavir F4: Porto-portal bridging fibrosis (marked by arrow), portal tract (marked by star) with cirrhotic nodule (marked as N)

Reticulin stain highlights the single cell plate framework (which runs continuously around cell plate seen as dark brown black linear pattern covering hepatic cell plate marked by arrow) separated by sinusoids (marked by arrowhead).

1  Basic Liver Histology

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1.9.3 PAS (Periodic Acid Schiff) Stain and PAS-D (Periodic Acid Schiff with Diastase) Stain

Reticulin stain

Reticulin stain showing reticulin condensation/collapse. In liver cell necrosis, there will be loss of hepatocytes resulting in apposition of reticulin fibers (marked by arrowhead).

• PAS stain highlights glycogen content seen in cytoplasm of hepatocytes. PAS stain also highlights the portal macrophages, polymorphonuclear cells, and basement membrane of biliary ductules and ducts. However, mononuclear cells like lymphocytes and plasma cells will be PAS negative. • PAS-D stain shows complete bleaching of hepatocytes. Diastase usually removes glycogen from hepatocytes resulting in bleaching/ clearing of cytoplasm. • As PAS stain highlights basement membrane of biliary ductules, PAS stain will be of additional benefit for the assessment of presence/ absence of bile ducts in conditions presenting with ductopenia like Alagille syndrome, vanishing bile duct syndrome and ductopenic rejection. This image shows highlighted PAS positive glycogen content of the hepatocyte cytoplasm.

Reticulin stain

Fragmentation/loss of reticulin framework (marked by arrows) seen in hepatocellular carcinoma. This is a case of HCC which shows nucleomegaly and few prominent nucleoli with loss and fragmentation of reticulin framework. PAS Stain

This image shows complete clearing of cytoplasm of hepatocytes due to washout of glycogen content with use of diastase reagent (when compared to previous image). Also noted is part of portal tract (PT) with pink portal macrophages (marked by arrow).

Reticulin stain

1.9  Special Stains

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Orcein Stain

PAS - D stain

This image shows part of portal tract with PAS stain highlighting the basement membrane of biliary ductules (marked by arrowhead) and neutrophils (marked by circle) of the inflamed portal tracts. Whereas lymphocytes (marked by arrows) are PAS negative.

Highlights copper associated proteins seen as black dot granules in the hepatocytes (marked by arrowhead).

Orcein Stain

PAS Stain

1.9.4 Orcein Stain • Usually highlights copper associated proteins, elastin content of fibrosis, HBV surface antigen inclusions, internal elastic lamina of hepatic arteriole.

1.9.5 Rhodanine Stain Rhodanine stain highlights copper within the hepatocytes. Copper deposits are usually seen as coarse granular pigments in cytoplasm of hepatocytes (marked by arrows).

Highlights internal elastic lamina of hepatic arteriole as corrugated, linear, and black structure (marked by arrow). Not visualized in venule (marked by arrowhead).

Rhodanine Stain

1  Basic Liver Histology

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1.9.6 Perl’s Stain Perl’s stain highlights iron pigments which are seen as granular green pigments. Iron deposition occurs in the Kupffer cells, portal macrophages, hepatocytes, and occasionally in the bile duct epithelial cells, depending on the disease condition (marked as arrow).

Perl’s Stain

Image shows dense coarse green colored granular pigment in hepatocyte and Kupffer cells with

Perl’s stain suggestive of severe hepatocellular and Kupffer cell iron overload. In this picture, there is increased iron deposition seen as green granular pigments within the hypertrophied Kupffer cells (Kupffer cells are not visualized separately, interspersed between hepatocytes within the sinusoids) and perinuclear within the hepatocyte cytoplasm (marked by arrow).

Perl’s Stain

2

Basic Liver Pathology Terminologies

2.1

Cholestasis

2.1.1 Bland Canalicular Cholestasis Acellular brown pigment of bile inside canaliculus with no inflammation or necrosis is called bland cholestasis. This image shows mild bland canalicular cholestasis. In addition, hepatocytes have clear cytoplasm because of detergent effect of bile which is compactly packed as rosettes (marked by dotted lines).

Causes of bland canalicular cholestasis include: • Progressive familial intrahepatic cholestasis­1/BRIC. • Drug induced cholestasis (NSAIDs, corticosteroids, oral contraceptives, etc.).

2.1.2 Hepatocellular Cholestasis and Canalicular Cholestasis Brown coarse bile pigment deposition within the hepatocyte is known as hepatocellular cholestasis. This image shows moderate to severe hepatocellular (marked by arrow) and canalicular (marked by arrowhead) cholestasis. In addition, there is focal lobular inflammation containing lymphocytes and neutrophils (marked by dotted lines).

Causes of hepatocellular and canalicular cholestasis: • • • •

Viral hepatitis Drug induced hepatitis Severe autoimmune hepatitis Wilson’s disease and inherited metabolic disorders

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 K. S. Mouleeswaran et al., Atlas of Basic Liver Histology for Practicing Clinicians and Pathologists, https://doi.org/10.1007/978-981-99-5762-0_2

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2  Basic Liver Pathology Terminologies

• Progressive familial intrahepatic cholestasis-2 and PFIC-3 • Extrahepatic biliary atresia and neonatal hepatitis syndrome • Post-transplant: preservation reperfusion injury (PRI), sepsis, biliary complications, and antibody mediated rejection

2.2 Pigment Deposits 2.2.1 Dubin-Johnson Pigments Brown coarse granular pigments in cytoplasm of hepatocytes seen in DJ syndrome. DJ pigments are both PAS and Masson Fontana positive unlike lipofuscin pigments (which is PAS positive and Masson fontana negative) or bile pigments (which are both PAS and Masson fontana negative).

Masson Fontana positive in DJ syndrome (characteristic of melanin) seen as coarse granular black brown pigments within the hepatocyte cytoplasm.

2.2.2 Iron Pigments

PAS positive pigment in DJ syndrome (characteristic of lipofuscin) seen as dark pink, coarse granular pigments within the hepatocyte cytoplasm in the background of pale pink glycogen content of hepatocyte cytoplasm. Also visualized in this picture are dilated sinusoids with anuclear RBCs (left side of image— marked by dotted lines) and polymorphonuclear cells (marked by blue arrowhead) and Kupffer cells (marked by red arrowheads).

Iron pigments are seen in Kupffer cells (Perls stain). Iron pigments are not visualized in routine H&E stain like bile pigments and requires special stains like Perl’s stain. Excessive iron will get deposited usually in the Kupffer cells, portal macrophages, hepatocytes, and occasionally in the biliary epithelial cells as green curvilinear pigment form. This image shows minimal iron deposition within the Kupffer cells seen interspersed in between the hepatocytes (marked by arrows). In addition, image shows hepatocellular cholestasis as light, yellow granular bile pigments within the hepatocyte (arrowhead).

2.2  Pigment Deposits

Causes of iron overload in liver: • • • •

Primary hemochromatosis Alcoholic and non-alcoholic cirrhosis Chronic viral hepatitis Metabolic disorders like tyrosinemia, Gaucher disease • Hematological disorder like thalassemia and repeated transfusion

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2.2.2.2 Hepatocellular and Kupffer Cell Iron Overload (Grade 3 Siderosis) Image shows dense coarse green colored granular pigment (marked by arrows) in hepatocyte and Kupffer cells with Perls stain suggestive of severe hepatocellular and Kupffer cell iron overload.

Grading of siderosis based on the histology: Grade 0 siderosis: Granules absent or barely discernible at >400× Grade 1 siderosis: Granules barely discernible at 250× and easily confirmed at 250× Grade2 siderosis: Discrete granules resolved at 100× Grade 3 siderosis: Discrete granules resolved at 25× Grade 4 siderosis: Masses visible at 10× or naked eye

2.2.2.1 Iron in Ductal Epithelial Cells (Perls Stain) This image shows iron pigments within the ductal epithelial cells (marked by arrows), likely to be in primary hemochromatosis.

2.2.3 Copper and Copper Associated Proteins Copper deposition is not visualized by routine H&E staining, unlike bile pigments. Special stains like rhodanine and rubeanic acid highlight copper and orcein stain highlights copper associated proteins.

2  Basic Liver Pathology Terminologies

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This image shows black fine granular pigments within cytoplasm of hepatocytes that are copper associated proteins (marked by arrows).

Rhodanine stain image showing coarse red granular pigments (marked by arrow) in cytoplasm of hepatocytes which represents increased copper in the cytoplasm.

2.3 Portal Inflammation 2.3.1 Mild Portal Inflammation

Copper is excreted through bile ducts, hence copper in liver is increased in conditions related to bile duct pathology (starting from canaliculi to larger bile ducts): • • • • • • • •

This image shows portal tract (containing portal vein-PV, arteriole on the right side-Ar) with moderate inflammation containing mainly lymphocytes and polymorphonuclear cells and numerous interlobular bile ducts at interface (ductular reaction, marked by arrows). Ductular reaction is characterized by proliferation of reactive bile ducts (neocholangioles). In addition, it also shows focal spillage of portal inflammation (interface hepatitis, marked by star) into adjoining hepatocyte parenchyma (HP).

Wilson’s disease Autoimmune cholangitis Primary biliary cholangitis Primary sclerosing cholangitis Extrahepatic biliary atresia Progressive familial intrahepatic cholestasis Large duct obstructive cholangiopathy Drug induced cholestatic hepatitis

Note: In cirrhosis, copper is increased periportally, irrespective of etiology.

2.2.3.1 Rhodanine Stain for Copper Deposits

Note: Neocholangioles are formed due to metaplastic transformation of the injured hepatocytes at the interface/portal tract junction. (CK7 is a marker of bile duct epithelial cells. Weak CK7 uptake in the periportal hepatocyte indicates its metaplastic nature).

2.3.1.1 Portal Inflammation with Ductular Reaction IHC staining CK7 for bile duct highlighting the ductular reaction. CK7 highlights the cytoplasm of bile ductules as dark brown color, whereas the nucleus of bile ductules appears light blue (marked by arrows).

2.3  Portal Inflammation

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2.3.3 Severe Portal Inflammation and Interface Hepatitis This image shows part of portal tract (PT) with predominant lymphocytic infiltration and spillage of lymphocytes into the adjoining hepatic parenchyma, interface hepatitis (marked by star).

Causes of portal inflammation include: • • • • •

Viral hepatitis Autoimmune hepatitis Alcoholic hepatitis Drug induced hepatitis Acute cellular rejection in post-transplant liver

2.3.2 Moderate Portal Inflammation This image shows expanded portal tract (PT) containing moderate inflammatory cell infiltrate.

