Oxford Textbook of Clinical Nephrology [4 ed.] 9780199592548, 0199592548

Table of contents :
Cover......Page 1
Oxford Textbook of Clinical Nephrology......Page 4
Copyright......Page 5
Summary of Contents......Page 8
SECTION 1: Assessment of the patient with renal disease......Page 26
1 Epidemiology of kidney disease......Page 28
2 Clinical assessment of the patient with renal disease: overview......Page 45
3 Presentations of renal disease......Page 47
4 Kidney disease-focused history taking......Page 52
5 Kidney disease-focused features on examination......Page 54
6 Urinalysis......Page 60
7 Assessment of renal function......Page 69
8 Tubular function......Page 87
9 Renal radiology: overview......Page 91
10 Ionizing radiation and radiation
protection......Page 93
11 Plain radiography, excretion radiography, and contrast radiography......Page 99
12 Intervention......Page 107
13 Ultrasound......Page 114
14 Computed tomography......Page 126
15 Magnetic resonance imaging......Page 134
16 Radioisotopes in diagnostic imaging in nephrology......Page 142
17 Immunological investigation of the patient with renal disease......Page 152
18 The renal biopsy......Page 167
19 Clinical trials: why and how in nephrology......Page 186
SECTION 2: The Patient with fluid, electrolyte, and renal tubular disorders......Page 194
20 An overview of tubular function......Page 196
21 Sodium transport and balance: a key role for the distal nephron......Page 214
22 Water homeostasis......Page 222
23 Potassium homeostasis......Page 229
24 Renal acid–base homeostasis......Page 243
25 Phosphate homeostasis......Page 250
26 Calcium homeostasis......Page 256
27 Magnesium homeostasis......Page 268
28 Approach to the patient with hyponatraemia......Page 274
29 Approach to the patient with hypernatraemia......Page 286
30 Approach to the patient with oedema......Page 297
31 Approach to the patient with salt-wasting tubulopathies......Page 307
32 Approach to the patient with polyuria......Page 316
33 Clinical use of diuretics......Page 324
34 Approach to the patient with hypo-/hyperkalaemia......Page 348
35 Approach to the patient with metabolic acidosis or alkalosis......Page 364
36 Approach to the patient with renal tubular acidosis......Page 388
37 Approach to the patient with hypercalcaemia......Page 397
38 Approach to the patient with
hypocalcaemia......Page 403
39 Approach to the patient with hypo-/hyperphosphataemia......Page 409
40 Approach to the patient with hypomagnesaemia......Page 422
41 Approach to the patient with renal Fanconi syndrome, glycosuria, or aminoaciduria......Page 437
SECTION 3: The patient with glomerular disease......Page 448
42 The glomerulus and the concept of glomerulonephritis......Page 450
43 The renal glomerulus: the structural basis of ultrafiltration......Page 461
44 Function of the normal glomerulus......Page 476
45 Mechanisms of glomerular injury: overview......Page 484
46 The patient with haematuria......Page 488
47 Loin pain haematuria syndrome......Page 494
48 Nutcracker syndrome and phenomenon......Page 498
49 Exercise-related pseudonephritis......Page 501
50 Proteinuria......Page 503
51 Postural proteinuria (benign orthostatic proteinuria)......Page 510
52 Nephrotic syndrome......Page 512
53 Pathophysiology of oedema in nephrotic syndrome......Page 521
54 Idiopathic nephrotic syndrome: overview......Page 524
55 Minimal change disease: clinical features and diagnosis......Page 526
56 Minimal change disease: treatment and outcome......Page 531
57 Primary focal segmental glomerulosclerosis: clinical features and diagnosis......Page 540
58 Primary focal segmental glomerulosclerosis: treatment and outcome......Page 550
59 Pathogenesis of proteinuria in minimal change disease and focal segmental glomerulosclerosis......Page 558
60 Membranous glomerulonephritis: overview......Page 562
61 Membranous glomerulonephritis: clinical features and diagnosis......Page 564
62 Membranous glomerulonephritis: treatment and outcome......Page 569
63 Secondary membranous glomerulonephritis......Page 582
64 Membranous glomerulonephritis: pathogenesis......Page 585
65 Immunoglobulin A nephropathy: overview......Page 590
66 Immunoglobulin A nephropathy: clinical features......Page 591
67 Immunoglobulin A nephropathy: diagnosis......Page 597
68 Immunoglobulin A nephropathy: treatment and outcome......Page 602
69 Immunoglobulin A nephropathy: pathogenesis......Page 611
70 Crescentic (rapidly progressive) glomerulonephritis......Page 617
71 Antiglomerular basement membrane disease: overview......Page 623
72 Antiglomerular basement membrane disease: clinical features and diagnosis......Page 624
73 Antiglomerular basement membrane disease: treatment and outcome......Page 631
74 Antiglomerular basement membrane disease: pathogenesis......Page 634
75 Alport post-transplant antiglomerular basement membrane disease......Page 644
76 Post-infectious glomerulonephritis: overview......Page 646
77 Post-streptococcal glomerulonephritis......Page 648
78 Immunoglobulin A-dominant post-infectious glomerulonephritis......Page 658
79 Glomerulonephritis associated with endocarditis, deep-seated infections, and shunt nephritis......Page 661
80 Membranoproliferative glomerulonephritis and C3 glomerulopathy......Page 666
81 Fibrillary and immunotactoid glomerulopathy......Page 674
82 Drug-induced and toxic glomerulopathies......Page 681
SECTION 4: The patient with interstitial disease......Page 692
83 Acute tubulointerstitial nephritis: overview......Page 694
84 Drug-induced acute tubulointerstitial nephritis......Page 703
85 Other toxic acute tubulointerstitial
nephritis......Page 712
86 Chronic tubulointerstitial nephritis: overview......Page 715
87 Drug-induced chronic tubulointerstitial nephritis......Page 720
88 Heavy metal-induced tubulointerstitial nephritis......Page 727
89 Aristolochic acid nephropathy caused by ingestion of herbal medicinal products......Page 734
90 Balkan endemic nephropathy......Page 739
91 Radiation nephropathy......Page 744
92 Urate nephropathy......Page 749
93 Immune-mediated tubulointerstitial nephritis......Page 751
SECTION 5: The patient with reduced renal function......Page 764
94 Chronic kidney disease: definition, classification, and approach to management......Page 768
95 Chronic kidney disease in the developed world......Page 780
96 Chronic kidney disease in developing countries......Page 787
97 Chronic kidney disease long-term outcomes: progression, death, cardiovascular disease, infections, and hospitalizations......Page 792
98 Cardiovascular disease and chronic kidney disease: overview......Page 802
99 Recommendations for management of high renal risk chronic kidney disease......Page 805
100 Hypertension as a cause of chronic kidney disease: what is the evidence?......Page 812
101 Diet and the progression of chronic kidney disease......Page 818
102 Lipid disorders of patients with chronic kidney disease......Page 825
103 Smoking in chronic kidney disease......Page 832
104 Analytical aspects of measurements and laboratory values in kidney disease......Page 838
105 Effect of lifestyle modifications on patients with chronic kidney disease......Page 849
106 Malnutrition, obesity, and undernutrition in chronic kidney disease......Page 855
107 Left ventricular hypertrophy in chronic kidney disease......Page 862
108 Sudden cardiac death in chronic kidney disease......Page 878
109 Epidemiology of calcium, phosphate, and parathyroid hormone disturbances in chronic kidney disease......Page 894
110 The role of inflammation in chronic kidney disease......Page 902
111 Vascular stiffness in chronic kidney disease: pathophysiology and implications......Page 909
112 Oxidative stress and its implications in chronic kidney disease......Page 920
113 Abnormal endothelial vasomotor and secretory function......Page 928
114 Endothelins and their antagonists in chronic kidney disease......Page 935
115 Chronic kidney disease-mineral and bone disorder: overview......Page 941
116 Imaging for detection of vascular disease in chronic kidney disease patients......Page 948
117 Pathophysiology of chronic kidney disease-mineral and bone disorder......Page 959
118 Management of chronic kidney disease-mineral and bone disorder......Page 964
119 Fibroblast growth factor 23, Klotho, and phosphorus metabolism in chronic kidney disease......Page 972
120 Vascular calcification......Page 982
121 Fractures in patients with chronic kidney disease......Page 995
122 Spectrum of bone pathologies in chronic kidney disease......Page 1001
123 Clinical aspects and overview of renal anaemia......Page 1008
124 Erythropoiesis-stimulating agents in chronic kidney disease......Page 1016
125 Iron metabolism in chronic kidney
disease......Page 1023
126 Iron management in renal anaemia......Page 1035
127 Pleiotropic effects of vitamin D......Page 1041
128 Immunity......Page 1063
129 The epidemiology of hepatitis viruses in chronic kidney disease......Page 1074
130 Gastroenterology and renal medicine......Page 1077
131 Cutaneous manifestations of end-stage renal disease......Page 1089
132 The patient with reduced renal function: endocrinology......Page 1097
133 Sexual dysfunction......Page 1116
134 Health-related quality of life and the patient with chronic kidney disease......Page 1124
135 Coagulopathies in chronic kidney disease......Page 1127
136 Mechanisms of progression of chronic kidney disease: overview......Page 1130
137 Proteinuria as a direct cause of progression......Page 1132
138 Nephron numbers and hyperfiltration as drivers of progression......Page 1137
139 Podocyte loss as a common pathway to chronic kidney disease......Page 1143
140 Disordered scarring and failure of repair......Page 1151
141 Modality selection for renal replacement therapy......Page 1161
142 Patient education and involvement in pre-dialysis management......Page 1167
143 Preparation for renal replacement
therapy......Page 1173
144 Choices and considerations for
in-centre versus home-based renal replacement therapy......Page 1182
145 Conservative care in advanced chronic kidney disease......Page 1190
146 Palliative care in end-stage renal disease......Page 1205
147 Patient selection when resources are limited......Page 1212
148 Acidosis in chronic kidney disease......Page 1217
SECTION 6: The patient with another primary diagnosis......Page 1222
149 The patient with diabetes mellitus......Page 1224
150 Kidney involvement in plasma
cell dyscrasias......Page 1273
151 The patient with cryoglobulinaemia......Page 1276
152 The patient with amyloidosis......Page 1289
153 The patient with myeloma......Page 1301
154 Light-chain deposition disease......Page 1317
155 Other consequences from monoclonal immunoglobulins/fragments: membranoproliferative glomerulonephritis and acquired Fanconi syndrome......Page 1324
156 The patient with sarcoidosis......Page 1329
157 The patient with vasculitis: overview......Page 1332
158 The patient with vasculitis: pathogenesis......Page 1340
159 The patient with vasculitis: clinical aspects......Page 1357
160 The patient with vasculitis: treatment and outcome......Page 1369
161 The patient with systemic lupus erythematosus: overview and
pathogenesis......Page 1381
162 The patient with systemic lupus erythematosus: clinical features, investigations, and diagnosis......Page 1390
163 The patient with systemic lupus erythematosus: treatment and outcome......Page 1405
164 The patient with antiphospholipid syndrome with or without lupus......Page 1414
165 The patient with scleroderma: systemic sclerosis......Page 1420
166 The patient with rheumatoid arthritis, mixed connective tissue disease, Sjögren syndrome, or polymyositis......Page 1428
167 The patient with sickle cell anaemia......Page 1438
168 The obese patient (metabolic
syndrome)......Page 1449
169 The patient with hepatorenal syndrome......Page 1458
170 Kidney/ear syndromes......Page 1467
171 Kidney/eye syndromes......Page 1475
172 The patient with renal cell cancer......Page 1482
173 The patient with Wilms tumour......Page 1489
174 The patient with haemolytic uraemic syndrome/thrombotic thrombocytopenic purpura......Page 1494
SECTION 7: The patient with urinary tract infection......Page 1516
175 Urinary tract infection in the adult: overview......Page 1518
176 Infection of the lower urinary tract......Page 1520
177 Upper urinary tract infection......Page 1531
178 Complicated urinary tract infection......Page 1539
179 Urinary tract infection in a patient with a kidney transplant......Page 1542
180 Urinary tract infection in infancy and childhood......Page 1545
181 Schistosomiasis: the parasite and the host......Page 1557
182 Schistosomiasis: clinical impact......Page 1563
SECTION 8: The patient with infections causing renal disease......Page 1568
183 Malaria......Page 1570
184 Leishmaniasis and trypanosomiasis......Page 1575
185 Hepatitis B......Page 1577
186 Hepatitis C......Page 1585
187 HIV and renal disease......Page 1592
188 Hantaviral infections......Page 1601
189 Dengue and other viral haemorrhagic fevers......Page 1606
190 Yellow fever, severe acute respiratory
syndrome virus, and H1N1 influenza infections......Page 1609
191 Leptospirosis......Page 1612
192 Syphilis......Page 1615
193 Rickettsiosis......Page 1617
194 Schistosomiasis......Page 1621
195 Nematode infections......Page 1627
196 Mycobacterial infections: tuberculosis......Page 1629
197 Mycobacterial infections: leprosy and environmental mycobacteria......Page 1645
198 Renal involvement in other infections: diarrhoeal diseases, salmonella, melioidosis, and pregnancy......Page 1650
SECTION 9: The patient with urinary stone disease......Page 1654
199 Epidemiology of nephrolithiasis......Page 1656
200 Approach to the patient with
kidney stones......Page 1662
201 Calcium stones......Page 1670
202 Uric acid stones......Page 1686
203 Cystine stones......Page 1691
204 Cell biology of nephrocalcinosis/nephrolithiasis......Page 1696
205 Medical management of nephrocalcinosis and nephrolithiasis......Page 1722
206 Imaging and interventional treatment: urolithiasis from the surgeon’s point of view......Page 1738
SECTION 10: The Patient with Hypertension......Page 1748
207 The structure and function of renal blood vessels......Page 1750
208 Regulation of vasomotor tone in the afferent and efferent arterioles......Page 1754
209 Tubuloglomerular feedback, renal autoregulation, and renal protection......Page 1763
210 The kidney and control of
blood pressure......Page 1767
211 The effect of hypertension on renal vasculature and structure......Page 1775
212 Ischaemic nephropathy......Page 1785
213 Renal artery stenosis: clinical features and diagnosis......Page 1791
214 Renal artery stenosis: diagnosis......Page 1801
215 Renal artery stenosis: management and outcome......Page 1807
216 Malignant hypertension......Page 1817
217 Resistant hypertension......Page 1824
218 The hypertensive child......Page 1827
219 Treatment of hypertension in children......Page 1850
SECTION 11: The patient with acute kidney injury (and critical care nephrology)......Page 1854
220 Definitions, classification, epidemiology, and risk factors of acute kidney injury......Page 1856
221 Pathophysiology of acute kidney injury......Page 1869
222 Clinical approach to the patient with acute kidney injury: diagnosis and differential diagnosis......Page 1897
223 The role of novel biomarkers in acute kidney injury......Page 1912
224 Prevention of acute kidney injury: overview......Page 1921
225 Prevention of acute kidney injury: non-pharmacological strategies......Page 1927
226 Prevention of acute kidney injury: pharmacological strategies......Page 1940
227 Prevention of acute kidney injury:
drug- and nephrotoxin-induced acute kidney injury......Page 1950
228 Non-dialytic management of the patient with acute kidney injury......Page 1958
229 Fluid overload in acute kidney injury......Page 1966
230 Electrolyte and acid–base disorders in AKI......Page 1970
231 Coagulation disturbances in acute
kidney injury......Page 1979
232 Renal replacement therapy in the patient with acute kidney injury: overview......Page 1981
233 Intermittent acute renal replacement therapy......Page 1987
234 Continuous renal replacement therapy......Page 2000
235 Peritoneal dialysis in acute kidney injury......Page 2014
236 Scoring systems in acute kidney injury patients......Page 2023
237 Overall outcomes of acute kidney injury......Page 2034
238 Renal outcomes of acute kidney injury......Page 2042
239 Acute kidney injury in children......Page 2049
240 Acute kidney injury in the elderly......Page 2063
241 Acute kidney injury in the tropics......Page 2072
242 Acute kidney injury and hantavirus disease......Page 2084
243 Community-acquired pneumonia and acute kidney injury......Page 2092
244 Acute kidney injury in severe sepsis......Page 2093
245 Cardiovascular surgery and acute kidney injury......Page 2101
246 Contrast-induced acute kidney injury......Page 2109
247 Renal failure in cirrhosis: pathogenesis, diagnosis, and treatment......Page 2116
248 Acute kidney injury in heart failure......Page 2134
249 Acute kidney injury in pulmonary
diseases......Page 2140
250 Acute kidney injury in pregnancy......Page 2154
251 Acute kidney injury in the
cancer patient......Page 2158
252 Acute kidney injury in polytrauma and rhabdomyolysis......Page 2168
253 Acute kidney injury in patients with severe burn injury......Page 2176
SECTION 12: The patient on dialysis......Page 2184
254 Uraemic toxins: overview......Page 2186
255 Haemodialysis: overview......Page 2198
256 Haemodialysis: vascular access......Page 2201
257 Haemodialysis: principles......Page 0
258 Haemodialysis: prescription
and assessment of adequacy......Page 2213
259 Haemodialysis: acute complications......Page 2232
260 Haemofiltration and haemodiafiltration......Page 2243
261 Dialysis withdrawal and palliative care......Page 2252
262 Frequent haemodialysis......Page 2259
263 Peritoneal dialysis: overview......Page 2274
264 Peritoneal dialysis: principles
and peritoneal physiology......Page 2277
265 Peritoneal dialysis: adequacy
and prescription management......Page 2288
266 Peritoneal dialysis: non-infectious complications......Page 2295
267 Overview of dialysis patient management and future directions......Page 2304
268 Cardiovascular complications in end-stage renal disease patients: pathophysiological aspects......Page 2310
269 Bacterial and fungal infections in patients on haemodialysis......Page 2318
270 Bacterial and fungal infections in patients on peritoneal dialysis......Page 2324
271 Viral infections in patients on dialysis......Page 2328
272 Cognitive function, depression,
and psychosocial adaptation......Page 2330
273 Volume assessment
and management in dialysis......Page 2338
274 Nutritional screening and nutritional management in dialysis patients......Page 2348
SECTION 13: The transplant patient......Page 2357
275 The evolution of kidney transplantation......Page 2359
276 Pre-transplant assessment
of the recipient......Page 2372
277 Organ donation......Page 2380
278 Donor and recipient kidney transplantation surgery......Page 2392
279 Immunology, sensitization,
and histocompatibility......Page 2404
280 Immediate post-transplant care
and surgical complications......Page 2419
281 Immunosuppression: drugs
and protocols......Page 2427
282 Renal transplant imaging......Page 2440
283 Rejection......Page 2455
284 Infection: prophylaxis, diagnosis, and management......Page 2467
285 Cardiovascular disease: prophylaxis, diagnosis, and management......Page 2477
286 Chronic allograft dysfunction......Page 2485
287 Cancer after kidney transplantation......Page 2497
288 Metabolic bone disease after renal transplantation......Page 2505
289 Recurrent renal disease: prophylaxis, diagnosis, and management......Page 2516
290 Paediatric renal transplantation......Page 2523
SECTION 14: Renal disease at different stages of life (infancy, adolescence, pregnancy, old age)......Page 2533
291 Growth and development......Page 2535
292 The adolescent with renal disease: transition to adult services......Page 2541
293 Contraception in patients
with kidney disease......Page 2544
294 Pregnancy and renal physiology......Page 2551
295 Pregnancy in patients with chronic kidney disease and on dialysis......Page 2563
296 Pre-eclampsia and related disorders......Page 2570
297 Acute kidney injury in pregnancy......Page 2580
298 Specific renal conditions in pregnancy......Page 2584
299 Pregnancy after renal transplantation......Page 2591
300 The kidney in ageing: biology, anatomy, physiology, and clinical relevance......Page 2594
SECTION 15: The patient with genetic renal disease......Page 2603
301 Ethical aspects of genetic testing......Page 2605
302 Antenatal diagnosis and pre-implantation genetic testing......Page 2608
303 The molecular basis of ciliopathies and cyst formation......Page 2611
304 The adult with renal cysts......Page 2631
305 The child with renal cysts......Page 2635
306 Autosomal dominant polycystic kidney disease: overview......Page 2639
307 Autosomal dominant polycystic kidney disease: clinical features......Page 2641
308 Autosomal dominant polycystic kidney disease: diagnosis......Page 2648
309 Autosomal dominant polycystic kidney disease: management......Page 2652
310 Management of intracranial
aneurysms......Page 2661
311 Management of cystic liver disease......Page 2663
312 Autosomal dominant polycystic
kidney disease in children
and young adults......Page 2667
313 Autosomal recessive polycystic kidney disease......Page 2670
314 Bardet–Biedl syndrome
and other ciliopathies......Page 2679
315 Hepatocyte nuclear factor-1B......Page 2685
316 Nephronophthisis and medullary cystic kidney disease: overview......Page 2688
317 Nephronophthisis......Page 2691
318 Autosomal dominant interstitial
kidney disease including
medullary cystic disease......Page 2696
319 Oral-facial-digital type 1 syndrome......Page 2699
320 The molecular basis of glomerular basement membrane disorders......Page 2702
321 Alport syndrome: overview......Page 2709
322 Alport syndrome: clinical features......Page 2711
323 Alport syndrome: diagnosis......Page 2714
324 Alport syndrome: management......Page 2720
325 Thin glomerular basement membrane nephropathy and other collagenopathies......Page 2723
326 Nail patella syndrome......Page 2725
327 Molecular basis of nephrotic syndrome......Page 2728
328 Molecular basis of renal tumour syndromes......Page 2736
329 WT1 and its disorders......Page 2741
330 Tuberous sclerosis complex renal disease......Page 2746
331 Hypoxia-inducible factor
and renal disorders......Page 2753
332 Von Hippel–Lindau disease......Page 2757
333 Molecular basis of complement-mediated renal disease......Page 2761
334 Inherited metabolic diseases and the kidney......Page 2771
335 Fabry disease: overview
and pathophysiology......Page 2781
336 Fabry disease: clinical features......Page 2784
337 Fabry disease: diagnosis......Page 2790
338 Fabry disease: management and outcome......Page 2794
339 Cystinosis......Page 2803
340 Mitochondrial diseases and the kidney......Page 2812
341 APOL1 and renal disease......Page 2822
342 MYH9 and renal disease......Page 2825
SECTION 16: The patient with structural and congenital abnormalities......Page 2827
343 Human kidney development......Page 2829
344 Kidney stem cells......Page 2836
345 Anatomical types of congenital anomalies: overview of obstruction......Page 2842
346 Renal agenesis......Page 2844
347 Renal dysplasia......Page 2845
348 Renal hypoplasia......Page 2846
349 Normal variation in nephron numbers......Page 2847
350 Renal tubular dysgenesis......Page 2849
351 Congenital solitary functioning kidney......Page 2850
352 Duplex, ectopic, and horseshoe
kidneys......Page 2852
353 Pelviureteric junction obstruction and megaureter in children......Page 2854
354 Posterior urethral valves......Page 2856
355 Primary vesicoureteric reflux and reflux nephropathy......Page 2858
356 The patient with urinary tract obstruction......Page 2868
357 Retroperitoneal fibrosis......Page 2875
358 Branchio-oto-renal syndrome......Page 2879
359 Townes–Brocks syndrome......Page 2881
360 Renal coloboma syndrome......Page 2883
361 Ante- and postnatal imaging to diagnose human kidney malformations......Page 2885
SECTION 17: Drugs and renal disease......Page 2897
362 Drug-induced nephropathies......Page 2899
363 Drug dosing in chronic kidney disease......Page 2925
364 Drug dosing in acute kidney injury......Page 2933
SECTION 18: Nephrology in the future......Page 2939
365 A global curriculum for training the next generation of nephrologists......Page 2941
A......Page 2947
B......Page 2954
C......Page 2956
D......Page 2964
E......Page 2967
F......Page 2969
G......Page 2971
H......Page 2973
I......Page 2978
K......Page 2981
L......Page 2982
M......Page 2984
N......Page 2987
O......Page 2990
P......Page 2991
R......Page 2997
S......Page 3001
T......Page 3004
U......Page 3008
V......Page 3010
Z......Page 3012

Citation preview

Oxford Textbook of

Clinical Nephrology

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VOLUME 1

Oxford Textbook of

Clinical Nephrology FOURTH EDITION Edited by

Managing Editors Neil Turner Norbert Lameire David J. Goldsmith Christopher G. Winearls Jonathan Himmelfarb Giuseppe Remuzzi

1

Section Editors William G. Bennett Jeremy R. Chapman Adrian Covic Marc E. De Broe Vivekanand Jha Neil Sheerin Robert Unwin Adrian Woolf

1 Great Clarendon Street, Oxford, OX2 6DP, United Kingdom Oxford University Press is a department of the University of Oxford. It furthers the University’s objective of excellence in research, scholarship, and education by publishing worldwide. Oxford is a registered trade mark of Oxford University Press in the UK and in certain other countries © Oxford University Press 2016 The moral rights of the authors‌have been asserted First Edition Published in 1992 Second Edition Published in 1998 Third Edition Published in 2005 Fourth Edition Published in 2016 Impression: 1 All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted, in any form or by any means, without the prior permission in writing of Oxford University Press, or as expressly permitted by law, by licence or under terms agreed with the appropriate reprographics rights organization. Enquiries concerning reproduction outside the scope of the above should be sent to the Rights Department, Oxford University Press, at the address above You must not circulate this work in any other form and you must impose this same condition on any acquirer Published in the United States of America by Oxford University Press 198 Madison Avenue, New York, NY 10016, United States of America British Library Cataloguing in Publication Data Data available Library of Congress Control Number: 2015938370 ISBN 978–0–19–870858–2 (volume 1) 978–0–19–870859–9 (volume 2) 978–0–19–870860–5 (volume 3) 978–0–19–959254–8 (set) Printed and bound in Great Britain by Bell & Bain, Glasgow. Oxford University Press makes no representation, express or implied, that the drug dosages in this book are correct. Readers must therefore always check the product information and clinical procedures with the most up-to-date published product information and data sheets provided by the manufacturers and the most recent codes of conduct and safety regulations. The authors and the publishers do not accept responsibility or legal liability for any errors in the text or for the misuse or misapplication of material in this work. Except where otherwise stated, drug dosages and recommendations are for the non-pregnant adult who is not breast-feeding Links to third party websites are provided by Oxford in good faith and for information only. Oxford disclaims any responsibility for the materials contained in any third party website referenced in this work.