Causes of significant interface hepatitis include: • Autoimmune hepatitis/Wilson’s disease • Severe viral hepatitis • Severe acute cellular rejection

2.3.4 Interface Hepatitis and Focal Emperipolesis This image shows spillage of inflammatory cells into adjoining hepatic parenchyma called as interface hepatitis (marked as star). Emperipolesis is defined by presence of lymphocytes within the hepatocyte cytoplasm, appears as if lymphocytes are engulfed by the hepatocytes (marked by arrow).

Causes of interface hepatitis with emperipolesis

2  Basic Liver Pathology Terminologies

16

• Autoimmune hepatitis • Wilson’s disease

phocytic infiltrate. Lymphocytes are seen as mononuclear cells with clear cytoplasm and single round nuclei (few are marked by arrows).

2.3.5 Lobular Inflammation Lobular inflammation is collection or aggregates of lymphocytes/neutrophils/macrophages in the hepatic parenchyma usually associated with one or few hepatocytes loss. This image show collections of neutrophils with loss of hepatocyte (spotty necrosis, marked by dotted lines). In addition, it shows Mallory Denk bodies (arrowhead) in the surrounding hepatocytes in the right side corner. This is a case of alcoholic hepatitis (differential diagnosis— Wilson’s disease).

Causes of lymphocytic infiltration include: • • • • •

Viral hepatitis Autoimmune hepatitis Wilson’s disease Primary biliary cholangitis Acute cellular rejection

2.3.7 Portal Inflammation with Predominant Neutrophils

2.3.6 Portal Inflammation with Predominant Lymphocytes This picture shows part of portal tract containing central interlobular small bile duct with single layer of cuboidal lining (dysplastic duct of acute cellular rejection) and surrounding dense lym-

Image shows portal tract with inflammation containing predominantly polymorphonuclear cells (neutrophils, arrow) and numerous small bile ducts containing bile plugs (star) with destruction of ducts. There is spillage of inflammatory cells into adjoining parenchyma (interface hepatitis, arrowhead). Neutrophils are identified based on their multilobed nucleus (few are marked by arrows).

2.3  Portal Inflammation

17

Causes of portal inflammation with predominant eosinophils include:

Causes of portal inflammation with predominant neutrophils include: • • • • •

Sepsis Ascending cholangitis Large duct obstructive cholangiopathy Drug induced hepatitis Antibody mediated rejection and biliary complication in post-transplant liver

2.3.8 Portal Inflammation with Predominant Eosinophils Image shows portal tract with small portal venules (star) with single layer of endothelial lining and predominantly eosinophils (arrow). Eosinophils are seen with bilobed (spectacle shaped) nucleus and bright eosinophilic cytoplasm.

• Drug induced hepatitis • Parasitic infection • Moderate to severe acute cellular/acute antibody mediated rejection in post liver transplant

2.3.9 Lymphoplasmacytic Infiltration at Portal Tract Following image shows part of portal tract with mixture of lymphocytes and plasma cells. The plasma cells (marked by arrow) have eccentrically placed nucleus and moderate eosinophilic cytoplasm.

2  Basic Liver Pathology Terminologies

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Causes of lymphoplasmocytic infiltration include: • • • • •

Autoimmune hepatitis Wilson’s disease Drug induced hepatitis Acute cellular rejection Chronic hepatotropic viral hepatitis (commonly in HCV)

2.4 Bile Duct Pathology 2.4.1 Ductopenia (Absence of Duct in the Portal Tract)

2.4.2 Periductal Lamellar Fibrosis

This image shows portal tract in the center with portal venule (star) and arteriole (arrow), but no bile duct. Usually hepatic arteriole is accompanied by 1 or 2 bile ducts.

This image shows hilar portal tract (as nerve bundles (NB) are seen only in the hilar portal tract) with central small bile duct (BD) surrounding concentric layer of periductal lamellar fibrosis.

Causes of periductal lamellar fibrosis include: • Primary and secondary sclerosing cholangitis Note: Normal portal tract to duct ratio (0.9– 1.8), in ductopenia (0.0–0.4)

2.4.1.1 Ductopenia in PAS PAS stain highlights bile ducts if present (refer Chap. 1). This image shows portal tract with absent bile duct.

2.4.2.1 Periductal Lamellar Fibrosis (Masson Trichrome Stain) This shows part of portal tract with central small bile duct surrounding concentric layer of periductal lamellar fibrosis highlighted in bright blue color by Masson trichrome stain.

2.5  Fatty Change in Liver Disease

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2.4.4 Lymphocytic Ductulitis This image shows portal tract with small interlobular bile duct (tangential section, marked by dotted lines) with surrounding portal inflammation containing mainly lymphocytes. Bile duct is infiltrated by lymphocytes suggestive of lymphocytic ductulitis.

2.4.3 Neutrophilic Ductulitis High power image showing portal tract (marked by dotted line) in the lower half with surrounding hepatocytes. PT shows interlobular small bile duct with low cuboidal epithelium, infiltrated and destroyed by neutrophils (marked by arrows) suggestive of neutrophilic ductulitis. Preserved ductal epithelial cells show reactive atypia with nucleomegaly. In addition, this image shows bile duct lumen with thick bile plug.

Causes of lymphocytic ductulitis include: • Primary biliary cholangitis • Acute cellular rejection

2.5 Fatty Change in Liver Disease Steatosis or fatty change is seen in:

Causes of neutrophilic ductulitis include: • Acute cellular/antibody mediated rejection • Sepsis • Post liver transplant biliary stricture/ascending cholangitis

• • • • • •

Non-alcoholic fatty liver disease Alcoholic liver disease Drug induced Viral hepatitis (commonly in HCV) Wilson’s disease Occasionally PSC (primary sclerosing cholangitis) • Various inherited metabolic disorder

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2  Basic Liver Pathology Terminologies

2.5.1 Macrovesicular Fatty Change Macrovesicular steatosis: Large, single fat globule resulting in complete clearing of cytoplasm with pushing of nucleus toward periphery of the hepatocytes (few are marked by arrows).

2.6 Central Perivenulitis

2.5.2 Micro and Macrovesicular Fatty Change Microvesicular steatosis multiple small fat globules seen as clear cytoplasmic vesicles without displacement of centrally placed nucleus of hepatocytes.

This image shows central perivenulitis which encompasses dropout of zone 3 hepatocytes and perivenular mononuclear and/or polymorphonuclear inflammation.

2.7 Necrosis

Causes of central perivenulitis include: • Drug induced hepatitis • Acute cellular rejection

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2.7.2 Confluent Necrosis This image shows confluent necrosis (CN), means loss of more than 20 hepatocytes.

This image shows perivascular inflammatory cells with subendothelial lymphoid cell infiltration.

2.7.3 Lobular Inflammation with Spotty Necrosis

2.7 Necrosis 2.7.1 Bridging Necrosis

This image shows lobular inflammation with neutrophils and loss of hepatocytes (1–2) with spotty necrosis. In addition, there is macrovesicular steatosis.

This image shows large areas of hepatocytes loss (marked in between arrows), extending from one portal tract (PT) to another.

2.7.4 Submassive to Massive Necrosis in Explant Liver This image shows low power view of submassive to massive necrosis. The entire hepatic lobule is

2  Basic Liver Pathology Terminologies

22

necrosed with massive hepatocyte loss, leaving only indistinct eosinophilic remnants of the hepatocyte cords.

2.8 Extramedullary Hematopoesis This image shows collection of erythroid precursors in the sinusoids (marked by arrow), rarely myelocytes and megakaryocytes are also seen. Causes include neonatal hepatitis syndrome, extrahepatic biliary atresia, hepatoblastoma.

Causes include: • • • •

Acute hepatitis Drug induced/toxic hepatitis Acute on chronic liver failure ALF related to acute viral hepatitis

2.7.5 Submassive to Massive Necrosis in Core Needle Biopsy Necrosis of entire lobules is called submassive necrosis or most of the liver called massive necrosis and is usually accompanied by hepatic failure.

2.9 Multinucleated Giant Cell These figures show multinucleated giant cell which are hepatocytes containing 4–10 nuclei. Damaged hepatocytes coalesce to form giant cell.

2.11  Mallory Denk Bodies with Cholestasis

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2.11 Mallory Denk Bodies with Cholestasis This image shows bright eosinophilic/pink acellular inclusions called Mallory Denk bodies (arrow). MD bodies are damaged and clumped intermediate filaments within the hepatocytes. In addition, surrounding hepatocytes shows hepatocellular cholestasis (arrowhead).

Causes of multinucleated giant cell infiltration include: • Neonatal hepatitis syndrome • Progressive familial intrahepatic cholestasis • Extrahepatic biliary atresia(rare)

2.10 Ballooning Degeneration of Hepatocyte Three or four times the size of normal hepatocyte is known as ballooning of hepatocyte. As a result of severe cell injury, these ballooned hepatocytes (marked by dotted line) show perinuclear clumping of intermediate filaments (marked by arrow) and cytoplasmic clearing (marked by star). It is a telltale sign of liver injury. In addition, this image shows hepatocellular cholestasis (arrowhead). Ballooning degeneration is also called as feathery degeneration in the setting of chronic cholestasis. This feathery degeneration occurs due to detergent effect of bile pigments.

Cause of Mallory Denk bodies include: • • • •

Alcoholic hepatitis Non-alcoholic steatohepatitis Wilson’s disease Hepatocellular carcinoma

2.11.1 Mallory Denk Body in HCC Mallory Denk bodies seen as bright eosinophilic deposits (marked by arrows) in the cytoplasm of the malignant hepatocytes showing nucleomegaly and prominent nucleoli (marked by arrowhead).

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2.12 Adequacy of Liver Biopsy 1. Minimum total length 2 cm of liver tissue. 2. 11–15 portal tracts

2  Basic Liver Pathology Terminologies

Note: When there are less portal tracts, diagnosis and differential diagnosis can be given. Grading and fibrosis staging of chronic hepatitis of any etiology with inadequate biopsy may result in underscoring.

3

Pediatric Liver Pathology

3.1 Neonatal Hepatitis Syndrome • Includes all forms of liver dysfunction in the neonate presenting with jaundice due to conjugated hyperbilirubinemia after excluding structural or anatomical disorders of biliary tree. • Etiology: Intrauterine infections like TORCH infection, of which cytomegalovirus infection is the commonest, endocrine causes such as hypothyroidism/hypopituitarism, inherited diseases including chromosomal abnormalities such as Trisomy 18 and 21. • Idiopathic neonatal hepatitis is diagnosed after exclusion of known causes.

trypsin deficiency, primary bile acid synthesis, PFIC II, and idiopathic neonatal hepatitis. • Macrovesicular steatosis favor metabolic disorders. A case of neonatal hepatitis syndrome showing hepatocanalicular cholestasis with numerous multinucleated giant cells, pseudoglandular formation/hepatocyte rosetting (marked by arrows), and portal tract (PT) containing moderate inflammation. Mild fibrous expansion of the portal tract (marked by arrowhead).