Preface

We have almost completely rewritten the Oxford Textbook of Clinical Nephrology for its fourth edition. That was a big decision for a very successful book, but there were several important drivers. Huge developments in nephrology and medicine more broadly were only part of the reasoning. First, we wanted a text that would adapt easily to presentation in multiple formats—on paper and also electronically on different devices. Shorter (but more) chapters seemed an important part of this, but meant reviewing the organization of the entire book. However, this also helped us to meet our second objective, which was to make it easier to get quick answers to specific questions. And third, we wanted a structure that would aid updating in the future, both on paper and electronically.

Our authorship and editorship are substantially changed and even more international than before, with representation from every continent except Antarctica. Our coverage of global topics is substantially enhanced. There is an entirely new section on genetic diseases, and substantially increased coverage for the patient with a renal transplant. We have completely revised and combined previous sections on tubular disorders and electrolytes, achieving greater clarity and reduced duplication. There is new material on renal disease in childhood and old age, together with a completely new set of chapters on renal disease in pregnancy. The book is no longer but we believe it is substantially enhanced. This has been a huge project for the editors, project managers, and production team—but ultimately, a very rewarding one. We hope you will agree that it has been worthwhile.

Summary of Contents

VOLUME 1 SECTION 1

Assessment of the patient with renal disease  1 Edited by Christopher G. Winearls

SECTION 2

The patient with fluid, electrolyte, and renal tubular disorders  169 Edited by Robert Unwin and Pascal Houillier

SECTION 3

The patient with glomerular disease  423 Edited by Neil Turner

SECTION 4

The patient with interstitial disease  667 Edited by Adrian Covic

SECTION 5

The patient with reduced renal function  739 Edited by David J. Goldsmith

VOLUME 2 SECTION 6

The patient with another primary diagnosis  1197 Edited by Giuseppe Remuzzi

SECTION 7

The patient with urinary tract infection  1491 Edited by Neil Sheerin

SECTION 8

The patient with infections causing renal disease  1543 Edited by Vivekanand Jha

SECTION 9

The patient with urinary stone disease  1629 Edited by Marc E. De Broe

SECTION 10

The patient with hypertension  1723 Edited by Neil Turner

SECTION 11

The patient with acute kidney injury (and critical care nephrology)  1829 Edited by Norbert Lameire

VOLUME 3 SECTION 12

The patient on dialysis  2159 Edited by Jonathan Himmelfarb

viii

summary of contents

SECTION 13

SECTION 16

The transplant patient  2343

The patient with structural and congenital abnormalities  2813

Edited by Jeremy R. Chapman

Edited by Adrian Woolf

SECTION 14

Renal disease at different stages of life (infancy, adolescence, pregnancy, old age)  2519

SECTION 17

Drugs and renal disease  2883 Edited by William G. Bennett

Edited by Norbert Lameire and Neil Turner

SECTION 15

The patient with genetic renal disease  2589 Edited by Neil Turner

SECTION 18

Nephrology in the future  2925 Edited by Neil Turner

Contributors

Dwomoa Adu Department of Medicine, University of Ghana Medical School, University of Ghana, Accra, Ghana

Vicente Arroyo Liver Unit, Hospital Clinic, University of Barcelona, Barcelona, Spain

Behdad Afzali Medical Research Council Centre for Transplantation, King’s College London, London; NIHR Biomedical Research Centre, Guy’s & St Thomas’ NHS Foundation Trust, London; King’s College London, London, UK; and Lymphocyte Cell Biology Section, Molecular Immunology and Inflammation Branch, National Institutes of Arthritis, and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD, USA

Carla Maria Avesani Department of Applied Nutrition, Nutrition Institute, Rio de Janeiro State University, Rio de Janeiro, Brazil

Rajiv Agarwal Division of Nephrology, Department of Medicine, Indiana University School of Medicine, Indiana University; and Veterans Administration Medical Center, Indianapolis, IN, USA Tomas Thor Agustsson Department of Endocrinology and Metabolic Medicine, Landspitali; and The National University Hospital of Iceland, Reykjavik, Iceland Bassam Alchi Renal Unit, Addenbrooke’s Hospital, Cambridge, UK Helen Alderson Institute of Inflammation and Repair, University of Manchester, Manchester, UK

Fred E. Avni Department of Medical Imaging, University Clinics of Brussels—Erasme Hospital, Brussels, Belgium Seema Baid-Agrawal Division of Nephrology and Medical Intensive Care, Department of Medicine, Charite Medical University, Berlin, Germany Colin Baigent Clinical Trial Service Unit and Epidemiological Studies Unit, Nuffield Department of Population Health, University of Oxford, Oxford, UK Matthew A. Bailey The British Heart Foundation Centre for Cardiovascular Science, The University of Edinburgh, Edinburgh, UK Richard Baker Renal Unit, St. James’s University Hospital, Leeds, UK

Ased Ali Freeman Hospital, Newcastle upon Tyne, UK

Joanne Bargman University of Toronto, Toronto; University Health Network, Toronto; and Toronto General Hospital, Toronto, ON, Canada

Richard D. M. Allen University of Sydney, Sydney; and Royal Prince Alfred Hospital, Sydney, Australia

Rashad S. Barsoum Cairo University, The Cairo Kidney Centre, Cairo, Egypt

Michael Allon Division of Nephrology, University of Alabama at Birmingham, Birmingham, AL, USA

Philip Beales Molecular Medicine Unit, UCL Institute of Child Health, London, UK

Halima Amer King’s College London BHF Centre, The Rayne Institute, St Thomas’ Hospital, London, UK

Monica Beaulieu Division of Nephrology, University of British Columbia, Vancouver, BC, Canada

Corinne Antignac Molecular Genetics Laboratories and Paediatrics, Inserm U983, Hôpital Necker-Enfants Malades, Paris, France

Aminu K. Bello Division of Nephrology & Immunology, University of Alberta, Edmonton, AB, Canada

Mugurel Apetrii Clinic of Nephrology, ‘C. I. Parhon’ University Hospital, ‘Gr. T. Popa’ University of Medicine and Pharmacy, Iasi, Romania

Rinaldo Bellomo Department of Intensive Care, Austin Hospital, Heidelberg, VIC, Australia

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contributors Katherine Bennett-Richards Department of Nephrology, Whipps Cross University Hospital, London, UK Jo H. M. Berden Department of Nephrology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands Carsten Bergmann Center for Human Genetics, Bioscientia, Ingelheim; University Hospital Freiburg, Freiburg; and Department of Human Genetics, RWTH Aachen University, Aachen, Germany Jürg Biber Institute of Physiology, University of Zurich, Zurich, Switzerland Daniel G. Bichet University of Montreal; and Hôpital du SacréCœur de Montréal, Montréal, QC, Canada Scott D. Bieber Division of Nephrology, Department of Medicine, University of Washington, Seattle; and Northwest Kidney Centers, Seattle, WA, USA Patrick Biggar Division of Nephrology, Klinikum Coburg GmbH, Coburg, Germany Heiko Billing Department of Pediatrics I, University Children’s Hospital, Heidelberg, Germany Coralie Bingham Exeter Kidney Unit, Royal Devon and Exeter Hospital (Wonford), Exeter, UK Lesley-Anne Bissell Division of Rheumatic and Musculoskeletal Disease, Leeds Institute of Molecular Medicine, University of Leeds, Leeds, UK

Cormac Breen Department of Nephrology and Transplantation, Guy’s and St Thomas NHS Foundation Trust, London, UK Jeremiah R. Brown The Dartmouth Institute for Health Policy and Clinical Practice and Departments of Medicine and Community and Family Medicine, Audrey and Theodor Geisel School of Medicine at Dartmouth, Hanover, NH; and Providence Park Heart Institute, Novi, MI, USA Nele Brusselaers Department of Molecular Medicine and Surgery, Karolinska Institutet, Stockholm, Sweden; and Department of Intensive Care Medicine and Burn Unit, Ghent University Hospital, Ghent University, Ghent, Belgium Emmanuel A. Burdmann Division of Nephrology, University of Sao Paulo Medical School, Sao Paulo, Brazil Aine Burns University College London (UCL) Centre for Nephrology Royal Free Hospital, Royal Free NHS Trust, London, UK David A. Bushinsky University of Rochester School of Medicine; and Nephrology Division, University of Rochester Medical Center, Rochester, NY, USA Patrice Cacoub UPMC Univ Paris 06, UMR 7211, Paris; INSERM, UMR_S 959, Paris; CNRS, UMR 7211, Paris; AP-HP, Groupe Hospitalier Pitié-Salpêtrière, Department of Internal Medicine, Paris, France Beverly D. Cameron Diabetes Education Center, St. John Providence Hospital, Southfield, MI

John J. Bissler Division of Nephrology and Hypertension, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH, USA

Stewart Cameron Department of Nephrology, Dialysis and Transplantation, King’s College London (Guy’s Hospital), London, UK

Christopher R. Blagg John Radcliffe Hospital, Oxford, UK; and Northwest Kidney Centers, University of Washington, Seattle, WA, USA

Angel Candela-Toha Department of Anaesthesiolgy and Reanimation, Hospital Universitario Ramón y Cajal, Madrid; Consorcio de investigación del Fracaso Renal Agudo de la Comunidad de Madrid (CIFRA), Madrid; and Instituto Ramón y Cajal de Investigación Sanitaria, Madrid, Spain

Judith Blaine Division of Renal Diseases and Hypertension, Department of Medicine, University of Colorado Anschutz Medical Campus, Aurora, CO, USA Detlef Bockenhauer Great Ormond Street Hospital for Children NHS Foundation Trust, London; and Institute of Child Health, University College London, London, UK Davide Bolignano Hypertension Unit, Department of Nephrology and Urology and CNR-Institute of Clinical Physiology, Ospedali Riuniti, Reggio Calabria, Italy Gregory L. Braden Tufts University School of Medicine, Baystate Medical Center, Springfield, MA, USA Kate Bramham Division of Women’s Health, King’s College London; and Women’s Health Academic Centre KHP, St Thomas’ Hospital, London, UK

Giovambattista Capasso Department of Internal Medicine and Department of Cardio-thoracic and Respiratory Science, Second University of Naples, Naples, Italy Andrés Cárdenas GI Unit, Institut Clinic de Malalties Digestives i Metaboliques, Hospital Clinic, and University of Barcelona, Institut d’Investigacions Biomèdiques August Pi-Sunyer (IDIBAPS), Ciber de Enfermedades Hepáticas y Digestivas (CIBEREHD), Barcelona; and InstitutoReina Sofıa de Investigación Nefrológica (IRSIN), Spain Juan Jesús Carrero Divisions of Baxter Novum and Renal Medicine, Department of Clinical Sciences, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden

 

contributors

Paul Carroll Consultant Endocrinologist, Guy’s & St. Thomas’ NHS Foundation Trust, London, UK

Lewis M. Cohen Division of Psychiatry, Baystate Medical Center, Tufts University School of Medicine, Springfield, MA, USA

Marie Cassart Department of Medical Imaging, University Clinics of Brussels—Erasme Hospital, Brussels, Belgium

Marie Condon Imperial College NHS Healthcare Trust Lupus Centre, Hammersmith Hospital, London, UK

Daniel C. Cattran University of Toronto, Toronto; Toronto General Research Institute, University Health Network; and Division of Nephrology, University Health Network, Toronto, ON, Canada

Thomas Connor Imperial College, London, UK

Steven Chadban Renal Unit, Royal Prince Alfred Hospital, Sydney, Australia

Jeff S. Coombes School of Human Movement Studies, University of Queensland, Brisbane, Australia

Christopher T. Chan Division of Nephrology, University Health Network, Toronto; and University of Toronto, Toronto, ON, Canada

Bruce A. Cooper Department of Renal Medicine, Kolling Institute of Medical Research, Royal North Shore Hospital, University of Sydney, St Leonards, Australia

Jung-San Chang Department of Renal Care, Kaohsiung Medical University, Kaohsiung, Taiwan

Cathy Corbishley Department of Cellular Pathology, St George’s Hospital, London, UK

Jeremy R. Chapman Renal Unit, Westmead Hospital, Westmead; and Centre for Transplant and Renal Research, Westmead Millennium Institute, University of Sydney, Westmead, Australia

Josef Coresh Cardiovascular Epidemiology, Comstock and Welch Centers, Johns Hopkins University, Bloomberg School of Public Health, Baltimore, MD, USA

Christos Chatziantoniou Hôpital Tenon, Paris; and Université Pierre et Marie Curie, Paris, France Dominique Chauveau Service de Néphrologie et Immunologie Clinique, CHU de Rangueil, Toulouse, France

Terry Cook Imperial College London, Hammersmith Campus, The Commonwealth Building, Hammersmith Hospital, UK

Adrian Covic Department of Nephrology, University of Medicine and Pharmacy ‘Gr. T. Popa’, Nephrology Clinic, ‘Dr. C. I. Parhon’ Hospital, Iaşi, Romania Sue Cox Guy’s & St Thomas’ NHS Foundation Trust, London, UK

Diana Chiu Salford Royal NHS Foundation Trust, Salford, UK

Gilles Crambert Metabolism and Renal Physiology Laboratory, Sorbonne Universités, UPMC Univ Paris 06, INSERM, Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, CNRS ERL8226, Centre de Recherche des Cordeliers, F-75006, Paris, France

Bhavna Chopra Transplant Nephrology, Mayo Clinic, Rochester, MN; and Nephrology, University of Pennsylvania, Pittsburgh, PA, USA

Antonia J. Cronin Guy’s Hospital Renal Unit, Guy’s and St Thomas’ Foundation Hospital, King’s Health Partners AHSC, London, UK

Constantina Chrysochou Salford Royal NHS Foundation Trust, Salford, UK

Dinna N. Cruz Division of Nephrology-Hypertension, Department of Medicine, University of California, San Diego, San Diego, CA, USA; and International Renal Research Institute (IRRIV), Vicenza, Italy

Hung-Chun Chen Department of Renal Care, Kaohsiung Medical University, Kaohsiung, Taiwan

William R. Clark Renal Therapeutic Area, Baxter Healthcare, Deerfield, IL, USA Philip Clayton Renal Unit, Royal Prince Alfred Hospital, Sydney, Australia Jan Clement National Reference Laboratory for Hantavirus Infections, Clinical Virology, University Hospital Gasthuisberg, University of Leuven, Leuven, Belgium Anna Clementi Department of Nephrology, Policlinico Universitario, Catania, Italy Eric P. Cohen Medical College of Wisconsin, Milwaukee, WI, USA

Zhao Cui Renal Division, Peking University First Hospital, Institute of Nephrology, Peking University, Key Laboratory of Renal Disease, Ministry of Health of China, Beijing, P. R. China Gary C. Curhan Channing Laboratory and Renal Division, Department of Medicine, Brigham and Women’s Hospital and Harvard Medical School, Boston, MA; and Division of Nephrology and Transplantation, Maine Medical Center, Portland, ME, USA Darshana Dadhania Division of Nephrology, Department of Medicine, Department of Transplantation Medicine, Weill Medical College of Cornell University, New York, USA

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contributors Michel Daudon Service des Explorations Fonctionnelles, Hôpital Tenon, APHP, Paris, France Andrew Davenport UCL Centre for Nephrology, Royal Free Campus, University College London Medical School, London, UK Marc E. De Broe Department of Nephrology, University of Antwerp, Antwerp, Belgium Benjamin Dekel The Pediatric Stem Cell Research Institute and Division of Pediatric Nephrology, Edmond and Lili Safra Children’s Hospital, Chaim Sheba Medical Center, Tel Hashomer, Sackler Faculty of Medicine, Tel Aviv University, Israel Joris Delanghe Department of Clinical Chemistry, Ghent University Hospital, Gent, Belgium Olivier Devuyst Institute of Physiology, University of Zurich, Zurich; and Division of Nephrology, University Hospital Zurich, Zurich, SwitzerlandUniversité catholique de Louvain Medical School, Brussels, Belgium

Duska Dragun Department of Nephrology and Medical Intensive Care, Charité University Hospital, Campus Virchow-Klinikum, Berlin, Germany Joost P. H. Drenth Department of Gastroenterology and Hepatology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands Jeremy S. Duffield Division of Nephrology and Center for Lung Biology, Departments of Medicine & Pathology, and Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA, USA Sara Dunsmore Toronto General Hospital, Toronto, ON, Canada Jean-Claude Dussaule Service de Physiologie HUPE, Assistance Publique-Hôpitaux de Paris (AP-HP), Paris; Université Pierre et Marie Curie, Paris; and INSERM U702, Hôpital Tenon, Paris, France John Eastwood Department of Renal Medicine and Transplantation, St George’s Hospital, London, UK

Patrick C. D’Haese Laboratory of Pathophysiology, University of Antwerp, Antwerp, Belgium

Tim Eisen Department of Oncology, Cambridge University Hospitals NHS Foundation Trust, Cambridge, UK

Ramya Dhandapani Royal Liverpool University Hospital, Liverpool, UK

Grahame J. Elder Department of Renal Medicine, Westmead Hospital, Westmead; and Osteoporosis and Bone Biology Division, Garvan Institute of Medical Research, Sydney, Australia

Neeraj Dhaun British Heart Foundation Centre for Cardiovascular Science, Queen’s Medical Research Institute, University of Edinburgh; and Department of Renal Medicine, Royal Infirmary of Edinburgh, Edinburgh, UK Meletios A. Dimopoulos Department of Clinical Therapeutics, National and Kapodistrian University of Athens School of Medicine, Athens, Greece Salvatore Di Filippo Department of Nephrology, Dialysis and Renal Transplant, Alessandro Manzoni Hospital, Lecco, Italy Bradley P. Dixon Division of Nephrology and Hypertension, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH, USA

Marlies Elger Centre for Biomedicine and Medical Technology Mannheim, Department of Anatomy and Developmental Biology, Medical Faculty Mannheim, University of Heidelberg, Heidelberg, Germany Timothy Ellam Department of Cardiovascular Science, University of Sheffield Medical School, Sheffield, UK David H. Ellison Division of Nephrology & Hypertension, Oregon Health & Science University & Renal Section VAMC, Portland, OR, USA

Luis D’Marco Hospital Universitario Ruíz y Páez, Universidad de Oriente, Venezuela

Paul Emery Leeds Musculoskeletal Biomedical Research Unit, Leeds Teaching Hospitals NHS Trust, Leeds; and Rheumatic and Musculoskeletal Disease, Leeds Institute of Molecular Medicine, University of Leeds, Leeds, UK

Philippa Dodd Imperial College NHS Healthcare Trust Lupus Centre, Hammersmith Hospital, London, UK

Karlhans Endlich Department of Anatomy and Cell Biology, University Medicine Greifswald, Greifswald, Germany

Anthony Dorling MRC Centre for Transplantation, King’s College London, London; and Guy’s and St Thomas’ NHS Foundation Trust, Guy’s Hospital, London, UK

Fabrizio Fabrizi Division of Nephrology, Maggiore Policlinico Hospital, IRCCS Foundation, Milano, Italy

Alain Doucet Metabolism and Renal Physiology Laboratory, Sorbonne Universités, UPMC Univ Paris 06, INSERM, Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, CNRS ERL8226, Centre de Recherche des Cordeliers, F-75006, Paris, France

Robert G. Fassett Royal Brisbane and Women’s Hospital, Brisbane; and School of Human Movement Studies, University of Queensland, Brisbane, Australia Bengt Fellström University of Uppsala, Uppsala, Sweden

 

contributors

Javier Fernández Liver Unit, Hospital Clinic Barcelona, IDIBAPS, Ciberhed, Barcelona, Spain

Michael J. Germain Baystate Medical Center, Tufts University School of Medicine, Springfield, MA, USA

Charles J. Ferro Department of Renal Medicine, Queen Elizabeth Hospital and University of Birmingham, Birmingham, UK

Morie A. Gertz Division of Hematology, Mayo Clinic, Rochester, MN, USA

Fernando C. Fervenza Division of Nephrology and Hypertension, Mayo Clinic, Rochester, MN, USA

Tom J. G. Gevers Department of Gastroenterology and Hepatology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands

Alexander Fichtner Division of Paediatric Nephrology, Centre for Paediatrics and Adolescent Medicine, University of Heidelberg, Heidelberg, Germany Fredric O. Finkelstein Yale University, New Haven, CT, USA Susan H. Finkelstein Yale University, New Haven, CT, USA Martin Flamant Hopital Bichat, Paris; and Université Diderot, Paris, France Stewart Fleming University of Dundee, Ninewells Hospital, Dundee, UK Frances Flinter Genetics Department, Guy’s & St Thomas’ NHS Foundation Trust, London, UK Danilo Fliser Department of Internal Medicine IV, Saarland University Medical Hospital, Homburg/Saar, Germany Lukas Foggensteiner The New Queen Elizabeth Hospital Birmingham, Edgbaston, Birmingham, UK Agnes B. Fogo Pathology Department, Vanderbilt University Medical Center, Nashville , TN, USA Ted A. Foster Oregon Health and Science, University Division of Internal Medicine, Portland, OR, USA Brunella Franco Telethon Institute of Genetics and Medicine (TIGEM), Department of Pediatrics Medical Genetics Services, Federico II University of Naples, Naples, Italy Valeska Frank Center for Human Genetics, Bioscientia, Ingelheim, Germany Benjamin J. Freda Tufts University School of Medicine, Baystate Medical Center, Springfield, MA; and Continuous Renal Replacement Therapies, Western New England Renal and Transplantation Associates, Springfield, MA, USA Simon J. Freeman Plymouth Hospitals NHS Trust, Radiology Department, Derriford Hospital, Crownhill, Plymouth, UK Daniel P. Gale UCL Centre for Nephrology, Royal Free Hospital School of Medicine, London, UK Timur A. Galperin St. Louis University, Department of Dermatology, Anheuser Busch Institute, St. Louis, MO, USA Giorgio Gentile IRCCS—Istituto di Ricerche Farmacologiche Mario Negri, Bergamo; and Unit of Nephrology, Azienda Ospedaliera Papa Giovanni XXIII, Bergamo, Italy

Pere Ginès Liver Unit, Institut Clinic de Malalties Digestives i Metaboliques, Hospital Clinic, and University of Barcelona, Institut d’Investigacions Biomèdiques August Pi-Sunyer (IDIBAPS), Ciber de Enfermedades Hepáticas y Digestivas (CIBEREHD), Barcelona; and InstitutoReina Sofıa de Investigación Nefrológica (IRSIN), Spain Hector Giral Division of Renal Diseases and Hypertension, University of Colorado at Denver and Health Sciences Center, Aurora, CO, USA Anna Giuliani Department of Nephrology Dialysis & Transplantation, San Bortolo Hospital, and International Renal Research Institute (IRRIV), Vicenza, Italy Richard J. Glassock Geffen School of Medicine at UCLA, Los Angeles, CA, USA James F. Glockner Department of Radiology, Mayo Medical School, Mayo Clinic, Rochester MN, USA Griet Glorieux Nephrology Division, University Hospital Ghent, Ghent, Belgium Luigi Gnudi Unit for Metabolic Medicine, Cardiovascular Division, King’s College London, London, UK Rishi M. Goel Department of Gastroenterology, Guy’s & St Thomas’ NHS Foundation Trust, London, UK Eric Goffin Department of Nephrology, Cliniques Universitaires Saint-Luc, Université Catholique de Louvain, Brussels, Belgium M. Refik Gökmen Department of Nephrology & Transplantation, Guy’s & St Thomas’ NHS Foundation Trust, London, UK Stanley Goldfarb Renal Electrolyte and Hypertension Division, University of Pennsylvania, Pittsburgh, PA, USA David J. Goldsmith Consultant Nephrologist, Guy’s and St Thomas’ Hospitals, London, UK; Professor, Cardiovascular and Cell Sciences Institute, St George’s University of London, London, UK Michael S. Goligorsky Renal Research Institute, New York Medical College, Valhalla, NY, USA Thomas A. Golper Vanderbilt University Medical Center, Medical Center North, Nashville, TN, USA Olga Gonzalez-Albarrán Endocrinology Dapartment, Hospital Universitario Ramón y Cajal, Madrid, Spain