3.1.1 Histology Salient histological features: • Varying degree of cholestasis. • Giant cell transformation (common/dominant finding) and ballooning degeneration. • Extramedullary hematopoiesis (EMH). • Lobular and portal inflammation. • Progressive fibrosis in some cases. • Lobular inflammation with hepatocellular damage varies, usually severe in alpha-1 anti-

No significant ductular reaction/bile duct plugging/prominent hepatic arterioles in this case as compared to biliary atresia. This image shows loss of reticulin framework with hepatocyte rosetting (marked by arrows).

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3  Pediatric Liver Pathology

This rhodanine stained image highlights increase in copper/copper associated proteins in periportal hepatocytes (marked by arrow).

A case of neonatal hepatitis syndrome, image here, highlights pericellular/sinusoidal fibrosis (marked by arrows) indicative of degenerative changes with hepatocyte rosettes (marked by arrowhead).

The image here, shows lobular parenchyma with numerous multinucleated giant cells (marked by arrow), severe hepatocanalicular cholestasis (marked by arrowhead), and ballooning degeneration (marked by star).

3.2 Extra Hepatic Biliary Atresia • Complete or partial obstruction of biliary tree within first three months of life leading to cholestasis, fibrosis, and cirrhosis. • Commonest cause of cholestatic jaundice requiring liver transplant in infants. • Clinical features: Persistent jaundice, dark colored urine, pale colored stools, failure to thrive, and hepatomegaly within first few days of life. • Liver function test: Conjugated hyperbilirubinemia exceeding 100 μmol/L (5 mg/dL), ALP exceeding 600 IU/L, γ-GT exceeding 100 IU/L, ALT and AST between 100 and 200 IU/L. • Fasting ultrasound shows no gall bladder or contracted gall bladder. • Biliary scintigraphy (HIDA scan) shows no isotope secretion into the bowel 24 h after administration. • Treatment: surgical, Kasai portoenterostomy—excision of the obliterated biliary channel and anastomosis with proximal jejunal loop followed by medical management. • Almost 80% develop persistent jaundice requiring liver transplantation, which has more than 90% long-term survival. Gross Macroscopic appearance: Gross section of the liver from patient with biliary atresia. Enlarged/ green in color with multiple dilated hilar bile

3.2 Extra Hepatic Biliary Atresia

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ducts (marked by red arrows) resembling Caroli disease. This feature occurs after 3 months of life not amenable to surgical intervention. The hepatic parenchyma is cirrhotic (marked by thin arrows).

This image shows high power view of portal tract with angular, linear, and irregular neocholangioles (N) (periportal ductular reaction).

3.2.1 Histology 3.2.1.1 Salient Histological Features • Edematous fibroplasia of portal tract with periportal ductular reaction. • Neocholangioles are usually distorted and irregular, forming anastomosing network at the portal periphery (necklace pattern). • Cholestasis. • Prominent hepatic arterioles. • Giant cell transformation seen in some cases. • Edematous fibroplasia (loose fibrosis) is progressive, resulting in portal bridging fibrosis leading to secondary biliary cirrhosis. • Variable to marked copper accumulation in the liver cells, as seen in other bile duct pathology like PFIC, PBC, Wilson’s, etc. This image of EHBA shows portal tract in the center and periportal ductular reaction along the interface, necklace pattern (marked by arrows). There are prominent hepatic arterioles in the center of portal tract (marked by star) without associated interlobular bile duct (orphan arterioles).

This image of EHBA highlights mild increase in copper in periportal hepatocytes (marked by arrows) with ductular bile plugging (marked by star).

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3  Pediatric Liver Pathology

This image of EHBA shows edematous portal fibroplasia (PT) with porto-portal bridging fibrosis (marked by arrows).

3.2.1.2 Bile Duct Pictures for EHBA This image shows scanner view of cross section of the common hepatic duct which shows complete obliteration of lumen by fibrous tissue containing sparse inflammatory cells and bile casts. The high power view depicts the nature of the inflammatory cells which are predominantly lymphocytes with bile casts.

The following image is a bile duct of an EHBA case. This shows distorted bile duct (BD) lined by non-continuous columnar epithelium (marked by arrowhead) and surrounded by inflammation and peribiliary glands (marked by arrows). Inflammation composed of neutrophils (marked by circles) and lymphocytes (marked by thin arrow).

3.3 Paucity of Intrahepatic Bile Ducts: Alagille Syndrome • Alagille syndrome (AGS): Syndromic paucity of interlobular bile ducts is a rare autosomal dominant genetic syndrome that can affect liver, heart, skeleton, eyes, and kidneys.

3.3 Paucity of Intrahepatic Bile Ducts: Alagille Syndrome

• AGS is caused in majority by mutation of Jagged1 (JAG1) in chromosome 20 which codes for Notch1 and in AST), or increased echogenicity of liver on ultrasound imaging. Newer modalities of imaging like fibroscan, magnetic resonance elastography are useful in determining the degree of hepatic steatosis and fibrosis. Various clinical scoring systems have come into play to assess the severity of NAFLD related fibrosis. Example: NAFLD fibrosis score that includes age, body mass index (BMI), serum albumin levels, AST/ALT ratio, platelet count, and blood sugar levels is more commonly used at present. Treatment includes lifestyle modification like low fat/sugar diet, aerobic exercises 3–4 times/ week, and pharmacotherapy. Medications currently in use include vitamin E, PPAR agonists (pioglitazone, saroglitazar), incretin-based therapy-GLP1 agonists (exenatide and liraglutide), dipeptidyl peptidase 4 inhibitors, and obeticholic acid. Newer modalities of pharmacotherapy like anti-fibrotic agents are under trial.

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5.1.1 Histology Salient histological features: • Steatosis to steatohepatitis followed by fibrosis to cirrhosis.

5.1.1.1 Steatosis (Non-alcoholic Fatty Liver Disease, NAFLD) • Steatosis fatty change: Macrovesicular (scored 1–3 based on the percentage of liver cells involved in non-alcoholic liver disease activity score clinical research network—NAS CRN Score), see below table. • Steatosis in non-alcoholic fatty liver disease is usually accompanied by portal inflammation (typically mild). Portal inflammation composed of mononuclear inflammatory infiltrate. Mixed inflammation with neutrophil is less common. 5.1.1.2 Steatohepatitis (Non-alcoholic Steatohepatitis, NASH) • Lobular inflammation is common in steatohepatitis (NASH). • Composed of lymphocytes, sometimes mixed infiltrate with polymorphonuclear leukocytes may be a minor component. • Lipogranuloma is a frequent finding in NASH. Less common in ALD (alcoholic liver disease). • Neutrophilic satellitosis is less common in NASH than in ALD (alcoholic liver disease). • Lobular inflammation also scored 1 and 2 (non-alcoholic liver disease activity score clinical research network—NAS CRN score see table below). • In addition to steatosis, lobular inflammation and ballooning degeneration with or without Mallory Denk bodies are seen. • Glycogenated nuclei in periportal hepatocytes are seen as in Wilson. • Early stages show centrizonal sinusoidal fibrosis with or without concomitant portal tract fibrosis, progressing to bridging fibrosis and cirrhosis.

5  Fatty Liver Disease

• In pediatric non-alcoholic fatty liver disease—portal fibrosis without zone 3 centrizonal perisinusoidal fibrosis is a common finding. This image shows macrovesicular fatty change in >70% hepatocytes occupying all the zones (marked by arrows).

This image shows macrovesicular fatty change (marked by arrow). Portal tract is largely normal/ unremarkable (marked by dotted lines).

This image shows hepatic parenchyma (HP) with macrovesicular fatty change (marked by arrowhead) and lobular inflammation (lipogranuloma—marked by dotted lines) containing lymphocytes and epithelioid macrophages (marked by arrow)/Kupffer cells. The above features are seen in steatohepatitis phase of alcoholic or non-­ alcoholic liver disease.

5.1  Non-alcoholic Fatty Liver Disease (NAFLD)

This image shows ballooning degeneration with clustering of intermediate filaments (marked by arrow), seen in alcoholic steatohepatitis.

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F3 Fibrosis.

MT stain

NASH Clinical research network (CRN) scoring [adapted from MacSween]. This image shows part of central vein and zone 3 hepatic parenchyma with pericellular fibrosis extending in between the hepatocytes.

5  Fatty Liver Disease

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5.2 Alcoholic Liver Disease • Caused by injury to the hepatocytes from high quantity and prolonged consumption of alcohol. • As liver is the major site for ethanol metabolism, formation of acetaldehyde plays a major role of mitochondrial damage and oxidative stress results in hepatocyte damage. • More than 90% of chronic alcoholics will develop fatty liver, whereas only 10–20% will develop subsequent progression to alcoholic hepatitis, fibrosis, and cirrhosis. • The threshold for developing alcoholic liver disease in men is approximately more than 60  gm/day for 10  years, whereas it is still

•

•

•

• •

lower for women, obese individuals, smokers, and simultaneous exposure to other hepatotoxins. Usually remains subtle and is incidentally detected while investigating for unrelated medical ailments. Clinical presentation includes right upper quadrant pain, jaundice, nausea, and tender hepatomegaly. Liver function tests reveal two- to seven-fold increase in liver enzymes (AST > ALT), elevated GGT, and hyperbilirubinemia. Presence of hypoalbuminemia and coagulopathy indicates severe liver injury. Ultrasound imaging shows fatty infiltration of liver with increased echogenicity. CT/MRI

5.2  Alcoholic Liver Disease

• •

• •

•

may also reveal fatty liver, surface nodularity, presence of collaterals/splenomegaly. Severe alcoholic hepatitis might lead to early liver transplantation. Maddrey discriminant function (MDF) is calculated to assess severity of alcoholic hepatitis and decide who requires prednisolone therapy. If MDF is more than 32, prednisolone 40 mg/ day for 4 weeks f/b tapering has been found to be effective in reducing the hepatic inflammatory response. MDF formula  =  4.6  ×  [prothrombin time— control (s)] + serum bilirubin (mg/dL). Lille’s score in severe alcoholic hepatitis will help to identify non-responders of steroid therapy. Treatment mainly involves complete abstinence from alcohol, treatment of alcohol dependence/withdrawal and psychiatric counseling.

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5.2.1.3 Severe Alcoholic Hepatitis and Alcoholic Cirrhosis • Confluent necrosis, ballooning degeneration, and Mallory Denk bodies. • Progress to perivenular fibrosis and alcoholic cirrhosis (most commonly micronodular (5 indicates hepatocellular, AST) in ­hepatocellular pattern and modest elevation of ALP levels in cholestatic pattern.

7.1.1 Patterns of Drug Induced Liver Injury • The onset of clinical symptoms like fatigue and pruritis will occur after 2–3  months of drug exposure. • Liver function test reveals moderate elevated ALP levels, mild increase in ALT and normal GGT levels. • The injury is self-limiting and reversible within days to weeks following dose reduction/cessation of the drug.

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7  Toxin Induced Hepatitis

• In rare instances, progression to chronic cholestasis, peliosis hepatis, and hepatocellular carcinoma.

7.2 Steatohepatitis • Drug induced steatohepatitis is characterized by focal hepatocyte injury/fibrosis along with steatosis. • The most common drugs causing steatosis and steatohepatitis include methotrexate, amiodarone, tamoxifen, raloxifene, anastrozole, and letrozole followed by calcium channel blockers, methyldopa, and certain chemotherapeutic agents like irinotecan. • These drugs act synergistically in individuals with predisposition to NASH like obesity, dyslipidemia, and diabetes. • Tamoxifen, an estrogen receptor antagonist, has been found to cause steatohepatitis in nearly 30% women within a median time interval of 2 years. • Periodic clinical examination, LFT, and hepatic imaging are mandatory for early detection of NASH in women on tamoxifen/ SERM therapy. • Continuation of these agents may result in progression to massive hepatic necrosis, peliosis hepatis, acute hepatitis, and hepatocellular carcinoma. • In addition, certain drugs like methotrexate, arsenic, and vitamin A will directly stimulate hepatic stellate cells resulting in fibrosis. Drug induced steatosis with fibrosis (methotrexate toxicity): H&E image shows diffuse macrovesicular steatosis with moderate portal fibrosis and septae. This is the case of chronic rheumatoid arthritis on methotrexate.

7.2.1 Other Patterns of Drug Induced Liver Injury Cholestatic hepatitis and bland canalicular cholestasis. Cytolytic/necrolytic hepatitis. Granulomatous hepatitis.

7.3 Yellow Phosphorus Poisoning • Commonly available as metal phosphides like zinc/aluminum phosphide and is used in the manufacture of rodenticide, fertilizers, fireworks, and ammunition. • Accidental/suicidal consumption of metal phosphides is more common in southern states of India. • It acts as a protoplasmic toxin resulting in hepatocyte necrosis and death. • The clinical and biochemical features of liver injury will usually appear after 48–72  h of consumption and include nausea, vomiting, right upper quadrant pain, and elevation of serum transaminase levels.

7.3  Yellow Phosphorus Poisoning

• Extrahepatic manifestations include cardiotoxicity/circulatory failure, pancreatitis, acute kidney injury, and CNS dysfunction. • Poor prognostic indicators are jaundice, severe coagulopathy, hepatic encephalopathy resulting in cerebral edema/coma, lactic acidosis, and renal dysfunction. • Initial treatment involves gastric lavage with activated charcoal, N-acetyl cysteine, and symptomatic management. • Therapeutic plasma exchange has been found to be effective in improving transplant free survival in patients not fulfilling King’s College criteria. • Ingestion of lethal doses (more than 1 mg/kg body weight) usually results in fulminant hepatic failure and requires liver transplantation.

7.3.1 Histology 7.3.1.1 Salient Histological Features • Microvesicular fatty change and sinusoidal congestion appear as an early histological features. • Followed by inflammatory cell infiltrate, hepatic cell necrosis, and cholestasis. • Hepatic necrosis can vary from focal (periportal), multifocal, submassive, and panacinar necrosis. • Bile duct proliferation can be seen. Case of yellow phosphorous poisoning presented as acute liver failure. H&E stain, low power view of explant liver in yellow phosphorus poisoning showing bridging

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and confluent necrosis (marked by arrows) with micro and macrovesicular steatosis (marked by star) and portal/ parenchymal ductular reaction.

High power view of a case of yellow phosphorus poisoning with submassive necrosis, hepatocanalicular cholestasis, ballooning degeneration, and sparse mixed inflammatory infiltrate in the necrotic parenchyma.

8

Vascular Liver Disease

8.1

Budd Chiari Syndrome (BCS)

• Defined as obstruction of hepatic venous outflow anywhere along the small hepatic venules up to entrance of IVC into the right atrium. • Excludes causes of hepatic outflow obstruction such as cardiac disease, pericardial disease, or sinusoidal obstruction syndrome. • Classified into (1) primary—caused by thrombosis in the absence of compression by SOL or invasion by parasites or malignancy and (2) secondary otherwise. • In Asia, pure IVC or combined IVC/hepatic vein block predominates, while in west pure hepatic venous block is common. • Risk factors for primary BCS include prothrombotic conditions which can be acquired like myeloproliferative neoplasia (most common primary cause), inherited like factor V Leiden mutation or other factors like oral contraceptives, pregnancy, or inflammatory bowel disease. • Presentation can range from asymptomatic to all possible clinical features of acute or chronic liver disease like ascites, hepatomegaly, splenomegaly, jaundice, gastrointestinal bleeding, bacterial infections, and encephalopathy. • A triad of abdominal pain, fever, and ascites of rapid onset suggests an acute episode of

hepatic vein thrombosis or recent progression of long-standing disease. • Treatment includes medical treatment with anti-coagulation, angioplasty, stenting or intravascular thrombolysis, TIPS, and liver transplantation in a stepwise algorithm.

8.1.1 Histology Salient histological features: • Acute presentation marked congestion/centrilobular mid zonal sinusoids with hepatocyte necrosis to variable extend. • Linking of adjacent central veins with hemorrhage and necrosis seen. • Periportal region are spaced. • Portal tracts lack significant pathological changes. • Panlobular necrosis/hemorrhage also seen in BCS. • Over time affected parenchymal region shows centrilobular and perisinusoidal fibrosis. Explant liver of Budd-Chiari syndrome: All three images are low and high power view of marked centrilobular congestion with hemorrhagic necrosis—marked by star (centrizonal and bridging necrosis).

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8  Vascular Liver Disease

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Marked lobular congestion with hemorrhagic necrosis—marked by star (Nutmeg liver).

H & E stain

Low power view—Nutmeg liver, portal tract in the center which is unaffected (marked by star) and shows no portal fibrous expansion and mild inflammation.

H & E stain

Shows bridging necrosis.

H & E stain

H & E stain

9

Infiltrative Liver Disease

9.1

Sarcoidosis

• Systemic disease is characterized by granulomatous lesions, more commonly involving the lung followed by eyes, skin, joints, lymph nodes, spleen, liver, kidneys, and nervous system. • The exact pathogenesis is considered to be multifactorial with genetic predisposition, autoimmune triggered by environmental antigens and infections. • Involvement of liver is seen in about 70% cases on liver biopsy, out of which only 20% develop clinical manifestations of jaundice, pruritis, and hepatomegaly. • Rarely, it may result in hepatocyte dysfunction, cell death, and fibrosis, which become evident only on liver biopsy. • Rare hepatic manifestations include portal hypertension, Budd-Chiari syndrome, extrahepatic cholestasis due to portal hilar lymphadenopathy, bile duct inflammation, and secondary sclerosing cholangitis. • Liver function test will show elevated ALP levels. Serum ACE levels may also be elevated but not confirmatory for diagnosis. • Diagnosis is mainly based on histological evidence of non-caseating granuloma.

• CT/MRI imaging will reveal focal granulomas or diffuse isointense or hyperintense parenchyma. T2 hypointensity is characteristic of diffuse involvement of liver and helps to differentiate from metastatic or inflammatory disorders. • Treatment is mainly supportive as hepatic involvement does not respond well to steroids. Initiation of corticosteroids may be considered in patients with associated other organ involvement/systemic symptoms like fatigue.

9.1.1 Histology Core needle liver biopsy from a case of systemic sarcoidosis with generalized lymphadenopathy (histologically proven from cervical lymph node biopsy). Patient presented with deranged liver function test in the form of elevated alkaline phosphatase and MRI abdomen showed nodular liver with splenomegaly. H&E stain, high power view showing ill-­ defined non-caseating granuloma (marked by dotted lines) within the portal tract with surrounding dense fibrosis. Adjoining granuloma with giant cell (marked by arrow).

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9  Infiltrative Liver Disease

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H&E stain and MT stain of the above described sarcoidosis case showing (low power view) cirrhotic nodule with markedly fibrosed portal tract.

• Other less common manifestations include tuberculous cholangitis, biliary stricture, tuberculous pylephlebitis, and periportal tuberculous lymphadenopathy. • Clinical features are usually nonspecific and present as pyrexia of unknown origin with fever, night sweats, and occasionally weight loss. • Jaundice appears at later stages and rarely leads on to fulminant hepatic failure. • Liver function tests will reveal elevated ALP levels with/without increase in bilirubin and hypoalbuminemia. • Tuberculomas are usually difficult to differentiate from lymphoma/metastatic deposits/actinomycosis based on imaging and liver biopsy is mandatory to confirm the diagnosis. • Diagnosis is based on detection of acid-fast bacilli, culture, PCR based testing like Gene-­ Xpert, and histopathological examination in biopsy specimen. • Screening to rule out pulmonary and other extrapulmonary sites of involvement should be done before initiation of treatment. • Treatment usually is with anti-tuberculous medications: 2 months of intensive phase with four drugs followed by 4 months of continuation phase with two drugs and close monitoring of LFT for ATT induced liver injury.

9.2.1 Histology H&E stain showing non-necrotizing, well-­ defined granuloma (marked by dotted lines) in lobular parenchyma.

9.2 Tuberculosis • Tuberculous involvement of the liver may occur in various forms, namely isolated granulomas, diffuse granulomatous infiltration or as part of military tuberculosis with/without pulmonary involvement.

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9.2 Tuberculosis

This image shows part of the fibrosed portal tract with ill-defined granuloma containing scattered epithelioid histiocytes (marked by arrowhead) and multinucleated Langhans giant cells (marked by arrows).

MT stain with bridging fibrosis (marked by arrow) and lobular parenchyma based granuloma (G).

Differentiating features between granulomatous hepatitis due to tuberculosis, sarcoidosis, and invasive fungal infection. S. No. 1.