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contributors Tim Goodship Institute of Human Genetics, Newcastle University, International Centre for Life, Newcastle upon Tyne, UK Paul Goodyer Montreal Children’s Hospital Research Institute, Montreal, QC, Canada Morgan E. Grams The Johns Hopkins Hospital, Baltimore, MD, USA John Grange London Clinic Cancer Centre B2, London, UK Darren Green University of Manchester, Manchester, UK Simon Gruenewald Departments of Nuclear Medicine and Radiology, Westmead Hospital, Westmead, Australia Marie Claire Gubler INSERM U983, Hôpital Necker-Enfants Malades, Paris; and APHP, Centre de référence des Maladies Rénales Héréditaires (MARHEA), Paris, France Victor Gueutin Department of Nephrology, Pitié-Salpêtrière Hospital, Paris, France Mónica Guevara Liver Unit, Hospital Clinic Barcelona, IDIBAPS, Ciberhed, Barcelona, Spain Theresa A. Guise Endocrinology and Metabolism, Department of Internal Medicine, Indiana University School of Medicine, Indianapolis, IN, USA Orlando M. Gutiérrez Division of Nephrology, Department of Medicine, School of Medicine; and Department of Epidemiology, School of Public Health, University of Alabama at Birmingham, Birmingham, AL, USA

Choli Hartono Division of Nephrology, Department of Medicine, Department of Transplantation Medicine, Weill Medical College of Cornell University, New York, USA Dietrich Hasper Department of Nephrology and Medical Intensive Care, Charité University Hospital, Campus VirchowKlinikum, Berlin, Germany Nick Hastie MRC Human Genetics Unit, University of Edinburgh Western General Hospital, Edinburgh, UK Jean-Philippe Haymann Department of Physiology, Hopital Tenon (AP-HP/UPMC/UMRS 702), Paris, France Richard Haynes Clinical Trial Service Unit and Epidemiological Studies Unit, Nuffield Department of Population Health, University of Oxford; and Oxford Kidney Unit, Oxford University Hospitals, Oxford, UK Peter Heeringa Department of Pathology and Medical Biology, University Medical Center Groningen, Groningen, The Netherlands Björn Hegner Department of Nephrology and Medical Intensive Care, Charité University Hospital, Campus Virchow-Klinikum, Berlin, Germany Laurence Heidet AP-HP, Service de Néphrologie Pédiatrique, Hôpital Necker-Enfants Malades, Paris; and APHP, Centre de référence des Maladies Rénales Héréditaires (MARHEA), Paris, France Udo Helmchen Department of Pathology, University of Hamburg, Hamburg, Germany

Mark Haas Department of Pathology and Laboratory Medicine, Cedars-Sinai Medical Center, Los Angeles, CA, USA

Heather Henderson Division of Nephrology, St. John Hospital and Medical Center, Detroit, MI, USA

Reza Hajhosseiny BHF Centre of Cardiovascular Excellence, Guy’s & St. Thomas’ NHS Foundation Trust, King’s College Academic Health Partners, London, UK

Lorna K. Henderson Renal Unit, Westmead Hospital, Westmead, Australia

Andrew Hall Institute of Anatomy, University of Zurich, Zurich, Switzerland Michèle Hall Department of Pediatric Nephrology, University Clinics of Brussels—Erasme Hospital, Brussels, Belgium

William G. Herrington Clinical Trial Service Unit, Richard Doll Building, Oxford; Clinical Trial Service Unit and Epidemiological Studies Unit, Nuffield Department of Population Health, Richard Doll Building, Oxford; and Oxford Kidney Unit, Oxford University Hospitals NHS Trust, Churchill Hospital, Oxford, UK

Mitchell L. Halperin Li Ka Shing Knowledge Institute in St. Michael’s Hospital, Toronto, ON, Canada

Irene J. Higginson King’s College London, London, UK

Mark Harber UCL Centre for Nephrology, Royal Free Hospital, London, UK

Jonathan Himmelfarb Division of Nephrology, Department of Medicine, Kidney Research Institute, University of Washington, Seattle, WA, USA

Lorraine Harper School of Immunity and Infection, Centre for Translational Inflammation Research, Queen Elizabeth Hospital Birmingham, Edgbaston, Birmingham, AL, USA

Hallvard Holdaas Rikshospitalet, Uniiversity of Oslo, Oslo, Norway

David C. H. Harris Sydney Medical School-Westmead, University of Sydney, Westmead Hospital, Westmead, Australia

Ewout Hoorn Division of Nephrology & Transplantation, Department of Internal Medicine, Erasmus Medical Center, Rotterdam, The Netherlands

 

Eric A. J. Hoste Department of Intensive Care Medicine, Ghent University Hospital, Ghent, Belgium Pascal Houillier Paris Descartes University; and Hopital Europeen Georges Pompidou, Assistance Publique-Hopitaux de Paris, Paris, France

contributors

Sabina Jelen Division of Renal Diseases and Hypertension, Department of Medicine, University of Colorado Anschutz Medical Campus, Aurora, CO, USA Vivekanand Jha Department of Nephrology, Postgraduate Institute of Medical Education and Research, Chandigarh, India

Frédéric A. Houssiau Service de Rhumatologie, Département de Médecine Interne, Cliniques universitaires Saint-Luc, Université catholique de Louvain, Bruxelles, Belgium

Achim Jörres Department of Nephrology and Medical Intensive Care, Charité University Hospital, Campus Virchow-Klinikum, Berlin, Germany

Erin J. Howden School of Human Movement Studies, University of Queensland, Brisbane, Australia

Dennis Joseph Division of Endocrinology, Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, USA

Peter Howells Department of Medical Physics, Old Medical School, Leeds General Infirmary, Leeds, UK Jeremy Hughes University of Edinburgh, Renal Medicine, Edinburgh Royal Infirmary, Edinburgh, UK Beverley J. Hunt MRC Centre for Transplantation, King’s College London, London; and Guy’s and St Thomas’ NHS Foundation Trust, Guy’s Hospital, London, UK Alastair J. Hutchison Department of Renal Medicine, Manchester Academic Health Science Centre, Manchester, UK Young-Hwan Hwang Division of Nephrology, University Health Network and University of Toronto, Toronto, ON, Canada; and Department of Internal Medicine, Eulji General Hospital, Seoul, South Korea Lesley A. Inker Division of Nephrology, Tufts Medical Center and Tufts University School of Medicine, Boston, MA, USA Nadia Wasi Iqbal Division of Nephrology, Saint Louis University, St. Louis, MO, USA Hassan Izzedine Department of Nephrology, Pitié-Salpêtrière Hospital, Paris, France Bertrand L. Jaber Department of Medicine, Tufts University School of Medicine, St. Elizabeth’s Medical Center, Boston, MA, USA Michel Jadoul Department of Nephrology, Cliniques Universitaires Saint-Luc, Université Catholique de Louvain, Brussels, Belgium Kitty J. Jager Department of Medical Informatics, Academic Medical Center, Amsterdam, The Netherlands

Islam Junaid Department of Urology, The Royal London Hospital, London, UK Paul Jungers Département de Néphrologie, Hôpital Necker, APHP, Paris, France Philip A. Kalra Genomic Epidemiology Research Group, The Univeristy of Manchester, Manchester, UK Kamel S. Kamel Keenan Research Center in the Li Ka Shing Knowledge Institute in St Michael’s Hospital and Division of Nephrology, St. Michael’s Hospital, University of Toronto, Toronto, ON, Canada Mehmet Kanbay Division of Nephrology, Department of Medicine, Koc University School of Medicine, Istanbul, Turkey Duk-Hee Kang Division of Nephrology, Department of Internal Medicine, Ewha Womans University School of Medicine, Ewha Medical Research Center, Seoul, Korea Ea Wha Kang Division of Nephrology, Department of Internal Medicine, NHIC Ilsan Hospital, Goyang, Korea Efstathios Kastritis Department of Clinical Therapeutics, National and Kapodistrian University of Athens School of Medicine, Athens, Greece Akira Kawashima Department of Radiology, Mayo Medical School, Mayo Clinic, Rochester, MN, USA John A. Kellum Department of Critical Care Medicine, Center for Critical Care Nephrology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

Mirian C. Janssen Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands

Claudia Kemper Medical Research Council Centre for Transplantation, King’s College London and NIHR Biomedical Research Centre at Guy’s & St Thomas’ NHS Foundation Trust and King’s College London, UK

Alan G. Jardine BHF Cardiovascular Research Centre, University of Glasgow, Glasgow, UK

Steven Kennish Department of Radiology, Royal Hallamshire Hospital, Sheffield

David Jayne Vasculitis and Lupus Clinic, Addenbrooke’s Hospital, Cambridge, UK

Markus Ketteler Division of Nephrology, Klinikum Coburg GmbH, Coburg, Germany

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contributors Kaivan Khavandi BHF Centre of Cardiovascular Excellence, Guy’s & St. Thomas’ NHS Foundation Trust, King’s College Academic Health Partners, London, UK

Wolfgang Kühn Renal Division, University Freiburg Medical Center, Freiburg; and BIOSS Center for Biological Signalling Studies, University of Freiburg, Freiburg, Germany

J. Christopher Kingswood Sussex Renal Unit, The Royal Sussex County Hospital, Brighton, UK

Navneet Kumar Division of Cardiology, St. John Hospital and Medical Center, Detroit, MI, USA

Kazuhiro Kitajima Department of Radiology, Mayo Medical School, Mayo Clinic, Rochester MN, USA

Manjula Kurella Tamura Division of Nephrology, Stanford University School of Medicine, Stanford, CA; and VA Palo Alto Health Care System Geriatrics Research Education & Clinical Center, Palo Alto, CA, USA

Robert Kleta University College London, Royal Free Hospital, London, UK Alan S. Kliger Hospital of Saint Raphael, New Haven; and Yale University School of Medicine, New Haven, CT, USA Bertrand Knebelmann Service de Néphrologie, Hôpital NeckerEnfants Malades, Paris, France Simon R. Knight Nuffield Department of Surgical Sciences, University of Oxford, Oxford; and Centre for Evidence in Transplantation, Clinical Effectiveness Unit, The Royal College of Surgeons of England, London, UK Harbir Singh Kohli Post Graduate Institute of Medical Education and Research, Chandigarh, India

Dirk R. J. Kuypers Department of Nephrology and Renal Transplantation, University Hospitals Leuven, Leuven, Belgium Laura Labriola Department of Nephrology, Cliniques Universitaires Saint-Luc, Université Catholique de Louvain, Brussels, Belgium Helen J. Lachmann UK National Amyloidosis Centre, University College London School of Medicine, London, UK Robin Lachmann Charles Dent Metabolic Unit, National Hospital for Neurology and Neurosurgery, London, UK Kar Neng Lai University of Hong Kong, Hong Kong

Martin Konrad University Children’s Hospital, Pediatric Nephrology, Münster; and University Hospital Münster, Department of General Pediatrics, Pediatric Nephrology, Münster, Germany

Edmund J. Lamb Clinical Biochemistry, East Kent Hospitals University NHS Foundation Trust, Canterbury, Kent, UK

Stephen M. Korbet Department of Medicine, Rush University Medical Center, Chicago, IL, USA

Anne-Sophie Lambert Department of Medical Imaging, University Clinics of Brussels—Erasme Hospital, Brussels, Belgium

Maarit Korkeila Divisions of Renal Medicine and Baxter Novum, Department of Clinical Science, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden Camille Nelson Kotton Infectious Diseases Division, Massachusetts General Hospital/Harvard Medical School, Boston, MA, USA Jeannette Kathrin Kraft Clarendon Wing Radiology Department, Leeds General Infirmary, Leeds, UK Wilhelm Kriz Centre for Biomedicine and Medical Technology Mannheim (CBTM), Anatomy and Developmental Biology, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany Wilhelm Kriz Department of Anatomy and Developmental Biology, Medical Faculty Mannheim, Centre for Biomedicine and Medical Technology Mannheim, University of Heidelberg, Heidelberg, Germany Jean-Marie Krzesinski Department of Medicine, Nephrology Unit, B35, University of Liège, Belgium

Heather Lambert Newcastle University, Newcastle upon Tyne, UK

Norbert Lameire University Hospital Ghent, Ghent, Belgium Martin J. Landray Clinical Trial Service Unit and Epidemiological Studies Unit, Nuffield Department of Population Health, University of Oxford, Oxford, UK Christophe Legendre Université Paris Descartes and Hôpital Necker and INSERM U845, Paris; and Service de Transplantation Rénale Adulte, Hôpital Necker, Paris, France Rachel Lennon Wellcome Trust Centre for Cell-Matrix Research Michael Smith Building, University of Manchester, Manchester, UK Edgar V. Lerma Section of Nephrology, Department of Medicine, University of Illinois at Chicago College of Medicine, Chicago, IL, USA Andrew J. LeRoy Department of Radiology, Mayo Medical School, Mayo Clinic, Rochester, MN, USA Kieron S. Leslie Department of Dermatology, University of California San Francisco, San Francisco, CA, USA

 

Andrew S. Levey Tufts Medical Center and Tufts University School of Medicine, Boston; and US Department of Agriculture Jean Mayer Human Nutrition Research Center at Tufts University, Boston; and Clinical Research Graduate Program (PhD and MS), Sackler School of Graduate Biomedical Sciences, Boston, MA, USA Moshe Levi Division of Renal Diseases and Hypertension, Department of Medicine, University of Colorado Anschutz Medical Campus, Aurora, CO, USA

contributors

Rodger Loutzenhiser Department of Physiology and Pharmacology, University of Calgary, Calgary, AB, Canada Valerie A. Luyckx Division of Nephrology, University of Alberta, Edmonton, AB, Canada Bernadette Lynch Centre for Rheumatology, Royal Free Hospital, London, UK Naim Maalouf Department of Internal Medicine, University of Texas, Southwestern Medical Center, TX, USA

Adeera Levin Division of Nephrology, University of British Columbia, Vancouver; and BC Provincial Renal Agency, Vancouver, BC, Canada

Iain C. Macdougall Department of Renal Medicine, King’s College Hospital, London, UK

Elena N. Levtchenko Department of Pediatric Nephrology, University Hospitals Leuven, Leuven, Belgium

Iain M. MacIntyre Department of Clinical Pharmacology, Royal Infirmary of Edinburgh, Edinburgh, UK

Edmund J. Lewis Department of Medicine, Rush University Medical Center, Chicago, IL, USA

Friederike Mackensen Interdisciplinary Uveitis Center, Department of Ophthalmology, University of Heidelberg, Heidelberg, Germany

Fernando Liaño Consorcio de investigación del Fracaso Renal Agudo de la Comunidad de Madrid (CIFRA), Madrid; and Instituto Ramón y Cajal de Investigación Sanitaria, Madrid; and Department of Nephrology, Hospital Universitario Ramón y Cajal, Madrid; and Department of Medicine, Universidad de Alcalá, Alcalá de Henares, Spain

Finlay MacKenzie United Kingdom National External Quality Assessment Scheme (UKNEQAS), University Hospitals Birmingham NHS Foundation Trust, East Wing, Institute of Research and Development, Birmingham, UK

Aurora Lietor Department of Critical Care Medicine, Hospital Universitario Ramón y Cajal, Madrid, Spain Liz Lightstone Imperial College NHS Healthcare Trust Lupus Centre, Hammersmith Hospital, London; and Section of Renal Medicine and Vascular Inflammation, Department of Medicine, Imperial College London, London, UK Bengt Lindholm Divisions of Baxter Novum and Renal Medicine, Department of Clinical Sciences, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden Agnès Linglart Department of Pediatric Endocrinology, Reference Center for Rare Disorders of the Mineral Metabolism, Paris, France Francois Lionnet Sickle cell Center, Hopital Tenon (AP-HP), Paris, France Jeffrey Lipman Department of Intensive Care Medicine, Royal Brisbane and Womens Hospital, Herston, Australia Francesco Locatelli Department of Nephrology, Dialysis and Renal Transplant, Alessandro Manzoni Hospital, Lecco, Italy Gerard M. London Department of Pharmacology, Georges Pompidou European Hospital, National Institute of Health and Medical Research U970, Paris; and Department of Nephrology, Manhès Hospital, Fleury Mérogis, France Graham M. Lord Department of Experimental Immunobiology, King’s College London, London; and Guy’s & St Thomas’ NHS Foundation Trust, London, UK

Piet Maes National Reference Laboratory for Hantavirus Infections, Clinical Virology, University Hospital Gasthuisberg, University of Leuven, Leuven, Belgium Julien Maizel Department of Nephrology, Medical Intensive Care Center, Amiens University Medical Center, Amiens, France Francesca Mallamaci Nephrology and Renal Transplantation Unit, Department of Nephrology and Urology and CNR-IBIM, Ospedali Riuniti, Reggio Calabria, Italy Jolanta Malyszko 2nd Department of Nephrology, Medical University, Bialystok, Poland Veena Manjunath Section of Nephrology, Yale University School of Medicine, New Haven, CT, USA Celestina Manzoni Department of Nephrology, Dialysis and Renal Transplant, Alessandro Manzoni Hospital, Lecco, Italy Stephen D. Marks Department of Paediatric Nephrology, Great Ormond Street Hospital for Children NHS Foundation Trust, Great Ormond Street, London, UK David Marples Institute of Membrane and Systems Biology, Faculty of Biological Sciences, University of Leeds, Leeds, UK Mark R. Marshall Department of Renal Medicine, Counties Manukau District Health Board; and Faculty of Medicine and Health Science, University of Auckland, Auckland, New Zealand Kevin J. Martin Division of Nephrology, Saint Louis University, St. Louis, MO, USA

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contributors Philip Mason Oxford Kidney Unit, The Churchill Hospital, Oxford, UK

Elaine Murphy Charles Dent Metabolic Unit, National Hospital for Neurology and Neurosurgery, London, UK

Anne Massez Department of Medical Imaging, University Clinics of Brussels—Erasme Hospital, Brussels, Belgium

Fliss E. M. Murtagh King’s College London, London, UK

Patrick H. Maxwell Cambridge Biomedical Campus, University of Cambridge, Cambridge, UK Peter A. McCullough Baylor University Medical Center, Baylor Heart and Vascular Institute, Baylor Jack and Jane Hamilton Heart and Vascular Hospital, Dallas, TX; and The Heart Hospital, Plano, TX, USA Emily P. McQuarrie Renal Unit, Western Infirmary, Glasgow, UK Nicholas Medjeral-Thomas Imperial College, London, UK Rajnish Mehrotra Division of Nephrology, Department of Medicine, Kidney Research Institute, University of Washington, Seattle, WA, USA Ravindra L. Mehta Division of Nephrology and Hypertension, Department of Medicine, University of California San Diego, San Diego, CA, USA Giampaolo Merlini Amyloid Research and Treatment Center, Foundation Scientific Institute Policlinico San Matteo, University of Pavia, Pavia, Italy Alain Meyrier Department of Nephrology, University ParisDescartes, Hôpital Georges Pompidou and Broussais, Paris, France

Thangamani Muthukumar Division of Nephrology, Department of Medicine, Department of Transplantation Medicine, Weill Medical College of Cornell University, New York, USA Walter P. Mutter Department of Medicine, Division of Nephrology, Newton-Wellesley Hospital, Newton, MA Maarten Naesens Department of Nephrology and Renal Transplantation, University Hospitals Leuven, Leuven, Belgium Saraladevi Naicker Division of Nephrology, Department of Internal Medicine, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa Brian J. Nankivell Renal Unit, Westmead Hospital, Westmead, Australia John Neary Royal Infirmary, Little France, Edinburgh, UK Nathalie Neirynck Nephrology Division, University Hospital Ghent, Ghent, Belgium Catherine Nelson-Piercy St Thomas’ Hospital, London, UK Khai Ping Ng Department of Renal Medicine, Queen Elizabeth Hospital and University of Birmingham, Birmingham, UK Patrick Niaudet Hopital Necker-Enfants Malades, Paris, France

Eve Miller-Hodges MRC Human Genetics Unit, University of Edinburgh Western General Hospital, Edinburgh, UK

Peter W. Nickerson Canadian Blood Services, Winnipeg, MB, Canada

Christopher Mitchell Children’s Services, John Radcliffe Hospital, Oxford, UK (now retired)

Søren Nielsen Department of Biomedicine, The Water and Salt Research Centre, Aarhus University, Aarhus, Denmark

Orson Moe University of Texas Southwestern Medical Center, Dallas; and Charles and Jane Pak Center for Mineral Metabolism and Clinical Research, Dallas, TX, USA

Allen R. Nissenson DaVita Inc., Denver, CO; and David Geffen School of Medicine at UCLA

Gilbert W. Moeckel Department of Pathology, Yale University School of Medicine, New Haven, CT, USA Thomas Mone OneLegacy, Los Angeles, CA, USA John Moran DaVita Inc., Denver, CO, USA Peter J. Morris Centre for Evidence in Transplantation, Royal College of Surgeons of England, London, UK Marion Muche Division of Gastroenterology, Infectious Diseases and Rheumatology, Department of Medicine, Charite Medical University, Hindenburgdamm, Berlin, Germany Heini Murer Institute of Physiology, University of Zurich, Zurich, Switzerland

Marina Noris IRCCS-Istituto di Ricerche Farmacologiche Mario Negri, Bergamo, Italy Ali J. Olyaei Division of Nephrology and Hypertension, Oregon Health and Science University, Portland, OR, USA Albert C. M. Ong Department of Infection and Immunity, The University of Sheffield Medical School, Sheffield, UK Michael Oppert Emergency Department, Klinikum Ernst von Bergmann, Potsdam, Germany Stephan R. Orth Dialysis Centre, Bad Aibling, Germany Nadina Ortiz-Brüchle Department of Human Genetics, RWTH Aachen University, Aachen, Germany

 

Graham Paget Division of Nephrology, Department of Internal Medicine, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa Chirag R. Parikh Program for Applied Translational Research, Section of Nephrology, Department of Medicine, Yale School of Medicine, New Haven, CT, USA Kamal V. Patel Guy’s and St Thomas’ Hospitals London, UK Rajan K. Patel Institute of Cardiovascular and Medical Sciences, University of Glasgow, Glasgow, UK York Pei Division of Nephrology, University Health Network and University of Toronto, Toronto, ON, Canada

contributors

Robert Provenzano Division of Nephrology, Hypertensive and Transplant Medicine, St John Hospital and Medical Center, Detroit, MI; and Wayne State University, School of Medicine, Detroit, MI, USA John Prowle Adult Critical Care Unit, Royal London Hospital, Barts and the London NHS Trust, London, UK James M. Pullman Department of Pathology, Albert Einstein College of Medicine; and Montefiore Medical Center, Bronx, NY, USA May M. Rabadi Department of Anesthesiology, College of Physicians and Surgeons, Columbia University, New York, USA

Mark A. Perazella Section of Nephrology, Department of Medicine, Yale University, New Haven, CT, USA

Milan RadoviĆ School of Medicine, University of Belgrade, Belgrade; and Department of Nephrology, Cllinical Center of Serbia, Belgrade, Serbia

Lisa M. Phipps Department of Renal Medicine, Orange Base Hospital, Orange, Australia

Paolo Raggi Mazankowski Alberta Heart Institute, University of Alberta, Edmonton, AB, Canada

Rob Pickard Newcastle University, Newcastle upon Tyne; and Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK

Shamima Rahman Mitochondrial Research Group, Genetics and Genomic Medicine Programme, UCL Institute of Child Health, London, UK

Matthew C. Pickering Imperial College, London, UK

Ravindra Rajakariar Barts Health NHS Trust, London, UK

Michael L. Picton Department of Renal Medicine, Manchester Royal Infirmary and Manchester Academic Health Science Centre, Manchester, UK

Premil Rajakrishna Renal Medicine, Royal Infirmary, Edinburgh, UK

Arkadiy Pinkhasov Baystate Medical Center, Tufts University School of Medicine, Springfield, MA, USA Richard K. S. Phoon Westmead Clinical School, Westmead, New South Wales, Australia Yves Pirson Department of Nephrology, Cliniques Universitaires St Luc, Brussels, Belgium Henry C. C. Pleass University of Sydney, Sydney; and Westmead Hospital, Westmead, Australia Oren Pleniceanu The Pediatric Stem Cell Research Institute and Division of Pediatric Nephrology, Edmond and Lili Safra Children’s Hospital, Chaim Sheba Medical Center, Tel Hashomer, Sackler Faculty of Medicine, Tel Aviv University, Israel Anneleen Pletinck Nephrology Division, University Hospital Ghent, Ghent, Belgium Rutger J. Ploeg Nuffield Department of Surgical Sciences, University of Oxford, Oxford, UK Carol A. Pollock Department of Renal Medicine, Kolling Institute of Medical Research, Royal North Shore Hospital, University of Sydney, St Leonards, Australia Giuseppe Pontoriero Department of Nephrology, Dialysis and Renal Transplant, Alessandro Manzoni Hospital, Lecco, Italy

S. Vincent Rajkumar Division of Hematology, Mayo Clinic, Rochester, MN, USA Raja Ramachandran Department of Nephrology, Postgraduate Institute of Medical Education and Research, Chandigarh, India Wolfgang Rascher University Hospital Erlangen, Erlangen, Germany Brian B. Ratliff Department of Medicine, New York Medical College, Valhalla, NY, USA Gauthier Raynal Université Pierre et Marie Curie, Hôpital Tenon, Service d’urologie, Paris, France Lesley Rees Department of Paediatric Nephrology, Great Ormond Street Hospital for Children, London, UK Heather N. Reich University Health Network and University of Toronto, Toronto, ON, Canada Giuseppe Remuzzi Mario Negri Institute for Pharmacological Research, Bergamo, Italy Anna Richards Queen’s Medical Research Institute, University of Edinburgh, Edinburgh, UK James Ritchie Salford Royal NHS Foundation Trust, Salford, UK Ian S. D. Roberts Department of Cellular Pathology, John Radcliffe Hospital, Oxford, UK