Features Granulomas

2.

5.

Location—Portal tract (PT) or hepatic parenchyma (HP) Necrosis Other specific features Special stains

6.

Fibrosis

3. 4.

Tuberculosis Well defined with Langhans type multinucleated giant cell Both portal tract (pt) and hepatic parenchyma (HP)

Sarcoidosis Well or ill-defined, but large plump epithelioid histiocytes

Fungal Ill-defined with foreign body type multinucleated giant cells

Usually PT

Usually PT

(+) or (−) Gene-Xpert or TB PCR (+ve) AFB stain—May be positive

Mostly (−) May have Schaumann bodies and asteroid bodies Calcium stain von kossa for intracytoplasmic calcospherules, Perl’s stain for iron (+/−) More severe portal based fibrosis rarely cirrhosis

(+) or (−)

Portal fibrosis, mild to moderate

PAS/GMS for fungus

Absent

Neoplasms of Liver

10

10.1 Regenerative Nodule • Regenerative nodules are benign, single/multiple, rounded, and sharply circumscribed lesions usually seen in the background of cirrhosis. • Based on size of nodules, they are named as micronodules, macronodules, or giant regenerative nodules (>5 cm). • It results from an adaptive hyperplastic reaction to altered hepatic perfusion, secondary to cirrhosis and portal hypertension. • They are usually not visible on plain CT unless they are siderotic and show similar enhancement as normal liver parenchyma in portal and delayed phases with no enhancement on arterial phase. • May rarely progress to form dysplastic nodules and HCC.  Hence, close follow-up for increase in size of nodule and monitoring of viral markers is mandatory.

This image shows part of regenerative nodule (RN) in acute liver failure with ill-defined border (marked by dotted line) in between regenerative hepatocytes and adjoining necrotic liver (NL).

10.1.1 Histology Gross image shows a well circumscribed regenerative nodule which is solid, similar in consistency and color to the adjoining pale green liver parenchyma. Adjoining liver is smooth, shrunken, and no cirrhotic nodules noted (a case of acute liver failure with regenerative nodule).

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 K. S. Mouleeswaran et al., Atlas of Basic Liver Histology for Practicing Clinicians and Pathologists, https://doi.org/10.1007/978-981-99-5762-0_10

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This image shows regenerative nodule with hyperplastic hepatocyte with increased cell plate thickness.

•

• • •

This image shows regenerative nodule with entrapped portal tracts containing intact bile duct (BD), hepatic artery (marked by arrow), and portal venules (PV). Also, canalicular cholestasis is noted within the regenerating hepatocytes.

10.2 Focal Nodular Hyperplasia (FNH) • Focal nodular hyperplasia is a regenerative mass lesion of the liver due to local vascular abnormalities or malformation and the second most common benign liver lesion after hemangioma. • It has female preponderance (8:1) and usually occurs between third and fifth decade, and

association with oral contraceptive use has been suggested. FNH is asymptomatic and incidentally discovered during imaging, but in patients with large nodule, FNH may be responsible for vague, nonspecific abdominal discomfort or pain, feeling of fullness, or dyspepsia. Intratumoral bleeding is a rare complication. Diagnosis can be made by imaging studies like ultrasound, MRI, or CT. Treatment is usually conservative, and surgical treatment is indicated only for persistent pain or for lesions with unclear diagnosis after radiological and pathological investigations.

10.2.1 Histology Salient histological features: • The color is lighter than that of the adjacent normal liver with fibrous septa. • Microscopically, FNH is characterized by hyperplastic nodules that are separated by fibrous septa. • The fibrous septa and central scar contain thick-walled arteries and to a lesser extent, veins. • The scar and fibrous septa typically show a ductular reaction and variable lymphocytic inflammation, while interlobular bile ducts are absent. • The hepatocellular component is arranged in plates one to two cells thick, with intact reticulin framework. • Fat accumulation can be seen, especially in the patient with fatty liver disease. • On IHC, characteristic geographical or map-­ like pattern on glutamine synthetase (GS) staining, with strong cytoplasmic staining of broad anastomosing groups of hepatocytes. Gross image (hepatectomy specimen) showing tan lighter in color ill-defined lesion.

10.2  Focal Nodular Hyperplasia (FNH)

This image of FNH shows central fibrous scar (S) and adjoining hepatocytes (HP) showing focal fatty change (marked by blue arrow).

High power view of the lesion showing arterioles (Ar) in a thin fibrous septum and macrovesicular fatty change (marked by arrowhead) in some of the hepatocytes. Interlobular small ducts seen in normal portal tracts are usually absent, though there may be ductular reaction in the fibrous septa.

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The following image of CD34 IHC of focal nodular hyperplasia shows no sinusoidal expression in the lobular parenchyma of hepatocytes which is a differentiating point between FNH and HCC [Internal positive control uptake in portal venules/capillaries, central vein, and periportal sinusoids—marked by arrow]. Some FNH may show sinusoidal expression of CD 34. Reticulin stain and glutamine synthetase (GS) activity by IHC are more helpful.

Below IHC image shows increased glutamine synthetase (GS) activity in FNH (marked by arrows). GS IHC demonstrates broad zones of hepatocyte brown staining alternating with zones of nonstaining hepatocytes—map-like pattern. In normal liver, GS expression is limited to narrow rim of hepatocytes around the central vein (centrizonal region), which in turn relates to beta catenin activity in the hepatocytes. Expansion of beta-catenin activated centrizonal region leads to overexpression of GS in FNH.

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• Large cavernous hemangioma (>10  cm) can have foci of thrombosis resulting in extensive fibrosis. A case of hemangioma. This image shows variably sized, dilated, and thin-walled vessels filled with blood (marked by arrows) with surrounding hepatic parenchyma (HP).

10.3 Benign Tumors 10.3.1 Hemangioma • It is the most common primary, benign liver tumor, detected incidentally on imaging or surgery or on autopsy. On gross examination of liver, they appear as reddish or brownish lesions. • Patients with larger hemangiomas (>5  cm) may present with symptoms of abdominal pain, discomfort, abdominal lump, or hepatomegaly. • Rarely, large cavernous hemangiomas may be associated with consumptive coagulopathy, thrombocytopenia, and hypofibrinogenemia, as part of Kasabach–Merritt syndrome or von Hippel–Lindau disease. • Hemangiomas are easily identified in ultrasound or CT imaging. The center of the lesion remains hypodense, with variable thickness and enhancing peripheral zone. • Most hemangiomas do not require any intervention and surgical resection is attempted only in case of large hemangiomas with hemorrhagic complications.

10.3.1.1 Histology Salient histological features: • Histologically shows vascular channels lined by flat bland endothelial cells overlying the fibrous stroma.

This is a high power view of the vascular spaces (VS) separated by fibrous septa lined by a single layer of flat endothelial cells (marked by arrows).

10.3.2 Hepatic Adenoma • It is a benign neoplasm involving noncirrhotic liver, commonly seen in women between third and fifth decade of life. • It has been classified into four types based on etiology, histology, and molecular characteristics, namely hepatocyte nuclear factor

10.3  Benign Tumors

•

•

•

•

•

•

1α-inactivated (HHCA), inflammatory (IHCA), β-catenin-activated (BHCA), and unclassified. The pathogenesis is mainly based on exposure to estrogen, androgens, or anabolic steroids resulting in increased hepatocyte proliferation and increase in size of adenomas. Other predisposing conditions include maturity onset diabetes of the young (MODY), glycogen storage disorders, obesity/metabolic syndrome, and other inherited syndromes like familial adenomatous polyposis, Fanconi’s anemia, and Klinefelter syndrome. Most patients are diagnosed incidentally based on imaging and less than one-fourth present with severe abdominal pain secondary to hemorrhage into the tumor. Hepatic adenomas rarely turn malignant to HCC (4–8%), especially in the presence of risk factors like advanced age, male gender, use of anabolic steroids, metabolic syndrome, and large tumor size. Treatment mainly involves discontinuation of inciting medication, dietary modification in case of glycogen storage disorders, and resection in case of large tumors (size>5 cm). Chemoembolization serves as an alternative in candidates with high surgical risk and close follow-up with CT/MRI every 6 months.

10.3.2.1 Histology Salient histological features: • Histologically composed of hepatocytes arranged in thin cell plate (one to two cells thick) without cytological atypia. • Tumor cells area uniform in size and shape. Reticulin framework is intact without fragmentation. • Immunohistochemistry—Absence of CD34 IHC expression in the heaptic sinusoids, glypican 3 (negative) and HSP-70 (negative) will help to distinguish from hepatocellular carcinoma. A case of hepatic adenoma. This image shows hepatocytes arranged in trabecular pattern with mildly thickened cell plate, 1–2 cell thick without cytological atypia. Unlike HCC, reticulin framework is intact in hepatic adenoma.

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Another case of hepatic adenoma with hepatocytes showing diffuse macrovesicular steatosis (marked by arrow).

In hepatic adenoma, interlobular bile ducts are not made out in portal tract (PT). This image shows portal tract with portal venules and absent bile duct.

A case of hepatic adenoma, where IHC for beta catenin shows patchy nuclear positivity in

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84

hepatocytes (marked by arrow). Glutamine synthetase IHC staining (map like pattern in FNH vs diffuse staining in hepatic adenoma) with beta catenin may be useful in distinguishing FNH from hepatic adenoma in core needle biopsies where fibrous septa are absent.

• Uniformly shaped and spaced tubules or ductular structures without dilatation lined by a single layer of cuboidal to columnar cells with intact basement membrane. • These tubules are embedded in a fibrous stroma. • Normal portal tract can be identified within the lesion. The following low power and high power images show circumscribed lesion composed of largely uniform ducts lined by cuboidal epithelium. High power shows intervening fibrous stroma and lymphoid aggregates. No normal portal tract identified in this neoplasm.

10.3.3 Bile Duct Adenoma • Bile duct adenomas are tumor like lesions arising secondary to inflammatory response following localized injury, remote ischemic event, or in late stages of cirrhosis. • Less common when compared to biliary hamartomas. • Almost 80% are solitary lesions and are subcapsular in location. • Usually asymptomatic and incidentally detected during surgery. • Features that help to differentiate this lesion from adenocarcinoma are: –– Presence of pre-existing portal areas within the bile duct adenoma. –– Uniform spacing and configuration of the glands and tubules—rounded rather than angulated. –– Absence of destructive infiltration of desmoplasia, significant pleomorphism, and mitoses. Salient histological features:

H & E stain

H & E stain

PAS stain of the same case shows prominent bile duct basement membrane highlighted by pink linear hue around the bile ducts.