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contributors Bernardo Rodriguez-Iturbe Universidad del Zulia, Hospital Universitario de Maracaibo, Maracaibo; and Laboratorio de Inmunobiología, Instituto Venezolano Investigaciones Científicas (IVIC)-Zulia, Venezuala Claudio Ronco Department of Nephrology Dialysis & Transplantation and International Renal Research Institute (IRRIV), San Bortolo Hospital, Vicenza, Italy

Franz Schaefer Centre for Pediatrics and Adolescent Medicine, Heidelberg, Germany Eva Schepers Nephrology Division, University Hospital Ghent, Ghent, Belgium Miet Schetz Department of Intensive Care Medicine, University Hospital KU Leuven, Leuven, Belgium

Pierre M. Ronco INSERM UMR_S 702, Tenon Hospital, Paris, France

Laurent Schild Department of Pharmacology & Toxicology, Lausanne University, Lausanne, Switzerland

Mitchell H. Rosner Division of Nephrology, University of Virginia Health System, Charlottesville, VA, USA

Adalbert Schiller Department of Nephrology, University of Medicine and Pharmacy ‘Victor Babeş’, Nephrology Clinic, Emergency County Hospital, Timişoara, Romania

Hansjörg Rothe Division of Nephrology, Klinikum Coburg GmbH, Coburg, Germany Piero Ruggenenti IRCCS—Istituto di Ricerche Farmacologiche Mario Negri, Bergamo; and Unit of Nephrology, Azienda Ospedaliera Papa Giovanni XXIII, Bergamo, Italy

Karl P. Schlingmann University Children’s Hospital, Pediatric Nephrology, Münster, Germany Michiel F. Schreuder Radboud University Medical Center, Nijmegen, The Netherlands

Luis M. Ruilope Nephrology Department, Hospital Doce de Octubre, Madrid, Spain

Melvin M. Schwartz Department of Pathology, Rush University Medical Center, Chicago, IL, USA

Andrew D. Rule Mayo Clinic and Medical School, Rochester, MN, USA

Vedat Schwenger Department of Nephrology, University of Heidelberg, Heidelberg, Germany

David N. Rush Health Sciences Centre, Winnipeg, MB, Canada

Victor F. Seabra Department of Medicine, Tufts University School of Medicine, St. Elizabeth’s Medical Center, Boston, MA, USA

Khashayar Sakhaee Division of Mineral Metabolism, University of Texas, Dallas, TX, USA Vinay Sakhuja Post Graduate Institute of Medical Education and Research, Chandigarh, India Alan D. Salama UCL Centre for Nephrology, Royal Free Hospital, London, UK

Liviu Segall Department of Nephrology, University of Medicine and Pharmacy ‘Gr. T. Popa’, Iaşi, Romania Jang Won Seo Division of Nephrology, Department of Internal Medicine, Hangang Sacred Heart Hospital, College of Medicine, Hallym University, Korea

Moin A. Saleem University of Bristol, Children’s Renal Unit, Bristol Royal Hospital for Children, Bristol, UK

Sanjeev Sethi Division of Anatomic Pathology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, USA

Richard Sandford Academic Laboratory of Medical Genetics, Addenbrooke’s Treatment Centre, Addenbrooke’s Hospital, Cambridge, UK

Mehmet Şükrü Sever Department of Internal Medicine/ Nephrology, Istanbul School of Medicine, Çapa/ Istanbul, Turkey

Menaka Sarav University of Chicago Pritzker School of Medicine, Chicago, IL; and Division of Nephrology and Hypertension, NorthShore University HealthSystem, Evanston, IL, USA

Gaurav Shah Midwest Heart Specialists/Advocate Medical Group, Naperville, IL, USA

Minnie M. Sarwal California Pacific Medical Center, San Francisco, CA, USA Anjali Bhatt Saxena School of Medicine, Stanford University, Stanford, CA; and Santa Clara Valley Medical Center, San Jose, CA, USA John A. Sayer Institute of Genetic Medicine, Newcastle University, Newcastle, UK

Ron Shapiro Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, PA, USA Neil Sheerin Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK Anushree Shirali Section of Nephrology, Department of Medicine, Yale University, New Haven, CT, USA Seema Shrivastava MRC Centre for Transplantation, King’s College London, London; and St George’s Hospital, London, UK

 

Roslyn J. Simms Institute of Genetic Medicine, Newcastle University, Newcastle, UK

contributors

Bridget Sinnott University of Texas, Dallas, TX, USA

Eugene Teoh Department of Oncology, Oxford Cancer Imaging Centre, University of Oxford, Oxford, UK; and Radiology Department, Oxford University Hospitals NHS Trust, Churchill Hospital, Oxford, UK

Marijn Speeckaert Department of Nephrology, Ghent University Hospital, Gent, Belgium

Bernadette A. Thomas University of Washington, Seattle, WA, USA

Thimoteus Speer Department of Internal Medicine IV, Saarland University Hospital, Homburg/Saar, Germany

Nicola Thomas Public Health, Primary Care and Food Policy Department, School of Community and Health Sciences, City University, London, UK

Stuart M. Sprague University of Chicago Pritzker School of Medicine, Chicago, IL; and Division of Nephrology and Hypertension, NorthShore University HealthSystem, Evanston, IL, USA

Raj Thuraisingham Royal London Hospital, London, UK Marcello Tonelli University of Calgary, Calgary, Canada

Joanna Stachowska-Pietka Institute of Biocybernetics and Biomedical Engineering, Polish Academy of Sciences, Warsaw, Poland

Jonathan N. Townend Department of Cardiology, Queen Elizabeth Hospital and University of Birmingham, Birmingham, UK

Coen A. Stegeman Department of Nephrology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands

Olivier Traxer Université Pierre et Marie Curie, Hôpital Tenon, Service d’urologie, Paris, France

Peter Stenvinkel Division of Renal Medicine, Department of Clinical Science, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden

Francesco Trepiccione Department of Cardio-thoracic and Respiratory Science, Second University of Naples, Naples, Italy

Douglas Stewart Freeman Heart and Vascular Institute, Joplin, MO, USA

Neil Turner Queen’s Medical Research Institute, University of Edinburgh, Edinburgh, UK; and Department of Renal Medicine, Edinburgh Royal Infirmary, UK

Arohan R. Subramanya Renal-Electrolyte Division, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

Andrew A. Udy Department of Intensive Care Medicine, Royal Brisbane and Womens Hospital, Herston, Australia

Rita Suri Division of Nephrology, University of Montreal, London, ON, Canada

Mark L. Unruh Renal-Electrolyte Division, University of Pittsburgh Medical Center, Pittsburgh, PA, USA

Manikkam Suthanthiran Division of Nephrology, Department of Medicine, Department of Transplantation Medicine, Weill Medical College of Cornell University, New York, USA

Ashish Upadhyay Kidney and Blood Pressure Center, Tufts University School of Medicine, Boston, MA, USA

Teena Tandon Division of Nephrology, Department of Medicine, Indiana University School of Medicine, Indiana University, Indianapolis, IN, USA Sydney C. W. Tang Department of Medicine, University of Hong Kong, Queen Mary Hospital, Hong Kong Eric N. Taylor Channing Laboratory, Department of Medicine, Brigham and Women’s Hospital and Harvard Medical School, Boston, MA; and Division of Nephrology and Transplantation, Maine Medical Center, Portland, ME, USA Karthik K. Tennankore Division of Nephrology, University Health Network, Toronto; and University of Toronto, Toronto, ON, Canada Teresa Tenorio Consorcio de investigación del Fracaso Renal Agudo de la Comunidad de Madrid (CIFRA), Madrid; and Instituto Ramón y Cajal de Investigación Sanitaria, Madrid; and Department of Nephrology, Hospital Universitario Ramón y Cajal, Madrid, Spain

Tushar J. Vachharajani Wake Forest University School of Medicine, Section of Nephrology, Winston-Salem, NC, USA Wim Van Biesen Renal Division, University Hospital Ghent, Ghent, Belgium Johan van der Vlag Department of Nephrology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands Raymond Vanholder Renal Division, University Hospital Ghent, Ghent, Belgium Radovan Vasko Department of Nephrology and Rheumatology, Goettingen University Medical School, Goettingen , Germany Nosratola D. Vaziri Division of Nephrology & Hypertension, Departments of Medicine, Physiology & Biophysics, Schools of Medicine and Biological Science, University of California, Irvine, CA, USA

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contributors Anja Verhulst Laboratory of Pathophysiology, University of Antwerp, Antwerp, Belgium Benjamin A. Vervaet Laboratory of Pathophysiology, University of Antwerp, Antwerp, Belgium

I. David Weiner University of Florida College of Medicine, Gainesville, FL; and Nephrology and Hypertension Section,North Florida/South Georgia Veterans Health System, Gainesville, FL, USA

Udo Vester University of Essen, Essen, Germany

Toby Wells Abertawe Bro Morgannwg University Health Board, Radiology Department, Singleton Hospital, Swansea, UK

Katie Vinen King’s College Hospital NHS Foundation Trust, London, UK

Ulrich Wenzel Division of Nephrology, Department of Medicine, University Hospital Hamburg-Eppendorf, Hamburg, Germany

Sobhan Vinjamuri Royal Liverpool University Hospital, Liverpool, UK

Michael J. Weston Department of Radiology, St James’s University Hospital, Leeds, UK

Philip Vladica Departments of Nuclear Medicine and Radiology, Westmead Hospital, Westmead, Australia

Jack F. M. Wetzels Department of Nephrology, Radboud University Medical Center, Nijmegen, The Netherlands

Alexandra Voinescu Division of Nephrology, Saint Louis University, St. Louis, MO, USA

Caroline Whitworth Renal Medicine, Royal Infirmary of Edinburgh, Edinburgh, UK

Luminita Voroneanu Clinic of Nephrology, ‘C. I. Parhon’ University Hospital, ‘Gr. T. Popa’ University of Medicine and Pharmacy, Iasi, Romania

Thorsten Wiech Department of Pathology, University of Hamburg, Hamburg, Germany

Carsten A. Wagner Institute of Physiology, University of Zurich, Zurich, Switzerland

Kate Wiles Guy’s and St Thomas NHS Foundation Trust, and King’s College, London, UK

Stephen Waldek Sale, Greater Manchester, UK

Andrew D. Williams School of Health Sciences, University of Tasmania, Launceston, Australia

Stephen B. Walsh UCL Centre for Nephrology, Royal Free Hospital, London, UK

Christopher G. Winearls Oxford Kidney Unit, Oxford University Hospitals, Oxford, UK

Gerd Walz Renal Division, University Freiburg Medical Center, Freiburg; and BIOSS Center for Biological Signalling Studies, University of Freiburg, Freiburg, Germany

Charles S. Wingo Division of Nephrology, Hypertension, and Transplantation, Department of Medicine, University of Florida College of Medicine, Gainesville; and Nephrology and Hypertension Section, North Florida/South Georgia Veterans Health System, Gainesville, FL, USA

Haiyan Wang† Formerly, Renal Division, Department of Medicine, Peking University First Hospital; and Peking University Institute of Nephrology; and Key Laboratory of Renal Disease, Ministry of Health of China; and Key Laboratory of Chronic Kidney Disease Prevention and Treatment (Peking University), Ministry of Education, Beijing, China Jacek Waniewski Institute of Biocybernetics and Biomedical Engineering, Polish Academy of Sciences, Warsaw, Poland David J. Webb University/British Heart Foundation Centre for Cardiovascular Science, Queen’s Medical Research Institute, University of Edinburgh, Edinburgh, UK Catherine Weber Division of Nephrology, Department of Medicine, McGill University, Montreal, Quebec, Canada Stefanie Weber Pediatric Nephrology, University-Children’s Hospital Essen, Essen, Germany Angela C. Webster Sydney School of Public Health, University of Sydney, Sydney; and Centre for Transplant and Renal Research, Westmead Hospital, Sydney; and Centre for Kidney Research, Kids Research Institute, The Children’s Hospital at Westmead, Sydney, Australia

Paul Winyard Nephro-Urology Unit, UCL Institute of Child Health, London, UK Sarah Withers Cardiovascular Research Group, School of Biomedicine, University of Manchester, Manchester, UK Germaine Wong Sydney School of Public Health, University of Sydney, Sydney; and Centre for Transplant and Renal Research, Westmead Hospital, Sydney; and Centre for Kidney Research, Kids Research Institute, The Children’s Hospital at Westmead, Sydney, Australia Muh Geot Wong Department of Renal Medicine, Kolling Institute of Medical Research, Royal North Shore Hospital, University of Sydney, St Leonards, Australia Terry Wong Guy’s & St Thomas’ Hospital, London, UK Alexander Woywodt Lancashire Teaching Hospitals NHS Foundation Trust, Royal Preston Hospital, Preston, UK

 

Hong Xu Divisions of Baxter Novum and Renal Medicine, Department of Clinical Sciences, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden Muhammad M. Yaqoob Queen Mary University of London, London; and Barts and the London NHS Trust, London, UK Soo Young Yoon Division of Nephrology, Department of Internal Medicine, Kwandong University College of Medicine, Goyang, Korea Nadia Zalunardo Division of Nephrology, University of British Columbia, Vancouver, BC, Canada Klaus Zerres Department of Human Genetics, RWTH Aachen University, Aachen, Germany

†We

contributors

Luxia Zhang Renal Division, Department of Medicine, Peking University First Hospital; and Peking University Institute of Nephrology; and Key Laboratory of Renal Disease, Ministry of Health, Beijing, China Ming-hui Zhao Renal Division, Peking University First Hospital, Institute of Nephrology, Peking University, Key Laboratory of Renal Disease, Ministry of Health of China, Beijing, P. R. China Robert Zietse Division of Nephrology & Transplantation, Department of Internal Medicine, Erasmus Medical Center, Rotterdam, The Netherlands Carmine Zoccali CNR-IBIM Clinical Epidemiology and Pathophysiology of Renal Diseases and Hypertension, Ospedali Riuniti, Reggio Calabria, Italy

record with sadness the death of Professor Hai-Yan Wang on 11 December 2014.

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SECTION 1

Assessment of the patient with renal disease

1 Epidemiology of kidney disease  3 Aminu K. Bello, Marcello Tonelli, and Kitty J. Jager

2 Clinical assessment of the patient with renal disease: overview  20 Christopher G. Winearls

3 Presentations of renal disease  22 Christopher G. Winearls

4 Kidney disease-focused history taking  27 Christopher G. Winearls

5 Kidney disease-focused features on examination  29 Christopher G. Winearls

6 Urinalysis 35 Walter P. Mutter

7 Assessment of renal function  44 Marijn Speeckaert and Joris Delange

8 Tubular function  62 Marijn Speeckaert and Joris Delange

9 Renal radiology: overview  66 Michael J. Weston

10 Ionizing radiation and radiation protection  68 Jeannette Kathrin Kraft and Peter Howells

11 Plain radiography, excretion radiography, and contrast radiography  74 Akira Kawashima and Andrew J. LeRoy

12 Intervention 82 Steven Kennish

13 Ultrasound 89 Toby Wells and Simon J. Freeman

14 Computed tomography  101 Eugene Teoh and Michael J. Weston

15 Magnetic resonance imaging  109 Kazuhiro Kitajima, Akira Kawashima, and James F. Glockner

16 Radioisotopes in diagnostic imaging in nephrology  117 Ramya Dhandapani and Sobhan Vinjamuri

17 Immunological investigation of the patient with renal disease  127 Jo H. M. Berden and Jack F. M. Wetzels

18 The renal biopsy  142 Ian S. D. Roberts, Philip Mason, and Agnes B. Fogo

19 Clinical trials: why and how in nephrology  161 Richard Haynes, Martin J. Landray, William G. Herrington, and Colin Baigent

CHAPTER 1

Epidemiology of kidney disease Aminu K. Bello, Marcello Tonelli, and Kitty J. Jager Basic epidemiology principles in nephrology Introduction Epidemiology is the study of the distribution, determinants, and frequency of disease in populations or settings (Rothman, 1981, 2002). Therefore, epidemiological studies assess the extent of disease, risk/causal factors, natural history, prognosis, prevention/ treatment strategies, and the potential for new policies to prevent disease or improve outcomes (Rothman, 2002). Epidemiological research helps to inform evidence-based medicine by identifying risk factors for disease and to determine optimal treatment approaches; it is the cornerstone of public health research and of preventive medicine. The identification of unbiased causal relationships between exposures (risk factors or interventions) such as hypertension or the use of antihypertensive medication and outcomes like morbidity and mortality is therefore an important aspect of epidemiology. This section will discuss some epidemiological concepts, methods, and their application to clinical research in nephrology.

Research questions Defining an appropriate research question requires familiarity with knowledge gaps in the subject area to judge if a question is feasible, interesting, novel, ethical, and relevant (FINER criteria) (Hulley et al., 2007). Whereas the FINER criteria highlight general aspects of research questions, the development of a specific research question may follow the PICO format, in which P stands for the population (or problem) of interest, I for the intervention (or any other exposure), C for the comparison group, and O for the outcome of interest. Some suggest a PICOT approach, adding a T for the follow-up time to assess outcome (Haynes et al., 2006). An example of a question framed according to PICOT is ‘In dialysis patients (P), what is the effect of statins (I) compared to placebo (C) on cardiovascular mortality (O) after 4 years of follow-up (T)?’ A research question has implications for the choice of the study design—which will in turn determine the analytical methods.

Study designs Fig. 1.1 shows an algorithm for the classification of study designs in clinical research (Grimes and Schulz, 2002). Study design is an important aspect of study quality. Studies can be classified into experimental and observational ones depending on whether or not exposures like therapy were assigned by the investigators. Random allocation of exposures is important to prevent selection bias by the clinician (also known as ‘confounding by indication’) (Stel et  al., 2007)  occurring, when clinicians provide a specific

therapy because of preconceived ideas about which therapy is best. Therefore, when it comes to studies on the effects of therapy or other interventions, randomized controlled trials (RCTs) are the gold standard. This was exemplified by RCTs showing a lack of effect or even harmful effects of using high target haemoglobin level in chronic kidney disease (CKD) patients as opposed to using a lower target (Drueke et al., 2006; Singh et al., 2006), whereas the majority of observational studies had suggested that higher haemoglobin levels were associated with favourable outcomes. On the other hand, observational studies may answer questions on aetiology, diagnosis, prognosis, and adverse effects. In addition, they may provide answers on the effects of therapy where RCTs are not possible, inappropriate, inadequate, or unnecessary (Black, 1996). The effect of transplantation as compared to dialysis cannot be determined through an RCT, as allocation of renal grafts depends on other factors like HLA-matching. Where there is no comparison (control) group (as in case reports or case series), observational studies are called descriptive and where there is a comparison group they are referred to as analytical. Finally, the temporal direction of analytical observational studies determines the type of study. Cohort studies like the Dialysis Outcomes and Practice Patterns Study (DOPPS) determine the exposure of subjects to risk factors at the start of inclusion and then look forward in time to observe the occurrence of outcomes. They may provide a wealth of data which enable the investigator to study not only multiple outcomes but—unlike RCTs—also multiple exposures. In contrast, case–control studies compare cases (those with the disease or other outcome of interest) with controls (those without the outcome of interest) and then look back in time for exposures that might have caused the outcome. In nephrology, case–control studies are uncommon. Nevertheless, this study type is very efficient for studying potential risk factors for rare outcomes that may take a long time to develop, for example, CKD. By going back in time and looking for particular exposures like analgesics one may find associations between outcomes and these exposures. In such a case, prospective cohort studies are less efficient as one will need a very high number of subjects and a very long time to acquire an equal number of cases. Finally, cross-sectional studies examine the presence of an exposure and that of the outcome at the same moment in time. In most cases this simultaneity makes it difficult to determine which is the cause and which is the consequence, in other words, this design may induce a chicken-and-egg problem. Table 1.1 describes the strengths and weaknesses of different study designs (Jager et al., 2007).

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assessment of the patient with renal disease Table 1.1  Strengths and weaknesses of frequently used study designs

Did investigator assign exposures?

Study design No

Yes Experimental study

Observational study

Random allocation?

Comparison group?

Yes

No

Randomized controlled trial

Exposure

Nonrandomized controlled trial

Analytical study

◆

First form of publication for new diseases, rare adverse events, or manifestations of disease ◆ Fast and inexpensive ◆ Hypothesis generating

◆

Cross-sectional study

◆

Can assess prevalence and burden of disease ◆ Fast and inexpensive ◆ Hypothesis generating

◆

Case–control study

◆

Efficient study design Very suitable for studying rare outcomes and outcomes that take a long time to develop ◆ Can study multiple exposures ◆ Relatively inexpensive ◆ Hypothesis generating

◆

Cohort study

◆

Can study multiple exposures, uncommon exposures and multiple outcomes ◆ Hypothesis generating

◆

Randomized controlled trial

◆

◆

No Descriptive study

Exposure and outcome at the same time Outcome Case–control study

Crosssectional study

Fig. 1.1  Algorithm for the classification of study designs in clinical research. Reprinted from The Lancet, 359(9300), Grimes DA, Schulz KF, An overview of clinical research: the lay of the land, 57–61, 2002, with permission from Elsevier.

Knowledge derived from different studies published on a specific topic can be summarized in a systematic review (Noordzij et  al., 2009). In contrast to narrative reviews, systematic reviews use explicit and reproducible methods for searching the literature and a critical appraisal of individual studies. This systematic methodology minimizes bias. Sometimes systematic reviews include a meta-analysis. This is a mathematical synthesis of the results of those individual studies in which more weight is given to results of studies with more events and sometimes to studies of higher quality. Criteria for the reporting of studies using particular designs have been summarized in statements like CONSORT (Schulz et al., 2010) for RCTs, STROBE (von Elm et al., 2007) for observational studies, and PRISMA (Moher et  al., 2009)  and MOOSE (Stroup et  al., 2000)  for their respective meta-analyses. Such statements assist authors in writing such reports, help editors and peer reviewers in reviewing manuscripts for publication, and aid readers in critically appraising the quality of published studies.

Weaknesses

Case report/ case series

Direction?

Outcome

Exposure Cohort study

Yes

Strengths

◆

Important potential to make causal inferences

Very limited potential to make causal inferences, unless in dramatic cases ◆ Selection bias

Very limited potential to make causal inferences, because the time order of exposure and outcome cannot be determined ◆ Selection bias ◆ Survival bias Some potential to make causal inferences ◆ Can study only one outcome at the time ◆ Choice of controls needs careful attention ◆ Selection bias ◆ Recall bias Some potential to make causal inferences ◆ If done prospectively, more expensive ◆ If done prospectively, may take a long time to complete ◆ Selection bias Can study multiple outcomes, but only one exposure ◆ Very expensive ◆ Limited generalizability when making use of restrictive in- and exclusion criteria ◆ Selection bias

Reprinted by permission from Macmillan Publishers Ltd: Kidney Int. Jager KJ, Stel VS, Wanner C, Zoccali C, Dekker FW. The valuable contribution of observational studies to nephrology, 72(6):671–5, 2007.

Measures of disease occurrence Different measures may be used to describe how often a disease (or other health outcomes) occurs in a population (Jager et al., 2007). Incidence expresses the development of new cases and is mostly used against the background of prevention, to assess disease aetiology or to determine risk factors. Depending on the study question, incidence may be reported as risk or as incidence rate. The latter is preferred when studying a dynamic population or when the observation period is sufficiently long for competing risks (like death from causes other than the outcome under investigation) or loss to follow-up to play a significant role. Determining the incidence rate of

a treatment like renal replacement therapy (RRT) is straightforward. When determining the incidence of a chronic condition like CKD, problems may arise, as it is unfeasible to identify newly developed CKD in all individuals who were initially free from this disease. Prevalence on the other hand is the number of existing cases. It reflects the burden of the disease in a population and can be used for the planning of healthcare facilities. Again, the assessment of the prevalence of a treatment like RRT is relatively uncomplicated. However, the assessment of the prevalence of stages 3–5 of CKD according to the National Kidney Foundation Kidney Disease

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Outcomes Quality Initiative (NKF-KDOQI) 2002 and Kidney Disease:  Improving Global Outcomes (KDIGO) 2004 definition only needs an estimated glomerular filtration rate (eGFR), whereas that of stages 1–2 CKD also requires albuminuria testing. Finally, measures of disease occurrence are used to study causes of disease or effects of risk factors. One way to estimate the size of such an effect is the calculation of the ratio between the disease occurrence (e.g. cardiovascular disease) in those being exposed to the risk factor (e.g. diabetes mellitus) and those not exposed to the risk factor. The resulting ‘relative risk’ is an example of this measure of effect.

epidemiology of kidney disease

Table 1.2  Types of regression analysis Outcome variable (dependent variable)

Exposure variable (independent variable)

Measure of effect

Linear

Continuous

Continuous or categorical

Relative risk

Logistic

Categorical

Continuous or categorical

Odds ratio

Survival analysis

Time-to-event

Continuous or categorical

Hazard ratio

Bias and confounding Bias is a systematic error in the design or conduct of a study (Tripepi et al., 2008). It may affect study validity in several ways, for example, through methods used by the investigator in recruiting study subjects or through factors affecting study participation (selection bias) or through systematic distortions in the collection of exposures or outcomes (information bias). Selection bias may, for example, occur when investigators performing an RCT use very strict inclusion criteria so that only relatively healthy subjects will be included in the study. The study results in this ‘selected’ group may not necessarily be generalized to very sick patients. Information bias on the other hand may, for instance, be induced when inaccurate instruments systematically over- or underestimate exposures like blood pressure or when in case–control study diseased individuals (cases) much better remember potentially harmful exposures than non-diseased individuals (controls). The latter is called recall bias. Confounding is a ‘blurring’ of effects, obscuring the ‘real’ effect of an exposure on outcome (Jager et al., 2008). For example, the inverse association between total cholesterol level and mortality in dialysis patients was likely the result of confounding by systemic inflammation and malnutrition (Liu et  al., 2004). For this reason, in aetiological studies, investigators do their best to prevent confounding, for example, by randomization, or to control confounding. A  frequently used method to control for confounding is including the confounding variable into multivariate analysis. However, before a variable qualifies as a confounder it should fulfil three criteria: (1) be a risk factor for the outcome studied, (2) be associated with the exposure, and (3) not be an effect of the exposure (be in the causal pathway). The last criterion is important to avoid ‘over-adjustment’ through which an investigator may take away part of the real effect of an exposure and may introduce bias instead of preventing it. For example, in a study on the association between body mass index (BMI) and the risk of ESRD (Hsu et al., 2006) the authors were right in not adjusting for blood pressure, as blood pressure can be considered in the causal pathway between BMI and end-stage renal disease (ESRD). Therefore, before including them in a multivariate analysis, the criteria for confounding should be checked carefully for every variable in every association studied.