10.3  Benign Tumors

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PAS stain

10.3.4 Biliary Cystadenoma

• Gross: Multilocular smooth cystic lesion with serous/mucinous/gelatinous secretion. • Histologically the cyst wall lined by a single layer of cuboidal epithelium, usually of mucinous type (strong positive for IHC on CK7 and CK19). • Rarely intestinal/gastric/squamous metaplasia in seen. • Secondary to rupture of cyst adjoining areas show xanthogranulomatous reaction, scarring and calcification. • Underlying stroma resembles ovarian stroma (positive for IHC on ER, PR, and inhibin). This image of biliary cystadenoma shows multiple cystic spaces lined by single layer of columnar/cuboidal epithelium (marked by arrow).

• Biliary cystadenomas are rare (constitute 3/more cell thick).

10.4  Malignant Tumors

• Other patterns such as pseudoglandular, pseudoacinar pattern, peliotic pattern, and macrotrabecular pattern are seen. • The hepatocellular carcinoma can be well to poorly differentiated. The well differentiated resemble as normal hepatocytes with abundant cytoplasm to poorly differentiated with increased nuclear cytoplasmic ratio, angulated nuclei with coarse chromatin. • Reticulin stain shows fragmented reticulin network of the hepatic plate is one of the most useful techniques for the diagnosis of HCC. • Immunohistochemistry: Arginase, Heppar-1 (hepatocyte paraffin 1), Glypican-3 is the confirmatory immunohistochemistry for hepatocellular carcinoma.

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H&E stain, high power view showing tumor cells arranged in cohesive nest and trabecular pattern (marked by arrow).

Gross Image

This image shows cut section from explant liver with pale, firm solid well defined lesion- HCC and background cirrhotic liver made out well along the capsular surface.

The following image shows grade II hepatocellular carcinoma with tumor cells showing hyperchromatic angulated nuclei and moderate eosinophilic cytoplasm.

H&E stain, low power view: Moderately differentiated hepatocellular carcinoma with tumor cells (marked by arrow) arranged in trabecular and macrotrabecular pattern.

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The following image shows moderately differentiated hepatocellular carcinoma clear cell variant with tumor cells arranged in solid nests and trabecular pattern with abundant clear cytoplasm and hyperchromatic nuclei.

IHC CD34 shows sinusoidal pattern of expression of CD34 in between the tumor cells in the sinusoids (marked with arrow).

IHC arginase image shows strong nuclear and cytoplasmic positivity (marked by arrow) of tumor cells.

IHC Heppar 1 shows the tumor cells with strong granular cytoplasmic positivity (marked with arrow) of the malignant hepatocytes.

10.4.1.2 Fibrolamellar Hepatocellular Carcinoma • It usually occurs in the absence of chronic liver disease/cirrhosis and more commonly seen in younger individuals. • Serum AFP levels are usually normal and these tumors differ from classic HCC in histology, immunophenotype, and molecular level.

10.4  Malignant Tumors

• Diagnosis of fibrolamellar variant is mainly based on the following three microscopic criteria, namely large polygonal tumor cells with granular eosinophilic cytoplasm, prominent nucleoli, and lamellar fibrosis. • Fibrolamellar variant results from deletion mutation in chromosome 19 resulting in formation of a chimeric fusion protein DNAJB1, which causes overexpression of key regulatory kinase PRKACA. • The clinical presentation and radiological features are usually indistinguishable from classic HCC. • Regional lymph nodal metastasis is seen in 3/ fourth of patients requiring lymph node dissection during lobectomy. • Although FLC is an aggressive tumor, the prognosis and outcome are similar to that of classic HCC and mainly depend on resectability of the lesion.

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H&E stain, high power view showing tumors cells (marked by arrow) with abundant granular eosinophilic cytoplasm and round to oval nuclei surrounded by dense collagen bundles (marked by star).

IHC for CK7: Strong membranous and cytoplasmic positivity (marked by arrow).

Histology Salient histological features: • Large polygonal tumor cells with abundant eosinophilic granular cytoplasm, round to oval nuclei with eosinophilic macronucleoli. • There is lamellar fibrosis (parallel stacks of collagen) interspersing the tumor cells. • Immunohistochemistry: The tumor cells are positive for glypican-3 and CK7. H&E stain, low power view showing sheets and cords of tumors cells (marked by arrow) interpersed by dense collagen bundles (marked by star).

IHC for Heppar 1: The tumor cells show strong cytoplasmic positivity (marked by arrow).

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10.4.1.3 Hepatoblastoma • Most common primary liver malignancy in pediatric population, it accounts for 1% of all pediatric malignancies. • Usually affects children younger than 3 years. • Typically, presents as abdominal distension and tumor may rupture due to rapid growth leading to hemoperitoneum. • Sexual precocity may be present due to ectopic gonadotropin synthesis. • Genetic syndromes associated with hepatoblastoma include Beckwith-Wiedemann syndrome, Edwards syndrome, and familial adenomatous polyposis. • Serum AFP is used for diagnosis, prognosis. and treatment monitoring. • Doppler ultrasonography will help to identify the lesion followed by contrast enhanced CT or MRI with hepatobiliary specific contrast to look for the extent of the disease and respectability. • Diagnosis should be confirmed by biopsy. • Treatment is surgical resection with or without adjuvant/neoadjuvant chemotherapy. • Liver transplantation can be considered with extensive disease where resection is not feasible. • Prognosis is better compared to HCC but worse compared to other pediatric malignancies.

10  Neoplasms of Liver

nuclear cytoplasmic ratio, prominent nucleolus, and frequent mitosis (resemble eighth week of embryonic liver). • Other pattern small cell undifferentiated and cholangioblastic (hepatoblastoma with ductular differentiation). • Mixed epithelial and mesenchymal admixed with mesenchymal component includes fibrous tissue, osteoid like tumor hyaline cartilage. • Rare variant teratoid variant (epithelial with neural/neuroectodermal differentiate with mature brain, primitive neuroepithelial cells like tubules and rosettes). Immunohistochemistry: PanCK, Heppar-1 (hepatocyte paraffin 1), polyclonal CEA, AFB PGC, nuclear beta-catenin (only in embryonal). H&E Stain: Hepatoblastoma—Fetal pattern shows tumor cells arranged in sheets of polygonal cells with clear cytoplasm (marked by arrows) along with fragment of adjacent normal liver parenchyma (marked by star).

Histology Salient histological features: • Histologically classified into epithelial/mixed epithelial—mesenchymal type. • Epithelial is the most common with different subtypes that includes: –– Fetal sheets of thin trabeculae of polygonal cells resembling fetal hepatocytes with small round nuclear, inconspicuous nucleolus, and clear to finely granular cytoplasm. • Embryonal sheets and clusters of single small, angulated hypochromatic cells with increased

The following image is the case of hepatoblastoma—Fetal pattern at higher magnification shows sheets of polygonal cells with small round inconspicuous nucleus (marked by arrows) and clear to finely granular cytoplasm (marked by arrowhead).

10.4  Malignant Tumors

91

PAS staining highlights the clear cytoplasm (marked by arrow) in hepatoblastoma fetal pattern.

Hepatoblastoma: Fetal pattern at higher magnification shows foci of extramedullary hematopoiesis with collection of erythroid precursors in the sinusoids (marked by arrow).

Masson Trichrome: Hepatoblastoma—Fetal pattern (marked by star) in Masson trichrome stains highlights the adjacent normal liver parenchyma (marked by arrow) with bile duct and central vein.

IHC glutamine synthetase: Hepatoblastoma— Fetal pattern shows cytoplasmic staining for glutamine synthetase (marked by arrow).

10.4.2 Biliary Epithelial Origin 10.4.2.1 Cholangiocarcinoma • Arises from biliary epithelium of intra or extrahepatic bile ducts, with varied clinical features and outcome based on the location of the tumor. • Second most common primary hepatic malignancy with increasing incidence in recent years.

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• Based on their location within the biliary tract, divided into intrahepatic, perihilar, and distal subtypes. Perihilar subtype, also known as Klatskin tumor, is further classified into Bismuth Corlette types I–IV, based on site of involvement along the biliary tree. • Risk factors include conditions causing chronic inflammation within the biliary tree as in PSC, infections with Clonorchis/opisthorchis species, exposure to chemical carcinogens and Caroli’s disease. • Most patients with extrahepatic disease are asymptomatic during the initial phases and present lately with features of obstructive jaundice. • Intrahepatic cholangiocarcinoma patients develop vague abdominal pain and systemic symptoms like malaise, fatigue, and weight loss. • Investigations include deranged liver function tests (cholestatic pattern), elevated levels of CA 19.9  in serum (most sensitive tumor marker), and CT/MRCP imaging. • Intrahepatic cholangiocarcinoma will usually have an initial peripheral rim enhancement on arterial phase and progressive centripetal enhancement on delayed phase CT/MRI. • Diagnosis is mainly based on biopsy and histopathological confirmation. • Treatment of solitary intrahepatic cholangiocarcinoma is surgical resection—hepatic segmentectomy/ lobectomy. In case of unresectable tumors, chemoembolization can increase the median survival up to 12–15 months. • Primary treatment of perihilar and extrahepatic distal cholangiocarcinoma also involves surgical resection: partial hepatectomy or Whipple’s resection, respectively. • Treatment options for unresectable, locally advanced tumors includes chemotherapy with gemcitabine, radiotherapy, and targeted therapy with FDFR inhibitors.

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Histology Salient histological features: • Gross: 3 macroscopic types. –– Mass forming type (firm/hard tan mass). –– Periductal infiltrating type (similar to mass forming type but infiltrate along the ducts). –– Intraductal type (polypoidal within the duct). • Histologically the tumor cells are arranged in tubular and glands with cribriform, nest, solid, cords, or papillary pattern. • Mostly well differentiated with columnar to cuboidal epithelial cells with moderate amount of clear/slightly granular cytoplasm and small nuclei with inconspicuous nucleoli. • Poorly differentiated tumor is rarely seen. • Immunohistochemistry—CK7, CK19, CA19.9—Strongly positive. CK20—Variably positive, CDX-2—weak to moderate patchy nuclear staining in intestinal type of cholangiocarcinoma. Low power view: Cholangiocarcinoma with tumor cells arranged in glandular pattern (marked by arrow) surrounded by desmoplastic stroma (marked by star).

The following H&E stain shows tumor cells with round to oval, vesicular to hyperchromatic

10.4  Malignant Tumors

and mildly pleomorphic nuclei (marked by arrows), surrounded by dense desmoplastic stroma (marked by arrowhead).

IHC CK7: Scanner 40× magnification tumor cells show strong membranous positivity (marked by arrow).

10.4.2.2 Mixed HCC/ Cholangiocarcinoma • By definition, these tumors contain mixed cell types of both HCC and cholangiocarcinoma, based on both morphology and immunohistochemistry. • Clinical features are similar to that of HCC rather than cholangiocarcinoma. • Identification of Heppar 1, Arginase 1, or albumin ISH on morphology confirms the presence of hepatocellular component, whereas biliary component is confirmed by presence of discrete gland formation, desmoplastic stroma, and mucin production.

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• IHC for CK7, CK19, and EpCAM (MOC31) ae usually positive, whereas hepatocellular markers are negative. • Treatment of mixed tumors is quite challenging because of high frequency of nodal metastasis requiring lymph node dissection during surgical resection. • The presence of mixed components also eliminates the feasibility of liver transplantation, because of higher rates of recurrence. Histology The following image shows combined tumor— hepatocellular carcinoma and cholangiocarcinoma: Cores of tissue with tumor cells arranged in sheets (marked by arrows) and vague glandular pattern (marked by arrowhead).

The following H&E stain shows tumor cells with increased nuclear cytoplasmic ratio, moderate eosinophilic cytoplasm, mildly pleomorphic hyperchromatic to vesicular nuclei (marked by arrow).

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10.4.3 Mesenchymal Cell Origin

IHC CK19 high power 200× magnification: Combined hepatocellular carcinoma and cholangiocarcinoma: The tumor cells show strong membranous positivity—appears brown (marked by arrows).

10.4.3.1 Hemangioendothelioma • Rare, malignant vascular tumor of unclear etiology, comprising of dendritic or epithelioid cells, more commonly seen in females. • Clinical presentation: nonspecific symptoms of abdominal pain, nausea, vomiting, hepatomegaly. and rarely jaundice. • Slowly progressive when compared with angiosarcoma with prolonged survival. • Imaging characteristics: multiple lesions with or without calcifications. • Differentiation from HCC is mainly based on IHC staining for endothelial markers like CD31 and CD34. • Treatment: Surgical excision of the tumor or liver transplantation in case of unresectable tumors. Histology Salient histological features: • Histologically in the tumor cells are either epithelioid and dendritic cells or focal signet ring like cells with intracytoplasmic lumina with RBC. Background shows prominent myxoid to fibrous stroma. Ill-defined border merges into the adjoining normal hepatic parenchyma. • Immunohistochemistry—vascular markers— CD34, FLI-1, Factor VIII.

7)IHC Heppar 1 Low power 100× magnification shows combined hepatocellular carcinoma and cholangiocarcinoma: The tumor cells show strong granular cytoplasmic positivity—appears brown (marked by arrows).

The following image shows solid nests or pseudo-alveolar arrangement of epithelioid cells enmeshed in a fibrous stroma. Usually minimal mitotic activity, atypia, or necrosis.

10.4  Malignant Tumors

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This high power image shows solid nests or pseudo-alveolar arrangement (marked by arrow) of epithelioid cells enmeshed in a fibrous stroma (marked by star). Tumor cells have abundant, densely eosinophilic cytoplasm and can form vascular spaces.

IHC CD31: The tumor cells show strong cytoplasmic positivity—appears brown (marked by arrows).

The following image shows tumor cells with vesicular, round to oval, sometimes indented nuclei. Some tumor cells have intracytoplasmic, round, clear vacuoles representing small vascular lumina containing erythrocytes (marked by arrows).

IHC FLI 1: The tumor cells show strong nuclear positivity—appears as brown dots (marked by arrow).

IHC CD34: The tumor cells show strong cytoplasmic positivity—appears brown (marked by arrow).

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10.4.3.2 Gastrointestinal Stromal Tumor • Most common mesenchymal tumors of GI tract, composed of muscular and neural elements. • Commonly seen in stomach (60–70%), small intestine (20%) followed by esophagus, colon, and rectum. It may also occur in extraintestinal sites like omentum, pancreas, and retroperitoneum. • GIST lesions in liver are usually secondary to metastasis from primary GI sites and are often asymptomatic and detected on multidetector CT/somatostatin receptor scintigraphy. • Initial treatment option in metastatic GIST is usually with tyrosine kinase inhibitors like imatinib, sunitinib, or regorafenib, with very good response rates and often warrants life-­ long therapy. • Other treatment options include chemoembolization, radiofrequency ablation and in selected patients, debulking surgery may be attempted.

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The following high power image shows spindle cell type GIST: Short fascicles of bland spindle cells with syncytial pale eosinophilic cytoplasm, elongated nuclei, and inconspicuous nucleoli. Artifactual paranuclear vacuoles seen.

Histology Salient histological features: • Gross: 3 morphological subtypes: Spindle, epithelioid, and mixed. • Spindle—Bland spindle cells with faintly eosinophilic cytoplasm in a syncytial pattern. • Epithelioid round cells with clear to eosinophilic cytoplasm in sheets and nest. • Mixed: Both epithelioid and spindle type pattern will be seen. • Immunohistochemistry: CD117 and DOG-1. The following image showing liver parenchyma (marked by star) with adjacent gastrointestinal stromal tumor with spindled out areas in fascicles (marked by arrowheads).

Epithelioid GIST: Round cells with clear to eosinophilic cytoplasm in sheets.

10.4  Malignant Tumors

IHC C-KIT High power 200× magnification: The tumor cells show cytoplasmic positivity (marked by arrow).

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Histology Salient histological features: • Gross: 3 morphological subtypes: –– Well differentiated. –– Moderately differentiated. –– Poorly differentiated. • Well differentiated: well-formed glands/simple tubules with uniform basally oriented nuclei. • Moderately differentiated: complex tubules with loss in nuclear alignment. • Poorly differentiated, sheets of cells with less than 50% of glands.

10.4.3.3 Metastatic Tumors: Colorectal, Breast • Liver forms a major site of metastatic tumor deposits with nearly 40–50% incidence. • Hepatic metastasis commonly occurs from primary sites along the distribution of the portal venous system such as the pancreas, stomach, and colon, followed by lungs and breast. • The symptoms related to hepatic metastasis are usually masked by that of primary tumor, but some patients may present with jaundice, abdominal pain, and hepatomegaly. • Hepatic metastasis is usually multiple and detected better on multiphasic CT/MRI. • Prognosis depends on resectability of the primary tumor and extent of hepatic involvement by metastatic disease. • In patients with completely resectable primary malignancy, resection of metastatic lesions can be considered. • Other treatment options include chemoembolization, radioablation, and liver transplantation with/without chemotherapy for low grade neoplasms like neuroendocrine tumors.

The following image shows liver parenchyma with metastatic deposit of mucinous adenocarcinoma composed of clusters of tumor cells floating in large extracellular pools of mucin—marked by arrow (comprising more than 50% of tumor).

High power image shows signet ring cells with intracellular mucin and eccentrically pushed nuclei—marked by arrow.

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This image shows metastatic renal cell carcinoma: Liver parenchyma with tumor composed of compact nest and sheets of cells showing eosinophilic granular to vacuolated cytoplasm (marked by arrows).

The following image shows metastatic invasive ductal carcinoma of breast: Tubules, nests, and sheets of malignant cells with moderate nuclear pleomorphism (marked by arrows). Surrounding stroma shows desmoplasia (marked by star).

The following image shows metastatic small cell carcinoma of lung: The tumor cells (marked by arrows) are small to medium sized round to oval blue cells with high N:C ratio and scant cytoplasm.

10.4.3.4 Neuroendocrine Tumors • Arise from enterochromaffin cells: part of neuroendocrine system (neural crest cell origin). • Age of onset: third to sixth decade of life. • Most cases are sporadic, but may be associated with syndromes like multiple endocrine neoplasia 1 (MEN1), Von Hippel-Lindau syndrome (VHL), neurofibromatosis type 1 (NF1), tuberous sclerosis complex (TSC). • Most common site of NEC is the small intestine (25%), followed by the rectum (20%), appendix (17%), stomach (7%), pancreas, and colon. • Can be functional/non-functional tumors. • Hepatic NET: Usually secondary to metastasis from GI/pancreatic primary tumors. • Diagnosed based on somatostatin receptor uptake imaging like DOTANAC/DOTATATE. • Prognosis depends on the extent of metastasis and histological grading. • Treatment usually remains to be segmental resection if single or confined to one lobe along with resection of primary tumor. • Other treatment options include chemo/radio-­ embolization, radiofrequency ablation along with chemotherapy and long-acting somatostatin analogues.

10.4  Malignant Tumors

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Histology Salient histological features:

amount of cytoplasm (marked by arrows) and focal areas of necrosis (marked by star).

• Nested trabecular/microacinar pattern. • Tumor cells are small, uniform with granular salt and pepper chromatin. • Stromal hyalinization also seen. • Well differentiated neuroendocrine tumor graded according to mitosis and Ki-67 (mitotic proliferation index). –– Grade 1: 20%. • Neuroendocrine carcinoma based on morphology with high Ki67 index (usually more than 50%). –– Small cell neuroendocrine carcinoma. –– Large cell neuroendocrine carcinoma. • Immunohistochemistry: Synaptophysin, chromogranin, INSM1, Ki67.

The following image shows poorly differentiated neuroendocrine carcinoma: Sheets or nests of atypical cells with pleomorphic, hyperchromatic nuclei, and brisk mitosis (marked by arrow). “Salt and pepper” chromatin is lost. Necrosis often present.

This image shows cores of tissue with uniform, small, bland tumor cells in solid sheets (marked by arrows).

IHC synaptophysin: The tumor cells show strong cytoplasmic positivity—appears brown (marked by arrows). High power view of the same showing tumor cells with hyperchromatic nuclei and scant

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IHC chromogranin: High power 200× magnification: The tumor cells show cytoplasmic positivity—appears brown (marked by arrows).

10  Neoplasms of Liver

Liver Transplant Pathology

11.1 Acute Cellular Rejection • Acute cellular rejection (ACR) can be defined as T cell-mediated damage to the liver allograft. • The three components of the diagnostic triad for ACR are. –– Portal inflammation, –– Bile duct damage, and –– Venular endothelial inflammation.