Regression analysis This statistical technique describes the dependence of the outcome (‘dependent’) variable from the value of one or more exposure variables (‘independent’ variables). Regression can be used for univariate analysis (without controlling for confounding) and multivariate analysis (with adjustment for confounding). Table 1.2

shows that the choice for the type of regression analysis depends on the type of outcome studied, but not on the type of exposure variables that may be included in the analysis. Linear regression analysis demands that the outcome variable is continuous, for example, the 3-year risk of cardiovascular events. Logistic regression, on the other hand, is needed to study categorical outcomes such as whether or not patients are compliant with therapy prescription. Finally, survival analysis is used to study ‘time-to-event’ data like time to death, to first renal transplant, or to myocardial infarction. In survival analysis, the survival times of subjects not experiencing the event of interest are being censored. The Kaplan–Meier method, being the most popular method for survival analysis, provides estimates of survival probabilities and may compare survival between groups. The method can, however, only study the effect of one exposure at the time and it cannot provide an effect size. Multivariate survival analysis and effect estimation therefore need other techniques like Cox proportional hazards regression. The measure of effect of Cox regression is the hazard ratio. Both the odds ratio and the hazard ratio can be interpreted as relative risks.

Screening Investigating apparently healthy individuals to detect unrecognized early stages of disease allows measures to be taken to prevent or slow down progression and reduce (premature) death in those affected (Grootendorst et al., 2009). In 1968, the World Health Organization (WHO) published ten criteria to facilitate the selection of conditions that are suitable for screening (Wilson and Jungner, 1968) which have been supplemented with additional criteria thereafter (Andermann et al., 2008). These criteria should be taken into account when considering screening for conditions like CKD. Evaluation of screening programmes may suffer from bias. For example, those who volunteer to be tested for CKD may have a family history of kidney disease resulting in a high detection rate. In addition to this form of selection bias, those who are diagnosed by screening more frequently include those with slowly progressive disease, as those with poor prognosis are less likely to be picked up by screening because of their higher mortality risk. This length bias will provide a ‘better’ prognosis to those identified in a screening programme, even if screening has no effect on prognosis. Similarly, earlier diagnosis through screening may lead to an apparent increase in survival time. This phenomenon is known as lead-time bias (Grootendorst et al., 2009).

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Epidemiology of acute kidney injury (See also Chapter 220.)

Definition, classification, and evaluation of acute kidney injury Acute kidney injury (AKI) is a clinical syndrome characterized by a sudden onset of reduced kidney function, and is manifested by increased serum creatinine (SCr) or a reduction in urine output (Hou and Cohen, 1985; Bellomo et  al., 2004; Lameire et  al., 2005; Cerda, 2008). Until recently, there was no standard definition for AKI, leading to highly variable estimates of the incidence, prevalence, and prognosis of this syndrome (Lameire et al., 2005). Prior to 2004, the key definition in use was based on the WHO International Classification of Disease (ICD) codes, which are used mainly for administrative purposes in the healthcare systems of developed countries. Criteria based on various SCr values and/or the receipt of dialysis were also commonly used (especially by clinicians in hospital settings (Mehta and Chertow, 2003; Lameire et al., 2006))—although such definitions vary widely across settings, populations and institutions (Mehta and Chertow, 2003; Mehta et al., 2007). The absence of a standardized definition of AKI led to the development of consensus criteria by the Acute Dialysis Quality Initiative (ADQI), which are known as the Risk, Injury, Failure, Loss, and End-stage renal disease (RIFLE) 2004 criteria (Van Biesen et al., 2006; Mehta et al., 2007). The RIFLE scheme is based on two important parameters: changes in SCr or eGFR from baseline and urine output at specific points in time (Table 1.3) (Mehta et al., 2007). This classification was subsequently modified and the term ‘acute renal failure’ was replaced with ‘acute kidney injury’ to cover the entire spectrum of acute renal dysfunction from mild changes in renal function to the use of dialysis (Table 1.3) (Van Biesen et al., 2006). This standardization of terminology facilitates comparison between settings. However, the central role of quantitative changes in urine output in the scheme may make it most applicable to critical care units or other closely monitored settings. Although the RIFLE criteria were validated in over half a million AKI patients globally (Cerda et al., 2008b; Cruz et al., 2009), the system was subsequently revised by another international group (the Acute Kidney Injury Network (AKIN)). The AKIN scheme uses slightly broader criteria than the RIFLE system, and reflects the clinical significance of relatively small rises in SCr (Table 1.3) (Mehta et al., 2007; Cruz et al., 2009). Of note, it appears that RIFLE may be slightly better than AKIN for risk prediction and prognostication (Ricci et al., 2008). To reconcile these minor differences, the International Society of Nephrology (ISN) and KDIGO (Anonymous, 2012), has brought together international experts from multiple specialties to harmonize the RIFLE and AKIN criteria and produce a truly global definition and staging system (Anonymous, 2012)—further enabling future comparisons of the incidence, outcomes, and efficacy of therapeutic interventions for AKI (Table 1.3).

Limitations and pitfalls for AKI definition It is important to recognize that no single definition of a clinical syndrome is ideal; the purpose of establishing a consensus definition is to establish the presence/absence of disease, provide an index of severity, and relate these to prognosis and outcome

Table 1.3  Definitions/classification for AKI Classifications Criteria

Urine output criteria

KDIGO Stage 1

Increase ≥ 26 μmol/L within 48 h or increase ≥ 1.5 to 1.9 × reference SCr

< 0.5 mL/kg/h for > 6 consecutive h

Stage 2

Increase ≥ 2 to 2.9 × reference SCr

< 0.5 mL/kg/h for > 12 h

Stage 3

Increase ≥ 3 × reference SCr or increase 354 μmol/L or commenced on RRT irrespective of stage

< 0.3 mL/kg/h for > 24 h or anuria for 12 h

Risk

Increased creatinine × 1.5 or GFR decrease > 25%

< 0 .5 mL/kg/h × 6 h

Injury

Increased creatinine × 2 or GFR decrease > 50%

< 0 .5 mL/kg/h × 12 h

Failure

Increase creatinine × 3 or GFR decrease > 75% or creatinine 4 mg/dL (acute rise of 0.5 mg/dL)

< 0.3 mL/kg/h × 24 h (oliguria) or anuria × 12 h

Loss

Persistent ARF = complete loss of renal function > 4 weeks

ESRD

End-stage renal disease

RIFLE

AKIN Stage 1

Increase ≥ 26 μmol/L within 48 h or increase ≥ 1.5–1.9 × reference SCr

< 0.5 mL/kg/h for ≥ 6 h

Stage 2

Increase ≥ 2–2.9 × reference SCr

< 0.5 mL/kg/h for ≥ 12 h

Stage 3

Increase ≥3 × reference SCr or increase≥ 354 μmol/L with an acute rise of at least 44 μmol/L or initiation of RRT

< 0.3 mL/kg/h for ≥ 24 h or anuria ≥ 12 h

ICD 9 codes ARF

ARF without dialysis Any of the following: 584.5: ARF, with lesion of tubular necrosis 584.6: ARF, with lesion of renal cortical necrosis 584.7: ARF, with lesion of renal medullary (papillary) necrosis 584.8: ARF, with other specified pathologic lesion in kidney

ARF-D

ARF with dialysis: ARF code as above plus any of the following codes: 584.9: ARF, unspecified V39.95: haemodialysis V45.1: renal dialysis status (patient requires intermittent renal dialysis; presence of arteriovenous shunt) V56.0: extracorporeal dialysis (dialysis (renal) not otherwise specified) V56.1: fitting and adjustment of extracorporeal dialysis catheter

ARF = acute renal failure; ARF-D = acute renal failure with dialysis; GFR = glomerular filtration rate; KDIGO = Kidney Disease: Improving Global Outcomes; RRT = renal replacement therapy; SCr = serum creatinine.

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(Rothman, 2002). In addition to lack of standardization, definitions used before RIFLE/AKIN did not facilitate detection of milder forms of AKI—which are highly prevalent and also associated with adverse clinical outcomes (Yong et al., 2011). Despite the advantages of the RIFLE/AKIN criteria, these definitions also have some limitations (Van Biesen et al., 2006; Cerda et al., 2008b; Yong et al., 2011). First, the use of 6-hour and 12-hour urine output criteria make RIFLE difficult to use for retrospective studies of AKI, since such data are not routinely collected (Van Biesen et al., 2006; Srisawat et al., 2010). Second, there is not necessarily any correlation between the SCr and urine output criteria for severity of AKI, as patients often present with high SCr but good urine output or vice versa. Third, SCr/eGFR criteria are based on change from a baseline value (which is not always available). Fourth, eGFR is valid only in steady-state conditions—which is certainly not the case in AKI (Srisawat et al., 2010). Fifth, recent studies have shown that smaller changes in SCr than those specified by the RIFLE criteria (such as an absolute increase of 0.3 mg/dL) are also associated with poor outcomes (Srisawat et al., 2010). These limitations have been partially addressed by AKIN (Van Biesen et al., 2006; Srisawat et al., 2010)—which has eliminated the eGFR criterion and explicitly recognized the adverse prognostic values associated with very small changes in SCr (Van Biesen et al., 2006).

Risk factors and causes for AKI A partial list of predisposing factors (risk factors) and causes for AKI is shown in Fig. 1.2; the incidence and likelihood of which vary according to the clinical setting (hospital vs community; developed vs developing countries) (Joannidis and Metnitz, 2005; Lameire et al., 2005, 2006). For instance, acute tubular necrosis (ATN) due to sepsis is the most common cause of AKI in the critical care setting, accounting for up to 35–50% of all cases of AKI (Lameire et al., 2005). In a large-scale study, the Madrid Acute Renal Failure Study in Spain, ATN was the cause of AKI in 75.9% of intensive care unit (ICU) patients compared to 37.6% in non-ICU patients (Liano and Pascual, 1996). ATN in hospitalized patients is more likely to be multifactorial, with hypotension and nephrotoxins as important causes in addition to sepsis and surgery (Cerda et al., 2008b). For community-acquired AKI, prerenal or acute-on-chronic renal failure is common and usually occurs as the result of dehydration or drug-induced causes as seen with the use of non-steroidal anti-inflammatory drugs (NSAIDs), angiotensin-converting enzyme inhibitors (ACEI), and angiotensin receptor blockers (ARBs) (Cerda, 2008; Cerda et  al., 2008b). Elderly people and patients with multiple comorbidities such as diabetes are at particularly high risk of developing community-acquired AKI (Cerda, 2008)  (Fig. 1.2). AKI is being conceptualized as a spectrum as obtained in CKD from normal to increased risk, decreased function to kidney failure, and/or death (Fig. 1.3).

Descriptive epidemiology of AKI (See also Chapter 220.) There is a paucity of high-quality population-based data on the incidence of AKI—whether community- or hospital-acquired (Cerda et  al., 2008b; Yong et  al., 2011). Before the advent of the RIFLE/AKIN classification systems, the reported incidence of AKI varied from 1% to 30% across studies (Lameire et al., 2005; Waikar et al., 2008). As reviewed by Waikar et al. (2008), the incidence of AKI also shows considerable variation based on the clinical setting

epidemiology of kidney disease

(Lameire et al., 2005, 2006). The prevalence of hospital-acquired AKI is reported to be about five to ten times greater than community-acquired AKI, with reported rates of AKI of 5–7% of hospitalized patients (Hsu et  al., 2007; Waikar et  al., 2008; Yong et al., 2011). In country-specific data, the reported prevalence of AKI in the United States ranges from 1% (community acquired) to 7.1% (hospital acquired) of all hospital admissions (Lameire et al., 2005, 2006). The population incidence of AKI from UK data ranges from 172 pmp to 486–630 pmp (Yong et al., 2011). Early studies from the 1990s reported that the annual incidence rates of community-acquired AKI varied from 22 to 620 pmp, with most studies using the receipt of dialysis or SCr ≥ 300 or 500μmol/L to define AKI (Lameire et al., 2005; Yong et al., 2011). The reported incidence of AKI varies considerably and depends on how AKI was defined, the setting where it occurred, population studied, and also whether patients with de novo CKD developing AKI were included. For instance, the reported incidence varies from 140 to 620 pmp across countries (Feest et  al., 1993; Liano and Pascual, 1996; Stevens et al., 2001; Waikar et al., 2006). In the United Kingdom, the reported incidence was 620 pmp/year among patients with SCr ≥ 300 µmol/L (Stevens et al., 2001) and 140 pmp/year in those with SCr > 500 µmol/L (Feest et al., 1993). In Spain, the incidence of AKI was 209 pmp from data of patients in tertiary care hospitals and SCr > 177 µmol/L (Liano and Pascual, 1996). In the United States, the reported incidence was 288 pmp, but ICD-9 codes rather than SCr criteria were used to define AKI (Waikar et al., 2006). The incidence of community-acquired AKI appears to be increasing:  the incidence of non-dialysis-dependent AKI and dialysis-dependent AKI increased from 322.7 to 522.4 and 19.5 to 29.5 per 100,000 person-years, respectively, between 1996 and 2003 (Hsu et al., 2007). The rising incidence of community-based AKI over time likely reflects the ageing general population, the increasing prevalence of chronic comorbidity (including CKD), and increasing utilization of nephrotoxic agents (such as intravenous radiocontrast, aminoglycosides, NSAIDs, ACEIs/ARBs, and chemotherapeutic agents) among outpatients (Cerda, 2008; Yong et al., 2011). In ICU settings, an estimated 5–20% of patients experience an episode of AKI during the course of their illness, and AKI accounts for nearly 10% of all ICU bed-days (Joannidis and Metnitz, 2005; Yong et al., 2011). However, AKI usually coexists with other acute illnesses, since the incidence of AKI as single-organ failure in ICU patients is as low as 11% compared to 69% in non-ICU settings. There have been several studies on epidemiology of AKI in various settings (Waikar et al., 2008).

AKI in special populations (See also Chapters 239–241.) There are major differences in the epidemiology of AKI between geographic regions (especially in developing vs developed nations), and also across sociodemographic categories defined by age (elderly, children), gender, and social deprivation (Lameire et al., 2005, 2006, 2013; Zappitelli et al., 2008). Numerous reviews outline differences and similarities in the causes and consequence of AKI between developed and developing regions, analyse the practical implications of the identified differences, and make recommendations for management (Lameire et al., 2005, 2006, 2013). In developing countries, the reported incidence of hospital-based AKI is much lower than in developed countries (Lameire et al., 2006), likely because AKI is under-recognized in developing countries (Lameire et  al.,

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Social demographics Old age Female Non-white Low SES

Risk Factors

Co-morbidities CVD (MI, PVD, HV arrhythmias Anaemia Chronic lung diseases Active cancer Hypoalbuminaemia sepsis Liver disease Vascular Renovascular disease Malignant hypertension Scleroderma Atheroemboli Microangiopathics (HUS, TTP) Vasculitides

AKI

Community

Health center

Medications Contrast agents NSAIDs Calcineurin inhibitors ACEI/ARBs Antibioitics/Antifungals (aminoglycosides, amphotericin B) Chemotherapy (methotrexate, cisplatin)

Hospital

Infections Acute infections (eg. streptococcal Chronic infections

Intensive care Poisonings Snake/bee stings Herbal remedies Insecticides

Non-intensive care

Environmental Heat stroke Natural disasters (eg. tsunamis) Pregnancy Septic abortions Pre-eclampsia/eclampsia Genetics G-6-P deficiency Mechanical BPH Neurogenic bladder Intra/extra ureteric obstruction Retroperitoneal fibrosis

Fig. 1.2  Risk factors for AKI.

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epidemiology of kidney disease

Conceptual model of acute kidney injury (AKI) ANTECENDENTS INTERMEDIATE STAGE AKI OUTCOMES

NORMAL

INCREASED RISK

COMPLICATIONS

DAMAGE

DECREASED GFR

KIDNEY FAILURE

DEATH

Cerdá J et al. CJASN 2008;3:881–886

©2008 by American Society of Nephrology

Fig. 1.3  Conceptual model for AKI. Drawn from Cerda, J., Lameire, N., Eggers, P. et al. Epidemiology of acute kidney injury. Clin J Am Soc Nephrol, 2008; 3:881–886.

2006; Cerda et  al., 2008a) due to reduced access to care, delayed referral to specialist services, and lack of dialysis services (Cerda et al., 2008a). Patients with AKI in developing nations tend to be substantially younger, are more likely to have infection-related glomerulonephritis, and are more likely to be female compared with those in the developed world (Lameire et al., 2006; Cerda, 2008)— the latter may be partially due to a high incidence of obstetrics complications such as septic abortion (Cerda et al., 2008a). In addition, seasonal increases in the risk of AKI may be observed in developing nations—due to corresponding variation in the risk of malaria, heat stroke, animal envenomation, and diarrhoeal diseases (Lameire et al., 2006; Cerda et al., 2008a; Lameire et al., 2013). Thus, the prerenal and toxic factors predominate as AKI risk factors in developing countries (Lameire et al., 2006). Preventive opportunities are often missed because of failure to recognize the risk factors and late presentation for treatment (Lameire et al., 2006). For instance, a firm aetiology for AKI cannot be established in many instances because of a lack of appropriate laboratory and technical support (Lameire et al., 2006, 2013). Regardless of the setting, most cases of AKI in children are secondary to prerenal causes from acute dehydration, major surgeries, and sepsis (Lameire et al., 2005). In contrast, the elderly are more susceptible to AKI due to multiple comorbidities and lower renal functional reserve associated with ageing (Lameire et al., 2006; Yong et al., 2011).

Prognosis and outcomes (See Chapter 237.) AKI in hospitalized patients is associated with poor prognosis, and mortality ranges from 10% to 80% depending on the population studied (Waikar et  al., 2008). Patients who present with uncomplicated AKI have a mortality of < 10% (Waikar et al., 2007). In contrast, patients presenting with AKI and other organ failure have been reported to have short-term mortality of > 50% (Ricci et al., 2008; Waikar et al., 2008; Alkandari et al., 2011; Yong et al., 2011; Singbartl and Kellum, 2012), especially if acute dialysis is required, which may be associated with short-term mortality of up to 80% (Lameire et al., 2005; Yong et al., 2011). As in CKD, lower

eGFR and heavier proteinuria independently increase the risk of AKI. However, the excess risk of mortality associated with an episode of AKI is actually lower among people with lower baseline eGFR and heavier proteinuria, as compared to those with normal kidney function (James et al., 2010).

Epidemiology of chronic kidney disease Definition, classification, and evaluation of CKD (See Chapter 94.) As for AKI, several terminologies such as ‘chronic renal failure’, ‘chronic renal insufficiency’, ‘chronic kidney failure’, ‘progressive renal failure or insufficiency’, and ‘pre-dialysis’ were used in the past to denote CKD, with further subclassification mainly by the underlying aetiology (Anonymous, 2002, 2007; Levey et al., 2011; Levey and Coresh, 2012). In 2002, the NKF-KDOQI released guidelines for the diagnosis and classification of CKD (Anonymous, 2002). These criteria standardized the nomenclature of CKD and the laboratory evaluation of kidney disease, documented the associations between level of kidney function and multiple complications, and facilitated risk stratification for adverse outcomes of CKD (Anonymous, 2002). The KDOQI criteria defined CKD by structural or functional abnormalities of the kidney for at least 3 months, manifested by either kidney damage (often persistent albuminuria) with or without a decreased eGFR to a value < 60 mL/min/1.73m2, or a decreased eGFR with or without other evidence of kidney damage for at least 3 months (Anonymous, 2002) (Box 1.1). This staging was later modified and endorsed by the international KDIGO in 2004 according to severity and treatment (Levey et al., 2011). This was further updated in 2012 following a decade of further research and practice since the publication of the initial guidelines (Stevens et al., 2013) (Table 1.4). The new classification system now incorporates a three-dimension operational definition for CKD that includes cause, GFR category, and albuminuria category (CGA). The underlying cause of CKD was added to the definition in order to highlight the importance of this information for management and prognostication (Table 1.5). Offsetting these

9

10

Section 1  

assessment of the patient with renal disease

Box 1.1  Stages of CKD by NKF-KDOQI definition ◆ Stage 1: patients with normal glomerular filtration rate (GFR), but some evidence of kidney damage as manifested by albuminuria/proteinuria, haematuria, or histological changes ◆ Stage 2: mild CKD characterized by GFR of 89–60 mL/min/1.73 m2 with some evidence of kidney disease as manifested by albuminuria/proteinuria, haematuria, or histological changes ◆ Stage

3: moderate CKD with GFR 59–30

mL/min/1.73 m2

Table 1.5 Classificationa of CKD based on presence or absence of systemic disease and location within the kidney of pathologic anatomic findings Examples of systemic diseases affecting the kidney

Examples of primary kidney diseases (absence of systemic diseases affecting the kidney)

Glomerular diseases

Diabetes, systemic autoimmune diseases, systemic infections, drugs, neoplasia (including amyloidosis)

Diffuse, focal or crescentic proliferative GN; focal and segmental glomerulosclerosis, membranous nephropathy, minimal change disease

Tubulointerstitial diseases

Systemic infections, autoimmune, sarcoidosis, drugs, urate, environmental toxins (lead, aristolochic acid), neoplasia (myeloma)

Urinary-tract infections, stones, obstruction

Vascular diseases

Atherosclerosis, hypertension, ischemia, cholesterol emboli, systemic vasculitis, thrombotic microangiopathy, systemic sclerosis

ANCA-associated renal limited vasculitis, fibromuscular dysplasia

Cystic and congenital diseases

Polycystic kidney disease, Renal dysplasia, Alport syndrome, Fa bry medullary cystic disease, disease podocytopathies

◆ Stage 4: severe CKD with GFR 29–15 mL/min/1.73 m2 ◆ Stage 5: CKD with GFR < 15 mL/min/1.73 m2, or where patient survival depends on provision of RRT in the form of dialysis or transplant. potential advantages are challenges associated with ascertaining the cause of CKD, especially in primary care. The addition of albuminuria to the definition has substantial benefits for prognostication and will undoubtedly enhance the utility of the new scheme. The evolution of these classification schemes over the last decade reflects our more refined understanding of CKD epidemiology. This trend is likely to continue in the future with identification of better markers for CKD and refinement in molecular diagnostic techniques. Although less sensitive than definitions based on SCr or eGFR, ICD-9-CM codes have been used to classify CKD (Anonymous, Table 1.4  KDIGO Classification of CKD 2012 (Stevens et al 2013; Anonymous 2013) Category

GFR, mL/ AER ACR Equivalent, Descriptor min 1.73 m2 mg/d mg/g

GFR

–

–

ANCA, antineutrophil cytoplasmic antibody; CKD, chronic kidney disease, GN, glomerulonephritis Genetic diseases are not considered separately because some diseases in each category are now recognized as having genetic determinants.

G1

≥90

–

–

Normal or high

a Note that there are many different ways in which to classify CKD. This method of

G2

60–89

–

–

Mildly decreased*†

separating systemic diseases and primary kidney diseases is only one, proposed by the Work Group, to aid in the conceptual approach.

G3a

45–59

–

–

Mildly to moderately decreased

G3b

30–44

–

–

Moderately to severely decreased

G4

15–29

–

–

Severely decreased

Definition of CKD stage 5 and ESRD

G5

2200 mg/d [ACR >2220 mg/g])

◆ 585.5: chronic kidney disease, stage 5

Modified by NT 7th Aug 2015

◆ 585.6: end-stage renal disease

(Note that the other table on the page submitted with proof corrections was unchanged table 1.5 - NT)

◆ 585.9: chronic kidney disease, unspecified.

chapter 1 

(Levey et al., 2011). CKD stage 5 and ESRD are therefore not synonymous because not all patients with CKD stage 5 receive RRT, some patients with asymptomatic stage 4 CKD (GFR 15–29.9 mL/ min/1.73 m2) may receive RRT, and kidney transplant recipients often have an eGFR > 15 mL/min/1.73 m2. Most available data on the epidemiology of ESRD are derived from registries of patients receiving chronic RRT. Because of different access to RRT across regions and healthcare systems, the incidence and prevalence of ESRD may not be fully captured by renal registries, which actually focus on ‘receipt of RRT’ (discussed below).