Scores Portal inflammation

1 Lymphocytic infiltration involving few portal triads

Bile duct damage

Reactive atypia like high nuclear/cytoplasmic ratio and hyperchromasia and lymphocytic/neutrophilic ductulitis noted in few ducts Subendothelial lymphocytic infiltrate in few portal/central venules

Venular endothelial inflammation

11

Out of three, at least two are required for diagnosis of ACR. Based on the severity of portal inflammation, bile ductular damage, and venular endotheliitis, all three parameters are further given scores ranging from 1 to 3 as follows:

2 1+ marked expansion of portal tract with polymorphic infiltration with lymphocytes, neutrophils, and eosinophils Reactive atypia like high nuclear/ cytoplasmic ratio and hyperchromasia and lymphocytic/ neutrophilic ductulitis noted in most of the ducts Subendothelial lymphocytic infiltrate in most of the portal/ central venules

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 K. S. Mouleeswaran et al., Atlas of Basic Liver Histology for Practicing Clinicians and Pathologists, https://doi.org/10.1007/978-981-99-5762-0_11

3 2 + spillover of inflammatory infiltrates into interface (interface hepatitis) 2 + ductular disruption/ degeneration

2 + significant perivenular hepatocyte necrosis involving most of the perivenular region

101

11  Liver Transplant Pathology

102

The Rejection Activity Index (RAI) is calculated based on the scoring of all three parameters, portal, bile ductular, and venular endothelial inflammation, with minimum and maximum scores of 3 and 9, respectively. ACR is graded accordingly into • Mild (RAI 3–5). • Moderate (RAI 6–7). • Severe (RAI 8–9).

11.1.1 Portal Inflammation This image shows expansion of portal tract (marked by arrow) by inflammatory cells infiltrate without spillover of inflammatory cells into the surrounding hepatocytes.

This image shows portal tract with dense portal inflammation containing predominantly lymphocytes (marked by arrow) and scattered neutrophils and eosinophils (marked by ­arrowhead and circle, respectively). Usually, the inflammatory infiltrate in ACR is pleomorphic with predominance of lymphocytes as seen in this picture and occasionally accompanied by plasma cells.

11.1.2 Ductulitis This image shows high power view of portal tract containing mixed portal inflammation and ductulitis. This portal tract shows inflamed bile duct (BD) infiltrated by neutrophils (marked by arrow) and lymphocytes (marked by arrowhead). Duct epithelium shows degenerative changes including nucleomegaly and cytoplasmic vacuolation.

11.1.3 Venous Endothelial Inflammation 11.1.3.1 Central Perivenulitis This image shows central vein with single layer of flat endothelial cells. It is surrounded by mononuclear inflammatory cells. It also shows endothelial damage of central vein and few hepatocytes dropout (hepatocyte death seen as absence of hepatocyte) in zone 3.

11.2  Antibody Mediated Rejection

103

Acute AMR is diagnosed based on • Histopathological pattern of injury consistent with acute AMR. • Positive serum DSAs. • Presence of diffuse microvascular C4d deposition (C4d score = 3, described below). • Exclusion of other insults that might cause a similar pattern of graft injury including biliary obstruction and ascending cholangitis.

11.1.3.2 Portal Venulitis: Inflammation of Portal Vein This image shows portal venulitis—inflammed portal tract with portal vein in the center. There is subendothelial lymphoid cell infiltration with damage of endothelial lining and endothelial upliftment of portal vein suggestive of venous endothelial inflammation of portal vein.

Histopathological features that point to the presence of AMR are portal/periportal oedema, portal hemorrhage, neutrophil-rich inflammatory infiltration in the portal tracts and prominent ductular reaction, resembling those seen in biliary obstruction. Lobular parenchymal changes including cholestasis, spotty/confluent necrosis are more prominent than ACR, almost resembling sepsis, preservation reperfusion injury and obstructive biliary complications.

11.2.1 Portal Inflammation with Predominant Eosinophils A case of AMR. This image shows section of portal tract (high power view). Inflammatory cells are predominantly eosinophils(arrow) with scattered neutrophils and lymphocytes. There is portal venulitis with endothelial swelling, subendothelial lymphoid cell infiltration, and portal edema.

H & E Stain

11.2 Antibody Mediated Rejection Antibody mediated rejection is antibody and complement mediated damage of liver allograft and its consequences.

11  Liver Transplant Pathology

104

11.2.2 C4d Staining This image shows two portal tracts, both the portal tracts show strong circumferential C4d positivity in all the portal venules (marked by arrows).

This image shows high power view of the portal tract with strong linear C4d positivity involving the entire circumference of more than 50% of portal venules. Sometimes in AMR, periportal sinusoids may also show C4d positivity (not shown in this picture).

C4d staining of Central Vein. This image shows central vein with single layer of endothelium and C4d deposits.

C4d Score C4d 0 (negative) C4d 1 (minimal) C4d 2 (focal) C4d 3 (diffuse)

No C4d deposition in portal microvasculature 50% of portal tracts involved

C4d score of 3 is needed to make a definitive diagnosis of AMR in a recipient of an ABO compatible and incompatible graft which requires the presence of C4d staining in >50% of portal microvascular endothelia (portal veins, capillaries, often extending into portal inlet venules or periportal sinusoids) involving >50% of portal tract. Severe acute antibody mediated rejection can present with multifocal confluent hemorrhagic necrosis. The following images show multifocal confluent hemorrhagic necrosis with loss of hepatocytes and areas of parenchymal hemorrhage.

11.3  Chronic Rejection (CR)

105

•

•

•

•

•

recipient age, old donor age, and presence of high-titer DSAs of IgG3 subclass. Post-transplant factors include the severity and number of episodes of acute rejection, late presentation of acute rejection (>1  month post-transplant), cyclosporine use (versus tacrolimus), cytomegalovirus infection, HBV and HCV infection, and interferon therapy for viral hepatitis. Clinically, CR is characterized by progressive jaundice accompanied by cholestatic liver biochemistry. The transition from acute to chronic rejection may be associated with an elevation in aspartate transaminase (AST) levels. CR is a challenging complication to treat; it may respond to increased doses of tacrolimus, plasmapheresis, and intravenous immune globulin. When all other treatment fails, chronic rejection requires retransplantation.

11.3.1 Histology 11.3.1.1 Salient Histological Features • An immune mediated damage with potential irreversibly injury to bile ducts, arteries, and veins. CR is also called chronic ductopenic rejection, • Two main features: Dystrophic epithelial denotes immune mediated damage to the changes in the bile ducts with loss of bile duct allograft to cause irreversible injury and even(ductopenia) and obliterative vasculopathy of tual loss of bile ducts in portal tracts. medium and large sized arteries (not seen in Although it is called chronic it can happen core biopsies). within months of liver transplantation. • Bile duct loss is the most important feature in The incidence of CR is declining, because of core needle biopsies as it is also called ductomore effective immunosuppression and now penic rejection. results in graft failure in 2 g/dL)

12.3 Differentiating Features Between Granulomatous Hepatitis S. no. 1.

Tuberculosis Granulomas—well defined

2. 3. 4.

Location: both portal tract (PT) and hepatic parenchyma (HP) Necrosis (+) or (−) Gene-Xpert or PCR (+ve)

5. 6. 7.

Langhans type giant cell AFB (+) or (−) Fibrosis—focal

Sarcoidosis Well defined, but large plump epithelioid histiocytes Usually PT

Fungal Ill-­defined

Mostly (−) Schaumann bodies and asteroid bodies Calcium stain von kossa Iron’s stain perls—PBC Fibrosis (+) portal based fibrosis even leads to cirrhosis

(+) or (−) Foreign body type and giant cell PAS/GMS for fungus Not related

Usually PT

12.4 Differences Between AIH and Viral Hepatitis S. no. 1. 2.

3. 4.

5.

6.

Autoimmune hepatitis Significant plasma cells present in portal tracts Significant portal inflammation with moderate to severe interface hepatitis

Hepatitis A virus—acute hepatitis Variable

Significant perivenular necrosis with confluent/bridging necrosis Generally, no significant hepatocanalicular cholestasis including no perivenular cholestasis Fatty change: Usually absent except in atypical variant

Absence of perivenular necrosis

Interface hepatitis +++

More of periportal inflammation and interface hepatitis

HCV (chronic hepatitis) Variable plasma cells in portal tracts Usually moderate portal inflammation with variable interface hepatitis Perivenular spotty and confluent necrosis +/−

Perivenular cholestasis significant

Fatty change: Rare—Fulminant hepatitis However ballooning degeneration, multinucleated giant cells, microvesicular fatty change and panlobular submassive necrosis Interface hepatitis +

Fatty change: Macrovesicular fatty change common

Variable

115

12.5 Chronic Hepatitis: Differential Diagnosis

12.5 Chronic Hepatitis: Differential Diagnosis S. no. 1.

Features Serology

HCV HCV PCR (RNA), anti HCV antibodies

AIH Negative

Wilson Negative

Drug induced Negative

Increased (variable to more than 10 times) Variable Normal

Increased always >5 times than normal >3 g/dL Normal

Increased

Increased

3. 4.

Enzyme AST, ALT—Hepatotrophic enzyme raise compared to ALP and GGT Serum gamma globulin Ceruloplasmin

HBV IgG, HbsAg with hepatitis core antigen, Anti HBC, HBV PCR Others HDV–RNA, IgM AntiHBD, HEV–RNA, IgM AntiHBE Increased (variable to more than 10 times) Variable Normal

2.

Normal

5.

Urinary copper

Nil

Nil

Nil

Low (D/D can be seen in primary biliary cholangitis, primary sclerosing cholangitis) Urinary copper increased D/D (can be seen in primary biliary cholangitis, primary sclerosing cholangitis)

Histopathological findings 6. Plasma cells

Seen

Seen

Seen

Seen

7.

Periportal spotty necrosis and lobular necrosis Fatty change

Less likely

Less likely

Seen with periportal increase in plasma cells Seen

Seen

Variable

Variable to less common in HBV

Fatty change variable

Common

Fatty change uncommon except in atypical ones Uncommon

Fatty change variable

Hepatocanalicular bilirubinostasis Mallory Denk bodies and neutrophilic satellitosis of hepatocytes Increased copper in early stage of disease (before cirrhotic stage)

Fatty change in HCV– Geno3 Common

Variable

Variable

Not seen

Not seen

Not seen

Seen

Variable

Not seen

Not seen

Not seen

Seen

Variable

8.

9. 10.

11.

Nil

12  Classical Histological Differences: Comparison Tables

116 S. no. 12.

13.

Features Hepatocyte rosetting/ severe interface hepatitis (+/−) emperipolesis Autoimmune markers (in rare cases, autoimmune markers can be seen in NASH, drug induced and viral hepatitis)  

HBV Not seen

HCV Not seen

AIH Common

Wilson Not seen

Drug induced Not seen

Rare

Rare

Positive for ANA, LKM, SMA

Variable

Variable ANA— Commonly seen