Pitfalls and limitations of the definition/staging system for CKD The NKF-KDOQI classification scheme has been widely adopted around the world (Eckardt et al., 2009; Ikizler, 2009; Gansevoort and de Jong, 2010). This important initiative permitted a common nomenclature for clinicians and researchers, and facilitated an international effort to educate the public about the significance of CKD. Despite these benefits, the scheme also has limitations. For example, individuals with only minimal functional or radiological abnormalities might be classified with stage 1 CKD despite the uncertain clinical relevance of these findings; elderly individuals with only mild reductions in eGFR could be classified as having stage 3 CKD—raising concerns about inappropriate labelling of healthy individuals as diseased (Chen and Hsu, 2003). Moreover, the prognosis of earlier stage disease is not necessarily better than those with more advanced stages, and progression from lower to higher stages is not necessarily inevitable. In fact, little is known about the natural history of stages 1 and 2 CKD and increasing evidence is accruing that a substantial proportion of patients with stages 3 and 4 CKD have stable eGFR (Keith et al., 2004). It has been suggested as more appropriate to divide patients into those with certain urinary abnormalities such as isolated haematuria or microalbuminuria and those with impaired kidney function (Winearls and Glassock, 2009). The latter may warrant subclassifications based on the presence or absence of progression and associated risk factors such as hypertension and proteinuria (Glassock and Winearls, 2008a; Winearls and Glassock, 2009). Guidelines from the United Kingdom by the National Institute for Health and Care Excellence (NICE) have attempted to address some of these criticisms by dividing CKD stage 3 into 3A and 3B (eGFR of 45–59 and 30–55 respectively), and the addition of the suffix ‘p’ to CKD stages to denotes significant proteinuria (Anonymous, 2007). These issues have been the subject of vigorous debate in recent years (Levey et al., 2011), and revision to the NKF-KDOQI scheme by an international KDIGO working group has recently been published (Anonymous, 2013).

Risk factors and causes for CKD A large body of epidemiological and clinical evidence has linked certain risk factors to the initiation and progression of CKD. These can be classified into distinct categories based on the presence or absence of established causation; factors that have been proven to be causal (risk factors) and those that are associated with CKD in the absence of established causal relationship (risk markers) (Anonymous, 2002; Levey et al., 2011). Causes of CKD include diabetes mellitus, hypertension, ischaemia, infection, obstruction, toxins, and autoimmune and infiltrative diseases (Anonymous, 2002; Levey et  al., 2011). Although

epidemiology of kidney disease

it is important to identify the cause(s) of CKD so that specific therapy can be instituted, adverse outcomes (including cardiovascular events and progression to ESRD) often occur despite appropriate treatment and irrespective of the underlying cause (Levey et al., 2011). Risk factors for development of CKD have traditionally been classified as susceptibility factors and initiation factors. Susceptibility factors increase susceptibility to CKD, for example, older age, male gender, and familial/genetic predisposition. Initiation factors directly initiate kidney damage; such factors include diabetes, hypertension, chronic infections, drugs, and toxins. The progression factors are risk factors associated with worsening of already established kidney damage, such as high levels of proteinuria, hypertension, poor glycaemic control in diabetes, obesity, and smoking (Anonymous, 2002, 2007; Levey et al., 2011). However, the difficulty of detecting the early stages of CKD makes it difficult to determine whether the risk factors so far identified in the population relate more to susceptibility, initiation, or even progression. Therefore, this traditional classification may be somewhat artificial. These risk factors may also be subclassified as potentially modifiable/preventable and non-modifiable (Table 1.6). Finally, risk factors can also be classified as clinical (diabetes, hypertension, autoimmune diseases, systemic infections, drugs) or sociodemographic (age, race, poverty/low income, toxins).

Global CKD epidemiology Initial population-based estimates of the prevalence of CKD were obtained from the National Health and Nutrition Examination Survey (NHANES) in the United States. The prevalence of eGFR 60 mL/min (Glassock and Winearls, 2008a). Some of these limitations are being addressed with development of more accurate and precise estimating equations—for instance, the CKD Epidemiology Collaboration (CKD-EPI) equation performs better at higher true GFR, and leads to slightly lower population-based estimates of CKD prevalence. For example, using the CKD-EPI equation rather than the MDRD equation reduced the estimated prevalence of CKD in the United States to 11.5% (95% confidence interval (CI), 10.6–12.4%) from 13.1% (CI, 12.1–14.0%) using the MDRD equation (Matsushita et al., 2010a; Stevens et al., 2010).

CKD in special populations The epidemiology of CKD may differ in special populations such as ethnic minorities, aboriginal people, Roma, the homeless, and children (Esbjorner et al., 1997; Ardissino et al., 2003; Harambat et al., 2012)—perhaps due to differences in genetic, behavioural, and sociodemographic characteristics (Foster, 2008; Huong et al., 2009; Hoy et al., 2010). There have been several population-wide surveys on the prevalence of CKD in indigenous people particularly in Australia and Canada (Gao et al., 2007; Hoy et al., 2010). Aboriginal people tend to have a higher burden of CKD than found in the general population (Katz et al., 2006; Gao et al., 2007; Hoy et al., 2010). For instance, on a remote Aboriginal island community, 26% of adults had microalbuminuria and 24% had overt albuminuria when screened in 1992–1995 (Hoy et al., 1998). Data on CKD epidemiology among ethnic black people is predominantly US based (KEEP, 2002) and information about the incidence and prevalence of CKD in Africa is sparse. In both the United States and the few communities in Africa where studies were conducted, black people tend to have a higher burden of CKD than white people. For instance, in a community survey conducted in adults attending primary care clinics in Soweto, 35% of the people screened had proteinuria; 10% needed referral to a tertiary hospital, and of these, most had stages 3–5 CKD

epidemiology of kidney disease

(Katz et al., 2006). A recent conservative estimate on prevalence of CKD in a sub-Saharan African country was 36% (Sumaili et al., 2009), as compared to 13.1% in the United States (Coresh et al., 2007).

CKD screening: measures and utility Currently, screening for CKD is accepted practice only in people with hypertension or diabetes (Anonymous, 2002). The UK CKD guidelines also recommend at least annual screening of all adults at risk of obstructive kidney disease and those with prevalent CVD, while the US KDOQI guidelines extend screening to all those aged > 60  years with testing for albuminuria (using urine albumin/ protein), SCr, and estimation of GFR (Anonymous, 2002, 2007). Recommendations from the ISN are more liberal, and advocate proactive screening for markers of CKD in all subjects visiting general practitioners, similar to the screening for high blood pressure or cholesterol concentrations (Levey et  al., 2011). However, all these recommendations are based mostly on expert consensus rather than high quality evidence, and doubt remains as to whether population or targeted screening is justifiable and cost-effective (Boulware et al., 2003; Manns et al., 2010). In cardiovascular disease (CVD), it has been argued that population screening would be more effective than targeted approaches as most cases are not derived from the minority at highest risk (Levin and Stevens, 2011). However, CVD is more prevalent than CKD, and it is uncertain whether the same arguments apply. Before population-based screening for CKD could be recommended, further research is required to determine the true prevalence of early stages of CKD in different populations; the prevalence of associated risk factors; the attributable risk associated with such risk factors; their amenability to treatment; and the incremental benefit compared to case-finding by means other than organized screening.

Prognosis and outcome in CKD Overall, even a mild reduction in estimated eGFR is associated with adverse clinical outcomes, as is increased urinary protein excretion (reviewed in (Matsushita et al., 2010b; Gansevoort et al., 2011; van der Velde et al., 2011). Among subjects with normal kidney function, proteinuria is independently associated with an increased risk of poor outcomes, which is further amplified in the setting of reduced eGFR—and has been observed in multiple populations including those at high CVD risk (Gerstein et al., 2001; Mann et al., 2001; Rahman et al., 2006; Solomon et al., 2007; Yokoyama et al., 2008; Anand et al., 2009) and in the general population (Grimm et al., 1997; Clausen et al., 2001; Kasiske, 2001; Hillege et al., 2002; Go et al., 2004; Klausen et al., 2004; Tonelli et al., 2006; Hemmelgarn et al., 2010; Matsushita et al., 2010b).

Epidemiology of renal replacement therapy Definition of RRT (See Sections 12 and 13.) RRT comprises the different forms of haemodialysis, peritoneal dialysis, and kidney transplantation and may be used to overcome a short period of kidney failure. The majority of RRT patients, however, receive chronic RRT for the treatment of ESRD (Levey et al., 2011). As mentioned earlier, not all patients with ESRD receive RRT—for example, due to differences in access to RRT across healthcare systems, or because of cultural differences in referral

13

Section 1  

assessment of the patient with renal disease Rates

Rates 2000 Rate per million population

Rate per million population

2000 1500 1000 500 0

0–19 20–44 45–64 65–74 75+ All

1500 1000 500 0

80 84 88 92 96 00 04 08

94 96 98 00 02 04 06 08

Fig. 1.4  Incidence of RRT by age category in USRDS (left) and ERA-EDTA registry (right) 1998–2012 (ERA-EDTA Registry, 2014; US Renal Data System, 2014).

patterns and patient attitudes towards RRT. Therefore, estimates based on the incidence and prevalence of chronic RRT will underestimate the burden of ESRD.

Risk factors and causes of ESRD Risk factors for ESRD are similar to those for CKD—with the caveat that characteristics that reduce the risk of mortality may also increase the risk of ESRD (since only those who survive long enough can receive RRT). Worldwide, diabetes mellitus and hypertension are the leading causes of ESRD treated with RRT. Renal registries use different systems for coding primary renal diseases underlying ESRD. The US Renal Data System (USRDS) makes use of ICD-9-CM coding, other registries use ICD-10, but the majority make use of (modifications of) the European Renal Association–European Dialysis and Transplant Association (ERA-EDTA) coding system for primary renal diseases (van Dijk et al., 2001; Venkat-Raman et al., 2012).

Global epidemiology of RRT for ESRD The incidence of RRT as reported by registries is determined by the age and sex distribution of the general population, the prevalence of diseases underlying ESRD, factors related to progression of CKD, survival from competing risk like CVD, access to RRT (Caskey et al., 2011), the timing of starting RRT in relation to GFR, and the completeness of data within those registries. The prevalence of RRT results from the incidence of RRT and the survival of RRT patients. Registry data show substantial changes in the incidence rates of RRT over the last 15 years. Figs 1.4 and 1.5 provide examples of trends in the United States and in Europe by age and primary renal disease. Rates 160 120 80 40 0

80 84 88 92 96 00 04 08

The initial rapid growth in incidence rates was likely due to a combination of population ageing, increased prevalence of diseases underlying ESRD, and an increased acceptance of older and sicker patients. In addition, a substantial rise of the number of patients starting dialysis above 10 mL/min/1.73m2 has provided a significant contribution to this increase (Rosansky et al., 2009). Recent data indicate that in the United States and in Europe the incidence rate of RRT has decreased since 2009 (US Renal Data System, 2014; ). Current differences in the incidence of RRT across the world are striking (Table 1.8). Mexico, Taiwan, and the United States have the highest incidence rates, whereas Hungary, Portugal, and Greece rank first in Europe. The prevalence of RRT has continued to increase by 1–2% per year (Kramer et al., 2009; US Renal Data System, 2014). Although in some countries this may in part be due to increased patient survival on RRT, the increase in prevalence may merely reflect higher numbers of patients starting RRT than numbers of patient deaths. Countries with the highest burden of RRT patients include Taiwan, Japan, and the United States. In general, patients on RRT have an unfavourable prognosis with remaining life expectancies being reduced to 20–35% in dialysis patients and to 70–80% in transplant recipients (ERA-EDTA Registry, 2014; US Renal Data System, 2014). In dialysis patients both cardiovascular (Foley et al., 1998; de Jager et al., 2009) and non-cardiovascular (de Jager et  al., 2009; Sarnak and Jaber, 2000) mortality are highly increased compared with those in the general population. Determinants of patient survival on RRT include age and sex, the cause of ESRD, timing of referral to a nephrologist, genetic factors, general population mortality, access to Rates

Rate per million population

Rate per million population

14

160 120

Diabetes Hypertension Glomerulonephritis Cystic disease

80 40 0 94 96 98 00 02 04 06 08

Fig. 1.5  Incidence of RRT by primary renal disease in USRDS (left) and ERA-EDTA registry (right) 1998–2012 (ERA-EDTA Registry, 2014; US Renal Data System, 2014).

Table 1.8  Incidence and prevalence estimates of RRT in different countries in 2012 Country

Incidence rate of RRT (pmp)

Incidence of DM (%)

Incidence of DM (pmp)

Prevalence of RRT (pmp)

Prevalence of RRT Prevalence of HD as (pmp) on transplant % of dialysis )

Albania

67

10

7

325

73

Argentina

159

36

57

836

168

95

Australia

112

36

40

916

411

81

Austria

140

26

36

1023

516

91

Bahrain

258

36

93

328

51

88

Belgium

188

20

37

1225

514

92

Bosnia and

125

29

36

718

52

96

720

216

91

38

59

1183

500

83

1263

203

95

Brazil

172

Canada

156

Chile

170

Croatia

158

24

38

1033

384

93

Denmark

124

28

35

872

411

80

Estonia

81

22

17

554

328

86

Finland

81

34

27

808

482

82

France

154

22

33

1139

507

93

Georgia

200

24

47

546

49

97

Greece

210

26

54

1136

231

93

484

Hong Kong

165

48

79

1192

Hungary

234

39

91

633

27

Iceland

59

0

0

683

Indonesia

191

26

50

265

Iran

105

22

23

621

299

95

395

94

86 440

73 96

Israel

183

49

89

1125

Japan

285

45

128

2365

Latvia

89

20

18

538

290

74

Malaysia

225

61

137

1056

64

91

Mexico (Jalisco)

467

59

276

1409

526

50

Montenegro

24

27

7

310

135

94

Netherlands

120

16

20

923

539

85

New Zealand

116

49

57

901

344

69

Norway

103

17

17

887

639

84

695

331

92

97

Oman

110

48

53

Philippines

117

44

51

Poland

135

25

34

748

254

94

Portugal

220

31

69

1670

602

93

Qatar

83

280

2

77

Rep. of Korea

221

51

113

1353

272

87

Romania

151

13

20

766

58

89

Russia

48

17

8

214

44

92

95

(continued)

16

Section 1  

assessment of the patient with renal disease

Table 1.8 Continued Country

Incidence rate of RRT (pmp)

Incidence of DM (%)

Incidence of DM (pmp)

Prevalence of RRT (pmp)

Prevalence of RRT Prevalence of HD as (pmp) on transplant % of dialysis )

Saudi Arabia

126

39

49

730

245

91

Serbia

123

24

30

752

106

91

Singapore

285

66

188

1741

368

88

Slovenia

122

28

34

996

310

97

Spain

120

25

30

1092

555

89

Sweden

115

23

26

933

530

79

Taiwan

450

45

203

2902

Thailand

221

38

84

906

89

90 77

Ukraine

25

12

3

147

18

85

United States

359

44

159

1976

594

91

United Kingdom

107

24

25

867

435

86

Uruguay

150

33

40

1073

316

90

Sources: ERA-EDTA 2014 and USRDS 2014; where cells are empty, data are unavailable.

transplantation, and other aspects of quality of RRT care (Kramer et al., 2012). There exists considerable international variation in the survival of patients starting dialysis (Kramer et al., 2012). Differences in patient age, sex, primary renal disease, and the presence of co-morbidities explain only a small part of that international variation (Goodkin et al., 2003; van Manen et al., 2007). Even when general population mortality rates and differences in treatment characteristics are taken into account, a major part of the variation in dialysis mortality across countries remains unexplained (van Dijk et al., 2007). Recent data suggest that the mortality of dialysis patients is higher in countries with high expenditure on healthcare. Complementary explanations for the international variation in mortality on dialysis may therefore include a more liberal acceptance policy among richer nations, differences in access to transplantation as well as different patterns of healthcare spending (Kramer et al., 2012).

RRT in special populations (See Chapter 95.) The epidemiology of RRT differs in specific populations like ethnic minorities and children. Data from the USRDS (2014) show that in 2012 the age- and sex-adjusted incidence rates of RRT for African Americans, Native Americans, and Asians were 908, 412, and 379 pmp. This is 3.3, 1.5, and 1,4 times greater than the rate of 279 found in white people. Incidence rates from Europe, albeit lower, have shown an increased incidence rate of RRT in Asians and black people (Roderick et al., 1996)  and in non-Caucasians in general (van den Beukel et  al., 2010), whereas in Canada, Australia, and New-Zealand incidence rates are highly increased in indigenous people (Dyck, 2001; McDonald, 2010; McDonald et al., 2010). Increased incidence rates in ethnic minorities frequently come together with better patient survival rates. US and UK black patients (Roderick et al., 2009; US Renal Data System, 2014), UK South Asian (Roderick et al., 2009), and non-Caucasian Dutch RRT patients (van den Beukel et  al.,

2008) have better survival rates than their white and non-Caucasian counterparts. In contrast, in Aboriginal Australians and New Zealand Maoris patient survival is lower compared with that of non-indigenous people (McDonald and Russ, 2003). The background of these survival differences requires further elucidation. In 2008, the median incidence rate of RRT in children aged 0–19 years was 9 (range 4–18) per million age-related population (pmarp) with the highest incidence rates in adolescents (Harambat et al., 2012). In children the pattern of diseases underlying ESRD is quite different from that in adults. In developed countries congenital disorders, including congenital anomalies of the kidney and urinary tract (CAKUT) and hereditary nephropathies, are responsible for more than half of all cases starting RRT. Differences in the incidence and causes of RRT across races and ethnic minorities are already reflected in paediatric populations (Harambat et  al., 2012). The prevalence of RRT was around 65 pmarp in Australia, Canada, Malaysia, and Western-Europe, whereas the United States had a higher (85 pmarp) and Japan a lower (34 pmarp) prevalence. Although mortality of children on RRT is relatively low compared with that in adults (2-year survival 96% in Europe (ERA-EDTA Registry, 2014) and 95% in the United States (US Renal Data System, 2014), their risk of death is about 30 times higher than in their healthy peers (Groothoff et al., 2002; McDonald and Craig, 2004).

Future challenges A key future challenge is to improve the definition and characterization of various kidney disease entities (Gansevoort and de Jong, 2010). This is an area of intense interest: better methods for SCr measurement, calibration, and standardization have evolved over the years, and higher-performance estimating equations are being developed (Stevens et al., 2010). In addition, newer filtration markers that are independent of muscle mass (such as cystatin C) are gaining increasing recognition and are being tested as a future alternative to SCr. Other potential biomarkers such as neutrophil gelatinase-associated lipocalin, kidney injury molecule-1, monocyte

chapter 1 

chemotactic peptide, netrin-1, and interleukin-18 among several others are becoming available for early identification of AKI (Waikar et al., 2008; Soni et al., 2011). It seems likely that some of these new tools will become more widely available, and will help to address the limitations of existing epidemiological studies. A second key challenge is to develop epidemiologic surveillance systems for both AKI and the various aspects of CKD—especially non-dialysis-dependent disease (Levey et al., 2011), and in developing nations (Barsoum, 2006; Nugent et al., 2011). Third, better information is needed on who to screen for kidney disease (as well as what test should be used and when screening should be performed)—aiming to identify people in whom early intervention to prevent adverse outcomes is feasible and cost-effective. These initiatives will help to establish the prevention and treatment of CKD as an important public health issue, and potentially as part of the WHO strategy on chronic non-communicable diseases (Couser and Riella, 2011).

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assessment of the patient with renal disease

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Keith, D. S., Nichols, G. A., Gullion, C. M., et al. (2004). Longitudinal follow-up and outcomes among a population with chronic kidney disease in a large managed care organization. Arch Intern Med, 164, 659–63. Klausen, K., Borch-Johnsen, K., Feldt-Rasmussen, B., et al. (2004). Very low levels of microalbuminuria are associated with increased risk of coronary heart disease and death independently of renal function, hypertension, and diabetes. Circulation, 110, 32–5. Kramer, A., Stel, V. S., Caskey, F. J., et al. (2012). Exploring the association between macroeconomic indicators and dialysis mortality. Clin J Am Soc Nephrol, 7(10), 1655–63. Kramer, A., Stel, V., Zoccali, C., et al. (2009). An update on renal replacement therapy in Europe: ERA-EDTA Registry data from 1997 to 2006. Nephrol Dial Transplant, 24, 3557–66. Lameire, N., Van Biesen, W., and Vanholder, R. (2005). Acute renal failure. Lancet, 365, 417–30. Lameire, N., Van Biesen, W., and Vanholder, R. (2006). The changing epidemiology of acute renal failure. Nat Clin Pract Nephrol, 2, 364–77. Lameire, N. H., Bagga, A., Cruz, D., et al. (2013). Acute kidney injury: an increasing global concern. Lancet, 382,170–9. Levey, A. S. and Coresh, J. (2011). Chronic kidney disease. Lancet, 379, 165–80. Levey, A. S., De Jong, P. E., Coresh, J., et al. (2011). The definition, classification, and prognosis of chronic kidney disease: a KDIGO Controversies Conference report. Kidney Int, 80, 17–28. Levin, A. and Stevens, P. E. (2011). Early detection of CKD: the benefits, limitations and effects on prognosis. Nat Rev Nephrol, 7, 446–57. Liano, F., and Pascual, J. (1996). Epidemiology of acute renal failure: a prospective, multicenter, community-based study. Madrid Acute Renal Failure Study Group. Kidney Int, 50, 811–18. Liu, Y., Coresh, J., Eustace, J. A., et al. (2004). Association between cholesterol level and mortality in dialysis patients: role of inflammation and malnutrition. JAMA, 291, 451–9. Mann, J. F., Gerstein, H. C., Pogue, J., et al. (2001). Renal insufficiency as a predictor of cardiovascular outcomes and the impact of ramipril: the HOPE randomized trial. Ann Intern Med, 134, 629–36. Manns, B., Hemmelgarn, B., Tonelli, M., et al. (2010). Population based screening for chronic kidney disease: cost effectiveness study. BMJ, 341, c5869. Matsushita, K., Selvin, E., Bash, L. D., et al. (2010a). Risk implications of the new CKD Epidemiology Collaboration (CKD-EPI) equation compared with the MDRD Study equation for estimated GFR: the Atherosclerosis Risk in Communities (ARIC) Study. Am J Kidney Dis, 55, 648–59. Matsushita, K., van der Velde, M., Astor, B. C., et al. (2010b). Association of estimated glomerular filtration rate and albuminuria with all-cause and cardiovascular mortality in general population cohorts: a collaborative meta-analysis. Lancet, 375, 2073–81. McDonald, S. (2010). Incidence and treatment of ESRD among indigenous peoples of Australasia. Clin Nephrol, 74 Suppl 1, S28–31. McDonald, S., Excell, L., and Jose, M. (2010). End-stage kidney disease among Indigenous peoples of Australia and New Zealand. In ANZDATA Registry Report 2010, pp. 12-1–12-19. Adelaide: Australia and New Zealand Dialysis and Transplant Registry. McDonald, S. P. and Craig, J. C. (2004). Long-term survival of children with end-stage renal disease. N Engl J Med, 350, 2654–62. McDonald, S. P. and Russ, G. R. (2003). Current incidence, treatment patterns and outcome of end-stage renal disease among indigenous groups in Australia and New Zealand. Nephrology (Carlton), 8, 42–8. Mehta, R. L. and Chertow, G. M. (2003). Acute renal failure definitions and classification: time for change? J Am Soc Nephrol, 14, 2178–87. Mehta, R. L., Kellum, J. A., Shah, S. V., et al. (2007). Acute Kidney Injury Network: report of an initiative to improve outcomes in acute kidney injury. Crit Care, 11, R31. Moher, D., Liberati, A., Tetzlaff, J., et al. (2009). Preferred reporting items for systematic reviews and meta-analyses: the PRISMA statement. PLoS Med, 6, e1000097. Noordzij, M., Hooft, L., Dekker, F. W., et al. (2009). Systematic reviews and meta-analyses: when they are useful and when to be careful. Kidney Int, 76, 1130–6.

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CHAPTER 2

Clinical assessment of the patient with renal disease: overview Christopher G. Winearls Introduction The next chapters describe the presentations of renal diseases, their assessment from the history and physical examination, and their investigation by the use of laboratory tests, imaging, and histopathology. Nephrologists are referred patients by their colleagues in primary care or in other specialties when they believe that there is a kidney problem needing an expert. That judgement is not always correct, for example, oedema or haematuria often have causes outside the kidney. The reasons for referral are in three categories: 1. To explain abnormalities attributable to kidney disease that have been found in asymptomatic individuals including those at risk of familial conditions. 2. Symptomatic renal disease: renal failure, either acute or chronic; abnormalities of urination including poly- and oliguria, visible haematuria; unexplained loin pain; the classic renal syndromes (nephritic and nephrotic). 3. Renal consequences of systemic conditions: metabolic, inflammatory, infectious; drugs, malignancy, pregnancy, organ failure (especially cardiac and hepatic). When responding to referrals coming by letter, telephone, or email there are two questions to be asked: 1. Should one be involved in this patent’s care and if so how? In other words, will one be able to add value? The problems seldom come neatly packaged. They have often emerged into prominence in the context of complex other diseases, both acute and chronic. Sometimes the referral is for diagnosis, sometimes it is management, and sometimes it is both. These dilemmas are particularly poignant for nephrologists who have at their disposal two powerful tools. They can examine the diseased organ directly by examining tissue obtained by biopsy and can, theoretically, replace its function indefinitely. The question should sometimes be not, ‘Can I?’ but, ‘Should I be involved?’ It takes some courage to admit that one has nothing to offer, for example, the patient with oliguria and terminal heart failure or malignant disease. It is hubristic to suggest interventions directed at a consequence of irremediable disease which may prolong suffering for no more than a few days and delay a merciful death and so one should pause before doing so. The supplementary question is; ‘If I should be involved, should I be the clinician in overall charge?’ When the renal disease dominates the clinical problem, for example,

in acute dialysis dependent renal failure, there are advantages in one team determining the priorities and supervising drug prescription. There are others in which a supportive role is sufficient. To take charge of every patient with a renal component to their illness would overwhelm the service. This is a particular problem in patients with renal failure as an additional complication of complicated surgery. Such patients need to be managed in the service responsible for treating the root cause. Without so doing, the renal problem will not resolve either. 2. How soon does one need to be involved? Contact may need to be immediate, for example, when there is an acute uraemic emergency or delayed as a routine outpatient assessment. This ‘triaging’ is not always straightforward. Some patients with life- and kidney-threatening conditions can appear deceptively well. Two examples come to mind—acute cast nephropathy in myeloma and rapidly progressive glomerulonephritis. One should not be deceived by the automatic labelling of a patient with an abnormal creatinine that has been translated into an estimated glomerular filtration rate (eGFR), as having chronic kidney disease usually abbreviated in correspondence to ‘CKD’. Having accepted the referral the nephrologist should clear his/ her mind of the potentially misleading glib statements within referral letters and clinical notes, assumptions of the diagnosis, and the prejudices of the referrer or indeed the patient. Who has not seen the oedematous ‘nephrotic’ patient without proteinuria who actually has heart failure or the patient with a reduced eGFR labelled as having ‘CKD’ explained by anything from bodybuilding to hypothyroidism? A  good rule is to assume that the referral lacks several pieces of highly relevant information and includes some that are actually wrong—‘no nephrotoxic drugs’, ‘normal-sized kidneys’, and ‘no relevant family history’ are common false assertions. It is then time to apply the ‘clinical method’ which comprises the following steps: ◆ Identification of the presenting complaint and its immediate history. The history should be methodically retaken including the general history of the patient including past medical, drug, social, travel, illness in the family, and a systematic enquiry of the function of other systems. ◆ Performing a general physical examination. ◆ Arranging laboratory and imaging investigations—screening and targeted.

chapter 2  ◆ Constructing

a differential diagnosis. ◆ Formulating a plan for management: specific and general. ◆ Communicating the conclusions to the patient, referring colleagues and others who will be needed to contribute to care. There is no substitute for ‘the clinical method’. It is a false economy to move straight to investigations not only because these may not provide the answer but because their interpretation will depend

clinical assessment of patients with renal disease on the clinical context. Radiologists and renal pathologists rightly expect nephrologists to describe the problem and how the findings will alter management. Accurate diagnosis is paramount for it informs the medical management exactly but knowing everything else will dictate how this will be delivered. The way the clinical method is applied depends on the clinical presentation—these are described in Chapter 3.

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CHAPTER 3

Presentations of renal disease Christopher G. Winearls Introduction It is not patients who seek the advice and help of nephrologists but primary care physicians and medical and surgical colleagues, who have identified a problem that they believe calls for the expertise of a nephrologist. What are these circumstances?

Abnormalities in asymptomatic patients raising the suspicion of renal disease The referral usually follows the finding of an abnormality on clinical or laboratory examination that is not causing symptoms. These arise at routine medical examinations at school, for employment, military service, life and salary insurance, immigration, major surgery, potential live kidney donation, registration with a new family doctor, ‘well woman/man’ clinics and as part of health screening in at-risk individuals with systemic disease, at booking for pregnancy care, and assessment of the risks of methods of contraception. The potential for causing anxiety is significant, especially if the patient attends a renal unit for evaluation, walking past signs to the dialysis and transplant wards. This concern has to be balanced against the wish to make an early diagnosis of renal disease in case it is possible to halt it, delay its progress, and defer or prevent the onset of renal failure. We will consider first the common reasons patients are referred to a nephrologist for the first time.

Abnormal findings in patients at risk of renal disease and complications Many referrals will come from colleagues in other disciplines who are on the lookout for renal problems associated with the condition they are treating or monitoring. Indeed they will have been advised to measure the estimated glomerular filtration rate (eGFR) more or less frequently. ◆ Primary care physicians are encouraged to screen for renal dysfunction in patients with diabetes, hypertension, heart failure, and vascular disease (ischaemic heart disease, strokes, and peripheral vascular insufficiency) and those on long-term treatment with drugs known to cause kidney injury, for example, lithium, non-steroidal anti-inflammatory drug, and calcineurin inhibitors. ◆ Urologists will refer patients with neurogenic bladders, urinary diversion, kidney stones, retroperitoneal fibrosis, and even relieved obstruction. ◆ Gastroenterologists will refer patients with malabsorption who are prone to oxalate nephropathy, and those on 5-aminosalicylic acid drugs used for inflammatory bowel disease which can cause interstitial nephritis at any time after they are instituted.

◆ Cardiologists

and vascular surgeons will refer patients with atherosclerotic disease and abdominal aortic aneurysms may also suffer from renal vascular disease which comes to light when renin–angiotensin–aldosterone system (RAAS) blockade is instituted. ◆ Haematologists will refer patients with paraprotein disorders. ◆ Rheumatologists will refer patients with vasculitides in whom they are suspicious of relapse in the hope that a renal biopsy will confirm this and justify escalation of immunosuppression. ◆ Radiologists will often add as addendum to their reports of incidental findings of renal cysts, kidney size asymmetry, and parenchymal calcification, ‘suggest referral to a nephrologist’.

Invisible haematuria (microscopic haematuria) (See Chapter 46.) This is by definition asymptomatic and is found when urine is tested with urine dipsticks as part of clinical assessment. This problem is not referred to nephrologists in the first instance.

Asymptomatic proteinuria (See Chapter 50.) This may have been detected by dipstick testing either as a part of a routine assessment or under abnormal conditions, for example, an intercurrent illness. Dipsticks can be misleading because they are not calibrated for urinary concentration. An early morning spot urine protein or albumin creatinine concentration should be requested and the presence of orthostatic proteinuria excluded before the patient is formally referred. The threshold for full evaluation of significant proteinuria (defined as > 300 mg/day or 30 mg/ mmol creatinine) up to and including renal biopsy will depend on the clinical context and the ‘need to know’.

Abnormal constituents of urine (See Chapter 175.) These include asymptomatic bacteriuria, and leucocyturia (see Chapter 175). Asymptomatic bacteriuria without leucocyturia suggests sample contamination, with leucocytes, an asymptomatic infection or urinary tract colonization. Isolated leucocyturia is found after treatment for an infection, in subjects with renal stones, interstitial nephritis, renal tuberculosis, or papillary necrosis.

An ‘abnormal’ eGFR (See Chapter 94.) This has become one of the most common reasons for referral to a nephrologist. This has arisen from the application of the Modification of Diet in Renal Disease and Chronic Kidney Disease Epidemiology

chapter 3 

presentations of renal disease

Collaboration eGFR equations to routine plasma creatinine measurements which are then reported as an eGFR and an interpretation as to a possible stage of chronic kidney disease (CKD), for example, ‘an eGFR of 45–59 mL/min/1.73m2 indicates stage 3A kidney disease’. Such statements are neither useful nor legitimate not least because a diagnosis of CKD requires the abnormal GFR to have been present for at least 3 months but also because the explanation may be acute kidney injury (AKI). It is also misleading because an eGFR of > 60 mL/min/1.73m2 is not necessarily ‘normal’. Indeed, there is no definition of a normal eGFR. Although an eGFR of < 60 mL/min is likely to indicate impairment of kidney function across the age range, the significance is different in the young compared to the old. A decision as to whether individuals with a reduced eGFR need a formal nephrological assessment will depend on their underlying co-morbid conditions, age, and the rate of change. The new KDIGO CKD guideline has the explicit requirement that CKD should be described according to cause, GFR, and albumin excretion—the CGA system (see Chapter 94). When the GFR is unequivocally abnormal (< 45mL/ min/1.73m2) or lower than predicted for the age of the patient there will need to be a decision on the urgency of the assessment. These dilemmas are explored and recommendations made in the KDIGO clinical practice guideline (Kidney Disease:  Improving Global Outcomes (KDIGO) Acute Kidney Injury Work Group, 2012).

consequence of RAAS blockade with angiotensin converting enzyme inhibitors (ACEIs) or angiotensin receptor blockers (ARBs), or potassium-sparing diuretics such as spironolactone or amiloride. This is becoming a common problem in patients receiving such combinations as part of their treatment for heart failure. When renal function is relatively preserved, these abnormal K+ concentrations, usually approximately 6.5 mmol/L do not cause problems.

A raised blood pressure

Urinary symptoms

Patients are referred if there are indications that this may be secondary to renal disease (reduced eGFR, haematuria, and or proteinuria). They will require a full nephrological assessment. (See Chapter 210.)

Screening for an inherited disorder known to be present in the family Common examples include autosomal dominant polycystic kidney disease (ADPKD), Alport syndrome, reflux nephropathy, urate nephropathy, and nephronophthisis-medullary cystic disease complex. (See Sections 15, 16.)

Hyper- and hyponatraemia (See Chapters 28 and 29.) Hypernatraemia is seldom asymptomatic but hyponatraemia (sodium < 135 mmol/L) often is. This is usually a consequence of salt wasting with mainly water replenishment, psychogenic polydipsia, and the syndrome of inappropriate antidiuretic hormone secretion (SIADH) a full description of this problem is provided in Chapter 28.

Symptomatic urinary disease The patient has symptoms and signs that strongly suggest an underlying kidney or urinary tract disease. Patients in this group of referrals may be suffering urinary symptoms or from one of the recognized renal syndromes that trigger automatic referral to a nephrologist. These are listed in Table 3.1. Patients with problems of urination (micturition) are not referred to nephrologists in the first instance but these may be relevant in cases referred with haematuria, recurrent urinary tract infection. Such patients will be referred either in the hospital setting or as requests for advice from the primary care. There is often overlap with urology and choice and direction of the referral is often perverse and random. Table 3.1  Urinary syndromes and symptoms

Incidental electrolyte abnormalities

Syndrome

Definition

(See Section 2.)

Nephrotic

Oedema associated with hypoalbuminaemia and heavy proteinuria (usually > 3 g/day)

Nephritic

Hypertension and oedema associated with haematuria and proteinuria

Acute kidney injury

This is based on urine output and change in plasma creatinine concentration (Ftouh et al., 2013)

Chronic kidney disease

Abnormalities of kidney structure or function present for > 3 months with implications for health.

Chronic kidney failure

An eGFR < 15 mL/min causing uraemia requiring symptom control, dialysis, and or renal transplantation

Hypokalaemia (K+ < 3.5 mmol /L) (See Chapter 34.) This will often cause concern. When associated with hypertension, a number of disorders will have to be considered. These include renovascular hypertension and Cushing, Conn, and rarely Liddle syndromes. If the blood pressure is normal the simple explanations should be considered first, for example, thiazide and loop diuretic use, diarrhoea, and purgative use. The abuse of such drugs can be hard to prove. A rare explanation is spurious hypokalaemia caused by delay in potassium (K+) measurement in patients with a membrane pump abnormality. This is also seen in patients with acute myeloblastic leukaemia. All these may have been excluded before the nephrologist is asked to confirm or deny the presence of Gitelman or Bartter syndromes.

Hyperkalaemia (K+ > 5.5 mmol/L) (See Chapter 34.) This also causes concern and the risk-averse doctor will refer patients to hospital urgently for further blood tests. The explanations range from the trivial, a haemolysed or old blood sample especially if taken with a rare tight tourniquet, to the relevant, for example, a

Symptoms: Urinary symptoms Recurrent macroscopic haematuria Loin pain ± haematuria Hypertensive renal disease Dysuria, polyuria

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Dysuria This is an umbrella term covering different sensations of discomfort with micturition and experienced, therefore, several times a day. In men, burning in the penile urethra suggests urethritis but a deeper pain suggests problems in the prostate or bladder. If there are systemic symptoms and signs, the prostate should be considered and samples taken after prostatic massage and imaging organized. Acute prostatitis can progress to septicaemia and the treatment is different from cystitis. Cystitis is uncommon in men unless there is an underlying structural or predisposing cause such as a stone, outflow obstruction, or malignancy. In women, dysuria is often associated with urinary urgency and frequency suggesting a diagnosis of cystitis. This occurs more commonly in women during their sexually active years but also after menopause when the effects of oestrogen deficiency reduce the defences of the bladder. Dysuria also describes difficulty in actually passing urine. This is more common in men who will describe a collection of distressing symptoms pointing to bladder outflow obstruction. Because bladder emptying is incomplete, they notice frequency, including nocturia, urgency, difficulty in initiating micturition (waiting up to a minute for flow to start), a poor stream, and then dribbling after micturition is thought to be finished. The finding of a full bladder often palpable to the umbilicus, a large volume of post-micturition residual urine, and a thick-walled bladder on bladder scan give the diagnosis. The causes include urethral structure and prostatic occlusion of the urethra. This diagnosis is often made when these men, often elderly, are referred unexamined with ‘CKD’. Strangury is the symptom of very painful and difficult micturition often caused by a bladder stone at the internal urethral meatus or in the urethra itself.

Urinary frequency, polyuria, and nocturia It is essential to establish what the patient actually means when they describe ‘Having to go to the toilet all the time’. ‘How often and how much urine?’ are the questions that need answering. It is best to get a record of a day’s worth of micturition, charting time and volumes passed. A kitchen measuring jug will suffice. Frequent large volumes of urine point to a concentration defect and frequent small volumes to a micturition problem or bladder irritation or a contracted bladder volume, setting off detrusor contraction despite the presence of relatively small volumes. The causes of true polyuria > 3 L/day are many (see Table 3.2). The diuresis may be the result of excess water consumption

Table 3.2  Causes of polyuria Cause

Diagnosis

Primary polydipsia

History and/or water deprivation

Solute diuresis

Urinary glucose

Drugs

Diuretic use

Central diabetic insipidus

Water deprivation test

Nephrogenic diabetes insipidusa

History, kidney imaging, and a water deprivation test

a Common causes: post-obstructive, hypercalcaemia, long-term lithium use or excess dose

acutely, and many forms of interstitial kidney injury. (See Chapter 32.)

(polydipsia), obligate loss caused by solutes such as glucose, or failure of ‘antidiuresis’ at a central (posterior pituitary) or peripheral (the kidney) level. These are simply distinguished by direct questioning and occasionally a water deprivation test with and without administration of arginine vasopressin. Primary or congenital diabetic insipidus is rare and will manifest in early life. Nocturia is the disturbing need to wake and micturate during the sleep cycle. Provided the bladder has been emptied before retiring and fluids have been avoided in the preceding 1–2 hours, most healthy subjects can last until morning perhaps because sleep is associated with antidiuretic hormone (ADH) secretion. When the volumes of urine passed are low, the causes are those of frequency described earlier. Large volumes may be the continuation of the problem of polyuria but a reversed problem of micturition, that is, more at night than in the day is more problematic. It implies that there is more urine production at night than during the day and is attributed to preferential renal perfusion when the other calls on cardiac output are reduced. It is commonly seen in patients with CKD, congestive heart failure, and nephrotic syndrome.

Anuria and oliguria (See Chapter 219.) It is uncommon for patient to report these and curious that they do not notice that they have ‘stopped going’. Anuria and oliguria (urine output < 400 mL/24 hours or < 0.5 mL/kg per hour) are emergencies and in the otherwise symptom free patient point to relatively few causes. Anuria The differential diagnosis includes acute renal vascular occlusion, catastrophic renal injury, for example, antiglomerular basement membrane (anti-GBM) disease, and bilateral complete urinary obstruction or obstruction of a single kidney. Oliguria There are many causes, including those of anuria, but there is usually evidence of a systemic disorder with or without a further aggravant or precipitant, for example, dehydration in a patient with heart failure and an ACEI and diuretics. Both oliguria and anuria are observed in the hospital setting and will trigger an entry into an AKI algorithm examining prerenal, renal, and postrenal causes (Lameire et al., 2013).

Macroscopic haematuria This is an alarming symptom and because of association of ‘red’ with danger, is seldom ignored. In the United Kingdom, any adult patient with painless macroscopic haematuria would be referred to a urologist to be seen within 2 weeks to exclude malignancy. Referral to a nephrologist is usually after the common urological causes have been excluded. The obvious ‘medical’ renal causes include ADPKD and over-anticoagulation but three glomerular diseases can manifest as macroscopic haematuria: Immunoglobulin A (IgA) nephropathy, Alport syndrome, and anti-GBM disease. Patients with the nephritic syndrome describe brown cloudy urine which is less alarming than truly bloody urine.

Loin pain This problem is usually referred when a urological cause has already been sought but it is worth rehearsing the symptoms and causes associated with loin pain.

chapter 3 

Renal colic is a very severe pain during which no normal activity can be undertaken. It is sudden in onset, comes in waves, radiates anteriorly and into the genitalia, and is associated with nausea and vomiting. It can abate quite suddenly. The urine will often but not always test positive for blood. This description implies that a stone or clot or papilla is in the ureter which is trying to move it on by peristalsis. These episodes are usually managed in the community and are only referred if the problem is recurrent or there are ‘red flags’. These include fever (implying the possibility of infection behind an obstructing stone), known solitary kidney, pain resistant to standard analgesia, pregnancy, renal dysfunction, oliguria, or poor social support. It is more difficult to attribute pain confined to the loin to the presence of a stone. There are other causes such as bleeds into renal cysts, pyelonephritis, renal infarcts, pelvi-ureteric junction obstruction, and the loin pain haematuria syndrome. This is a curious condition in which patients present with very severe chronic loin pain with and without visible haematuria and few if any abnormalities are found by imaging or even renal biopsy; the description of the pain is vivid and by the time of referral many patients are taking very large doses of opiate analgesics. Occasionally a factitious cause of haematuria is proved. On examination the patients are exquisitely tender during attempts to palpate the kidney bi-manually. There will often be a request for a surgical solution ranging from auto transplantation to nephrectomy. It is considered a form of somatoform pain disorder (Winearls and Bass, 1994) (See Chapter 47). Acute glomerulonephritis is occasionally associated with loin discomfort but seldom with severe pain. Severe loin pain has also been attributed to the ‘nutcracker phenomenon’ when the left renal vein is compressed between the aorta and the superior mesenteric artery.

Nephrotic syndrome (See Chapters 48, 52.) This is a shorthand term for the combination of oedema, heavy proteinuria, and hypoalbuminaemia. Cut-off concentrations as diagnostic criteria are unhelpful as there is a poor correlation with the effects of the syndrome. Hypercholesterolaemia is an epiphenomenon and not helpful diagnostically. Usually the plasma albumin is < 30 g/L, the urine protein loss > 3 g/24 hours or > 350 mg/mmol creatinine. The diagnosis may be missed if the latter two components are not sought and the oedema misattributed to immobility, heart failure, and venous insufficiency. Adult patients need prompt assessment by a nephrologist and almost all will require a renal biopsy. The cost of guessing the pathology by the known hierarchy of causes in age groups is too high to be allowed.

presentations of renal disease

used today as most patients with this combination have recognized glomerular diseases such as IgA nephropathy, Henoch–Schönlein purpura (HSP), systemic lupus erythematosus (SLE) with a diffuse proliferative glomerulonephritis, or a systemic vasculitis.

Acute kidney injury (See Section 11.) AKI is defined as any of the following:  an increase in serum creatinine by > 0.3 mg/dL (> 26.5 μmol/L) within 48 hours; or an increase in serum creatinine to > 1.5 times baseline, which is known or presumed to have occurred within the prior 7 days; or a urine volume < 0.5 mL/kg/h for 6 hours (Kidney Disease:  Improving Global Outcomes (KDIGO) Acute Kidney Injury Work Group, 2012). In practice, AKI covers a spectrum from transient minor to catastrophic kidney injury arising in both hospital and the community and always represents a nephrological emergency. There are two tasks: one is to attempt to halt or reverse the injury and the other is to provide support to the patient to compensate for the effects of renal failure (Kidney Disease:  Improving Global Outcomes (KDIGO) Acute Kidney Injury Work Group, 2012; Lameire et al., 2013).

Symptomatic chronic kidney disease (See Section 5.) The overt symptoms of chronic disease are usually late in the illness and deceptively non-specific. Once CKD or kidney failure have been found by the reporting of the eGFR the nephrologist has to decide whether these symptoms are indeed a consequence of the kidney disease. The lower the eGFR or more advanced the stage the more likely this is true. Patients with CKD stage 5 are almost always symptomatic though they often only acknowledge the severity after they have been dialysed, but those with CKD stage 3A and 3B are not. At one extreme the symptom complex will include the full range of uraemic consequences affecting almost all systems: ◆ Dyspnoea

is explained by pulmonary oedema, anaemia, and acidosis ◆ Anorexia and weight loss ◆ Pruritus ◆ Cognitive decline ◆ Sexual dysfunction by the central effects of the elusive uraemic toxins ◆ Skeletal discomfort and proximal weakness by secondary hyperparathyroidism

Nephritic syndrome (See Chapter 46.) This term describes the combination of oedema, hypertension, glomerular haematuria, and proteinuria (not in the ‘nephrotic’ range) with or without a reduction in GFR. A  looser definition is glomerular haematuria with any of the features. Unlike the nephrotic syndrome the patient has evidence of a significantly expanded extracellular volume with a raised jugular venous pressure. This and the oedema are attributed to sodium and water retention caused by an acute inflammatory injury to the glomeruli. This is a classical complication of beta haemolytic streptococcal infection in children. It is rare in adults. The term is not much

The patient has a systemic disorder known to be complicated by renal involvement There are many conditions in which the kidney is the victim of collateral damage and this can be severe enough to mean that the nephrologist has to take responsibility for overall care.

Metabolic diseases and inherited disorders (See Chapters 149, 167, 334.) The most important is diabetes mellitus that is now the single most common cause of end-stage renal disease. CKD clinics

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assessment of the patient with renal disease

are now dominated by patients with earlier stages of diabetic nephropathy hoping that good conservative management will prevent or delay progression. Unfortunately many have passed beyond the point of reversibility so the major contribution of the nephrology clinic is in helping to find a tolerable and effective combination of blood pressure-lowering drugs. Tuberous sclerosis, sickle cell disease, and other rarer disorders such as Anderson–Fabry disease or cystinosis also cause renal failure and their care has to be shared with experts in their other manifestations.

Malignancy (See Chapters 60, 63, 150, 172, 251.) Apart from the obstructing effects of solid tumours on the renal tract there are non-metastatic effects such as membranous and membranoproliferative glomerulonephritis and hypercalcaemia. Chemotherapy with agents such as cisplatin has adverse effects on the kidney which if extreme make renal replacement necessary. The tumour lysis syndrome is less common now that the risks have been recognized but still occurs in patients with high tumour burdens (especially leukaemias) responding to effective chemotherapy. This is a renal emergency requiring prolonged dialysis to control potassium, urate, and phosphate concentrations. The most common joint malignancy-kidney problem is in paraprotein disorders the effects of which range from acute cast nephropathy to the deposition disorders such as AL amyloid, lightand heavy-chain deposition disease.

Infection (See Sections 8 and 11.) Sepsis and shock are the most common cause of AKI in the hospital setting but there are community acquired infections that present with life-threatening renal injury too. The best examples are falciparum malaria, Escherichia coli O157 causing haemolytic uraemic syndrome (HUS), leptospiral infection causing Weil disease, hantavirus infection, and post-infectious proliferative glomerulonephritis (usually a consequence of beta haemolytic streptococcal infection). Subacute and chronic infections can lead to glomerular injury too:  hepatitis B and C, HIV, and bacterial endocarditis are the most common. Renal tuberculosis is quite rare in the developed world but not so in emerging economies.

Auto-immune inflammatory disorders (See Chapters 156, 162, 165, 166.) The management of the renal consequences of the vasculitides especially SLE, HSP, systemic sclerosis, the polyangiitides are a significant part of the acute and long-term workload of clinical nephrologists. Sarcoidosis and Sjögren syndrome can involve the kidney and evidence of interstitial inflammation obtained from

renal biopsy provides justification for immunosuppressive treatment which is otherwise usually held in reserve.

Effects of drugs (See Chapter 362.) Most drugs are two-edged swords and the prescribers dread finding changes in renal function especially if they are uncertain as to whether the drug is really the cause. This is a particular problem in oncology (cisplatin and intravenous pamidronate), rheumatology, and infectious disease (antiretroviral and antituberculosis drugs, high-dose aciclovir and sulphonamides, and amphotericin are prime examples). Although drug withdrawal is an option, a definite diagnosis of the nature of the kidney injury is preferable. An allergic interstitial nephritis will require active treatment not just stopping the agent.

Pregnancy (See Chapter 250.) Pregnancy is a systemic state in which the kidneys are vulnerable especially if there is any underlying renal disease. The catastrophic effects of the later specific complications such as eclampsia, haemolysis, elevated liver enzymes, low platelets (HELLP) syndrome, and HUS almost always require nephrology input.

Other system failure (See Chapters 247–9.) Patients with the end stages of heart and liver failure are usually referred with oliguria, disproportionate plasma urea concentrations, and hyperkalaemia, all explained by a combination of the system failure and the valiant pharmacological attempts to ameliorate their condition with various diuretics, ACEIs, and spironolactone. They are usually hypotensive and the decision on whether to offer renal support is finely balanced, especially if the underlying cause is irremediable.

References Ftouh, S., Thomas, M., on behalf of the Acute Kidney Injury Guideline Development Group (2013). Acute kidney injury: summary of NICE guidance. BMJ, 347, f4930. Hole, B., Whittlestone, T., and Tomson, C. (2014). BMJ, 349, g6768. Kidney Disease: Improving Global Outcomes (KDIGO) (2012). KDIGO clinical practice guideline for glomerulonephritis. Kidney Int Suppl, 2, 1–274. Kidney Disease: Improving Global Outcomes (KDIGO) Acute Kidney Injury Work Group (2012). KDIGO Clinical Practice Guideline for Acute Kidney Injury. Kidney Int Suppl, 2, 1–138. Kidney Disease: Improving Global Outcomes (KDIGO) CKD Work Group (2013). KDIGO clinical practice guideline for the evaluation and management of chronic kidney disease. Kidney Int Suppl, 3, 1–150. Lameire, N. H., Bagga, A., and Cruz, D., et al. (2013). Acute kidney injury: an increasing global concern. Lancet, 382, 170–9. Winearls, C. G. and Bass, C. (1994). The loin pain haematuria syndrome. Nephrol Dial Transplant, 9, 1537–9.

CHAPTER 4

Kidney disease-focused history taking Christopher G. Winearls Introduction Each referral must be approached with an open mind. No assumptions should be made and every statement should be checked. One should not believe or disbelieve what has been asserted.

The presenting complaint This will usually be in the pick list described in Chapter 3. One should start with the simple introduction, ‘I have been asked to see you because [presenting complaint] but I would like to know what you believe to be the problem.’ What concerns the patient is not always what concerns the doctor. For example, a patient consults a primary care physician because of tiredness, and is found to be anaemic and have renal impairment. It is easy to accept this sequence as causes and effect but it is often not so. Ten per cent of the population have ‘chronic kidney disease’ (CKD) but a minority are anaemic so one has to dissect the problem not just agree with the proposition.

The history of the presenting complaint This involves a detailed exploration of the onset, duration, progression, alleviating and aggravating features, and associated symptoms. This is well illustrated by the visible haematuria of mesangial immunoglobulin (Ig)-A disease which is of acute onset, painless, may follow an infection, and is of short duration. It is quite different from that of a bladder tumour. The patient’s understanding of medical words should be explored. What do they mean by ‘UTIs’, cystitis, ‘kidney pain’, migraine, angina? The same applies to the phrases in the referral which will often include vague and often misleading terms such as chronic pyelonephritis, essential hypertension, and ‘PET’.

The past medical history One is greatly assisted by a good general practice record of attendances or the hospital notes which should be examined from the first page. The past history may come as a cryptic computer printout that has been assembled over years and passed from one family practitioner to another. Each should be asked about and the basis of the diagnosis confirmed. This needs to be exhaustive, with prompting because patients have

variable memories and notions of what constitutes a significant medical problem, for example, diabetes may be considered a longstanding background nuisance rather than a relevant condition ‘because I have had it for so long and it does not cause me any problem’. Childhood illnesses are particularly difficult as the adult patient will not be accompanied by a parent and may have ‘grown out’ of the problem. Reflux nephropathy is often suspected from the story of the individual being a sickly child, with frequent fevers, courses of antibiotics, or enuresis which eventually resolved. Every operation, every medication, every hospitalization should be recorded. In women the history of pregnancies can be very revealing, for example, proteinuria at booking or blood pressure concerns that do not fit pre-eclampsia. Men have fewer opportunistic medical assessments but findings at employment, insurance, or military service medicals are helpful.

The drug (medication) history One is often informed of the currently prescribed drugs, not those that may have triggered the problem or exacerbated it and have been discontinued. Examples of the missing culprits include lithium for bipolar disorder; 5-aminosalicylic acid for ulcerative colitis; proton pump inhibitors; gold or penicillamine for rheumatoid arthritis; intravenous bisphosphonates for breast cancer; cisplatin; non-steroidal anti-inflammatories; angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin receptor blockers (ARBs); and calcineurin inhibitors for non-transplant indications. There is seldom mention of the ‘over-the-counter’ drugs but the referrer may be unaware of these so the patient should be quizzed. Perhaps the most notorious example of this problem is Chinese herbal remedies which contain aristolochic acid.

The family history This is perhaps one of the most neglected elements of history taking, for doctors rely on the catch-all question, ‘Do any diseases run in your family?’ Instead one should ask about causes of death and disability in three generations. Adult polycystic kidney disease commonly declares itself because a parent is on renal replacement treatment but there are families in which the affected parent has died prematurely, has been estranged or has not yet been diagnosed.

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The social history Patients are quite surprised that one wants to know so much about their ethnicity, where they were born, where they have lived, where they have travelled, what work they have done, what their habits—personal, sexual, smoking, drug use illicit and recreational—are. These provide clues to disorders such as HIV-associated nephropathy, focal segmental glomerulosclerosis, Chinese herb nephropathy, renal tract tumours, and atherosclerotic renal arterial disease. Of course serious renal disease, especially when renal replacement treatment is required, has far reaching implications.

One has to explore employment, relationships and domestic arrangements.

The review of systems This can be tedious and unrewarding especially if the patient is trying to be helpful and agrees that they do have symptom such as chest pain or breathlessness ‘occasionally’. It is best to leave the question open allowing them to declare and describe a symptom spontaneously. ‘How is your breathing?’ is preferred to ‘Do you ever get breathless?’ This said, one does have to be quite direct on sensitive issues such as sexual function.

CHAPTER 5

Kidney disease-focused features on examination Christopher G. Winearls Introduction This depends on the context and clinical presentation and the emphasis will be different too. Tell-tale signs are often unnoticed in the general examination of the eyes (lecithin cholesterol acyltransferase deficiency (LCAT) deficiency, Fabry disease, corneal calcification), the skin (vasculitis, Anderson–Fabry disease), the optic fundus (haemorrhages and exudates, papilloedema), and the hands (nail patella syndrome, splinter haemorrhages of SLE and subacute bacterial endocarditis) (see Figs 5.1 and 5.2). In the cardiovascular system, one is most interested in the volume status (the pulse, jugular venous pressure, and presence of oedema and its extent); blood pressure measured accurately with the an appropriate-sized cuff (the patient rested and lying down for 10 minutes and then standing); the left ventricular (LV) impulse for signs of LV hypertrophy (LVH), murmurs, and a pericardial friction rub; and the peripheral pulses. In the respiratory system one is looking for pleural effusions, signs of chronic suppurative lung disease, and clues to bronchial malignancy. In the abdomen it is the prominence of the kidneys, the bladder, and the presence of renal bruits that one wants to record. Pelvic examination is not usually routine but the slightest hint of bladder outflow problems or obstruction requires an examination of the prostate or the uterine cervix. In the nervous system one is interested in cognitive function reflecting uraemia; peripheral neuropathy in uraemia and diabetes; and autonomic neuropathy in diabetes and AL amyloid.

Investigations These will be arranged after the completion of the examination but there are general principles that need stating (see Chapters 6–18). The rule is to start with the simple and avoid requesting a comprehensive and unselected set of blood tests, the results of which may be distract or be difficult to interpret. Before the consultation ends, one will want to know what the urine dipsticks show, for example, haematuria, proteinuria, leucocytes, and nitrites, and, if appropriate, urine microscopy (see Chapter 6). A full blood count and biochemical screen including calcium, phosphate, and liver enzymes, and a C-reactive protein are reasonable routine requests because they may provide a clue to common disorders that have rather non-specific symptoms or may be clinically silent.

Once the problem is described the investigations are directed very specifically at diagnosis and consequences. For example, if one is sure one is dealing with true chronic kidney disease (CKD) one wants imaging of the renal tract, a further estimate of GFR, urine protein quantification, an electrocardiogram and echocardiogram looking for LVH, and a parathyroid hormone test for evidence of mineral and bone disorder (MBD). The consultation is closed with a description of the working diagnosis, the way it will be resolved but avoiding a description of specific treatments, unless generic or justifiable, or indeed prognosis. This is more often the business of the follow-up. It is wise to counsel patients about the risks of searching the Internet which can cause unnecessary anxiety. There are good informative websites to which patients can be referred, for example, the website of the Edinburgh Renal Unit ().

Specific consultations The acute uraemic emergency (See Chapter 222.) In the hospital setting, the nephrologist will usually be consulted either because the diagnosis is obscure or there is a need for renal replacement treatment. This will need to be instituted urgently if there are any of the life-threatening consequences of renal failure: pulmonary oedema, metabolic acidosis, hyperkalaemia, and encephalopathy. The same rules apply to community-referred patient after which the diagnostic algorithm is followed seeking prerenal, renal, and postrenal causes. As emphasized earlier, the clinical method takes precedence over investigation especially as physical examination is not usually diagnostic. Laboratory tests tend to be confirmatory rather than immediately diagnostic except in the silent renal diseases such cast nephropathy and anti-glomerular basement membrane (GBM) disease. There should be a low threshold for performing a renal biopsy unless there is a very obvious cause.

The patient with CKD stages 3–5 (See Section 5.) These referrals are made for one of two reasons. First because an estimated GFR (eGFR) has been interpreted as a stage of CKD, for example, ‘an eGFR of 45–59 mL/min/1.73m2 indicates stage 3A kidney disease’. A decision as to whether individuals with a reduced eGFR need a formal nephrological assessment will depend on their

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(A)

(B)

(C)

(D)

(F)

(I)

(J)

(E) (G)

(K) (H)

Fig. 5.1  Physical signs on general examination of the hands and peripheries. (A) White bands in the nails—evidence of hypoalbuminaemia in a patient who had an episode of nephrotic syndrome. (B) Palmar crease hyperpigmentation in a patient referred with hyperkalaemia caused by adrenal failure (Addison’s disease). (C) Broad fingers in a potential living donor found to be hypertensive and acromegalic. (D) White shiny hands in a patient with scleroderma. (E) Limb purpura in a patient with the nephritic syndrome caused by type 2 essential cryoglobulinaemia. (F) A digital infarct in a patient with microscopic polyangiitis. (G) Widespread purpura in a young man with AKI and disseminated intravascular coagulation caused by meningococcal septicaemia. (H) Gouty tophus in a patient on long-term diuretics. (I) Finger pulp infarct in a patient with nephritic syndrome caused by endocarditis. (J) Digital ischaemia in a patient with AKI and disseminated intravascular coagulation following a dog bite. The infecting organism was Capnocytophaga canimorsus. (K) Angiokeratoma corporis diffusum I in the bathing trunk distribution in a patient with Anderson–Fabry disease.

underlying co-morbid conditions, age, and the rate of change. Is this actually acute kidney injury (AKI) in an early stage or CKD? The common trap is the older patient with non-specific symptoms and a lower than predicted GFR and a bland urine sediment attributed to ‘CKD’ who actually has AKI caused by interstitial nephritis or myeloma. These dilemmas are explored and recommendations made in the Kidney Disease: Improving Global Outcomes (KDIGO) clinical practice guideline (Lameire et al., 2013).

The second reason is that a patient has presented with symptoms, for example, of anaemia and routine testing has shown that renal function is more or less abnormal.

The first referral The first task is to confirm that the patient does indeed have CKD. The eGFR may misrepresent the true renal function so one needs to take account of ethnicity, intercurrent events, the consumption of

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(A)

(B)

(D)

(E)

(F)

kidney disease-focused features on examination (C)

(H)

(G)

(I)

(J)

Fig. 5.2  Signs in the eyes and the face in patients with kidney disease. (A) The puffy eyelid of patient with unexplained CKD G3a A1 who was found to have myxoedema. (B) Limbic calcification in a young woman on long-term haemodialysis with autonomous hyperparathyroidism. (C) Collapsed nasal bridge (saddle nose) in a patient with granulomatosis with polyangiitis. (D) Subconjunctival haemorrhages in a patient with Weil’s disease. (E) Skin ecchymoses and jaundice in a man with AKI and Weil’s disease. (F) Cloudy corneas in a woman with LCAT deficiency causing CKD. Courtesy of Prof. A. J. Rees. (G) Prominent subconjunctival blood vessels in a patient with Anderson–Fabry disease, who presented with proteinuria and acroparaesthesia. (H) Differing skin pigmentation in a young woman with membranous nephropathy who had used mercury containing skin-lightening cream. (I) Facial lipodystrophy in a woman with nephrotic syndrome caused by minimal-change glomerulonephritis. (J) The taut skin and telangiectasiae in a patient with scleroderma and AKI caused by hypertension as part of a scleroderma crisis.

drugs that alter creatinine concentration but not GFR (trimethoprim and cimetidine), or those that do alter GFR reversibly such as non-steroidal anti-inflammatory drugs. The second is to establish the chronicity of the CKD and its immediate relevance to the health of the patient. CKD has greater implications in younger subjects than in much older people with co-morbidity. The third is to find out the

rate of change which will inform the conversation about prognosis. One can establish a cause in approximately 80% of patients but in the older group one often has to resort to suggesting vascular and hypertensive disease. Intensive attempts to make a diagnosis are only justified if a remediable or modifiable cause is possible. Finally one documents the presence or absence of CKD complications.

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The follow-up patient Patients with CKD of at different stages constitute a majority of the attendees at general nephrology clinics. They seldom fail to attend and until the more advanced stages have few symptoms attributable to their kidney disease. They will nevertheless share a variety of problems fearing that they are manifestations of CKD or because they have been told so. When asked how they are feeling, they will generally say, ‘I am fine’. It is advisable to press them for symptoms of uraemia and if possible engage their accompanying relative. Some patients are in denial, others have accommodated their disabilities, and a few fear that any complaint will lead to the institution of dialysis. One needs to move on to individual elements of the CKD syndrome including its cause, the complications, and then the plans for the future. Much of this will rely on the laboratory tests, best undertaken before the actual visit. Measuring the blood pressure, the weight, and the demeanour are a good neutral start, setting the patient at their ease. That done, a conversation on their understanding of the nature of CKD, its potential for progression, and what one might do when ‘the kidneys are no longer doing enough to keep you healthy’ is needed. As patients dread reaching end stage, one reviews the serial eGFRs, answering the inevitable question, ‘What can I do to avoid the need for dialysis?’ This allows one to reinforce the need for good blood pressure control, adherence to a sensible diet, and forgoing tobacco. One then checks the complications including anaemia, MBD, and cardiovascular risk. Cardiovascular disease occurs prematurely in patients with CKD and many patients will die of it before reaching end-stage renal disease. Consideration should be given to measures to reduce cardiovascular risk such as lowering low-density lipoprotein (LDL) cholesterol and controlling blood pressure (which is already a focus of slowing progression of their kidney disease). Do not be misled by a ‘normal’ LDL cholesterol concentration: this predicts cardiovascular risk poorly in patients with CKD and lowering LDL cholesterol is effective regardless of the starting concentration. Depending on the stage (usually late stage 4)  one talks about options ranging from transplantation, through various forms of dialysis, to medical care without dialysis (sometimes called maximal conservative care—not a useful term because it suggests that there is minimal conservative care). In some services the patients approaching stage 5 are referred to ‘low clearance clinics’. These have the advantage of reliable adherence to process, easy access to dietetic, dialysis, vascular access and transplantation, and other advice and a well-mapped pathway. It does, however, separate the patient from his or her nephrologist who may have cared for the patient for many years. It is possible for each stage 4/5 patient to have a pro forma or their own logbook to record and ensure that all the basic elements of CKD care have been covered. Their drug regimens should be reviewed particularly if there may have been additions made by other disciplines. As the GFR falls it is worth asking whether the ACEI should be continued, especially if the K+ is drifting above and beyond the upper limit of normal.

The patient with unexplained haematuria (invisible and visible) The patient will be referred to a nephrologist in the first instance if the primary care physician has excluded a trivial or non-urinary tract cause (gynaecological), confirmed that the abnormality is persistent, the patient is under 50 years of age, and if there is collateral

evidence of kidney diseases such as proteinuria or renal dysfunction. Referral to a urologist first is advised in patients over the age of 34–40 years or if they are younger if there is no obvious renal cause. They will seek to exclude stones, infections including in the prostate, non-infective inflammation, and tumours. If a urologist has excluded an obvious structural cause, the issue for the nephrologist is to decide whether this finding is significant, that is, is there an underlying disorder of the kidney requiring a renal biopsy for diagnosis? This will be decided on the basis of the red cell appearances being suggestive of a glomerular origin, the presence of any collateral evidence of renal disease, a ‘need to know’, and the policies of the medical system (Hole, Whittlestone and Tomson, 2014). A nephrologist is not usually involved until primary care physicians and urologists have exhausted common causes which are infection, stones, inflammation, and a malignancy anywhere from the renal parenchyma to the urethra. The timing of the haematuria helps a little: observed early and then clearing suggests a urethral problem; at the end of micturition, a bladder cause; and throughout, an upper urinary tract cause. The appearance is important—fresh red haematuria with clots points to bleeding from the urinary tract, muddy brown (oxidized blood) suggests a renal parenchymal cause such as glomerulonephritis. When such patients do reach the nephrologist he/she will want to be reassured that the cause of ‘haematuria’ is indeed blood and not haemoglobin or myoglobin. The absence of red cells on urine microscopy will be the alert. There are other causes of discoloured urine but they do not look like haematuria. Patients on rifampicin produce orange urine; in porphyria the urine has to stand before changing colour. The common three renal causes of macroscopic haematuria are immunoglobulin (Ig)-A disease, especially during periods of infection; anti-GBM disease; and adult polycystic kidney disease. Finally there is the problem of factitious haematuria which may be part Munchhausen syndrome. The complaint of haematuria is intermittent and inconveniently (for the doctor) difficult to confirm. An immediate cystoscopy revealing clear urine within the bladder and from both ureteric orifices within hours of a bloody urine being provided is the best way to establish the diagnosis. The handling of this finding is challenging but it allows both urologist and nephrologist to desist from further intrusive investigation. This presentation is sometimes in combination with loin pain in what is blandly called the ‘loin pain haematuria syndrome’. This is a common entity in which the pain is out of proportion to the examination, imaging, and pathology findings which are minimal and non-specific. It is resistant to conventional analgesia and the doses and duration of opioid analgesics exceptional. The descriptions of the pain are graphic and appear exaggerated as the pain is claimed to be present at the time (Winearls and Bass, 1994).

The patient with proteinuria or the nephrotic syndrome (See Chapters 50 and 52.) This consultation will concentrate on the indication for a renal biopsy and dealing with the consequences of the nephrotic state. The key question in the asymptomatic patient with proteinuria is whether a biopsy will provide useful prognostic information and change management. The case for a biopsy is very strong in nephrotic patients as it is dangerous to offer trials of treatment with corticosteroids except

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in children. Serological tests can offer clues but seldom certainty. When tests for antiphospholipase A2 receptor antibodies become widely available the decision will be more difficult in those adults suspected of suffering from membranous nephropathy who have a positive test. The nephrotic patient will want relief from uncomfortable oedema and the nephrologist will have to make a judgement about the need or otherwise for anticoagulation KDIGO clinical practice guideline for glomerulonephritis.

The patient with diabetes mellitus (See Chapter 149.) There are specific extra tasks in patients with diabetes mellitus and renal disease. The first is to test the assumption that the renal disease is indeed a consequence of diabetes. It is assumed to be so especially if the patient has longstanding diabetes, retinopathy, and proteinuria. Even when these are present the explanation can be another disease. A sudden change in renal function, ‘nephrotic’ range proteinuria, haematuria, and new systemic symptoms should alert the nephrologist to another cause. Diabetes occurs in approximately 5% of the population so it is no surprise when they are found to have conditions such as IgA nephropathy or steroid-sensitive nephrotic syndrome. Control of diabetes in patients with reduced renal function can be difficult. Insulin requirements fall and sulphonylurea drugs have prolonged half-lives. The patients fearing hypoglycaemia may eat more to prevent this. Metformin can rarely cause lactic acidosis if the dose is not modified, or an additional illness supervenes. The advice on metformin use is different in the United States from Europe. Glipizide is the preferred sulphonylurea in renal failure because it is not excreted by the kidney. Tight control of diabetes does not improve outcomes in patients with established nephropathy and may actually have an adverse effect on survival so haemoglobin A1c targets are less stringent. Of all patients with CKD, diabetics have the worst cardiovascular risk so this aspect of management—smoking cessation, statin use, good blood pressure control, and a low threshold for investigation of ischaemic heart disease—should be emphasized. In older patients, target blood pressures should be a matter for clinical judgement and are > 120/80 mmHg. One should emphasize to the patient that blood pressure control offers the best chance of delaying the need for renal replacement and reducing the risk of cardiovascular events. This may require combinations of drugs but not angiotensin receptor blockers and angiotensin-converting enzyme inhibitors (ACEIs) as so-called dual renin–angiotensin–aldosterone system blockade is associated with an increased risk of adverse events. See the KDIGO guideline for further information (Kidney Disease: Improving Global Outcomes (KDIGO) CKD Work Group (2013).

The review of the haemodialysis patient (See Chapter 255.) Depending on the frequency of review this can be a short or a long consultation. The patient and the nephrologist are likely to have different agendas and it is best to start with theirs. One wants to gain an understanding of how dialysis is affecting their lives and whether they are actually gaining the maximum benefit. A  ‘Good Dialysis Index’ proposed by John Agar () can be adapted. For example, one can start with the open-ended questions to allow the patient to tell one how treatment is affecting their everyday life: ◆ ‘How are you feeling today?’ ◆ ‘What symptoms do you have during and after your treatment?’ ◆ ‘Can you eat and drink what you want?’ ◆ ‘How does having a dialysis treatment affect what you want to do?’ ◆ ‘If you are not already retired, are you working?’ ◆ ‘Have you been in hospital recently? If so, for what?’ One can then move to the nephrologist’s agenda: ◆ ‘How is your vascular access?’ ◆ ‘How many hours of treatment are you actually having?’ ◆ ‘How much weight do you need to take off on your first dialysis of the week?’ ◆ ‘How often do you have a problem with low blood pressure during treatment?’ ◆ ‘How many blood pressure tablets are you taking?’ ◆ ‘How much “Epo” do you need? Is your haemoglobin in range?’ ◆ ‘Is your phosphate a problem? How many binders do you need?’ ◆ (If appropriate) ‘Are you on the transplant waiting list?’ ◆ (If appropriate) ‘Can we talk about home haemodialysis?’ ◆ Has your cardiovascular risk been addressed? Are you on a statin?’ The examination is often neglected in busy clinics but should include a look at the fistula, the skin, the pulses, the muscles and the ease of rising from a chair, the heart sounds to be sure that calcific aortic stenosis is not stealthily developing, and the lung bases for signs of chronic pulmonary congestion. Only then should one scan the laboratory results reflecting good treatment of anaemia, bone mineral balance, and nutrition.

The review of the peritoneal dialysis patient (See Chapter 263.) These consultations are generally conducted with the peritoneal dialysis (PD) nurse specialists who will review the practical issues. They may have undertaken this in the patient’s home before nephrology review. ◆ Is the catheter working and is the exit site clean? ◆ Is the bowel habit regular? ◆ Is the regimen correct and controlling the concentration of uraemic markers and fluid balance? This is assisted by the peritoneal equilibration test and an ‘adequacy test’. They will assess nutrition both under and over. ◆ Is the technique being practised properly and is a refresher course of instruction needed? The nephrologist will concentrate on the overall well-being of the patient, checking that they are still content with their choice of modality and reassuring them that PD is delivering good renal replacement but mindful of the possibility that a switch to haemodialysis may become necessary in the future as residual function declines or the function of the peritoneal membrane wanes. The blood pressure and cardiovascular risk factors will be scrutinized along with the medication list, specifically asking whether the

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cardiovascular risk been addressed and that prescription of a statin has been considered. The aim of this is to reduce the tablet load, particularly blood pressure agents, analgesics, and often unnecessary drugs such as proton pump inhibitors (PPIs) and H2 antagonists. The need for and dose of an erythropoiesis-stimulating agent, iron repletion, and markers of MBD will be examined. Finally they will want to be sure that renal transplantation has been considered and if so the patient is listed

The review of the stable transplant patient (See Section 13.) These routine visits require a systematic review of the function of the renal transplant and the health of the patient. Unless specific issues are volunteered one moves to the checklist. The threshold for investigating for opportunistic infections and malignancy should be low. ◆ Graft function is assessed by the plasma creatinine and compared to the best achieved and the last measurement. The eGFR has not replaced the plasma creatinine in routine care. Urine is not routinely examined for evidence of infection but dipsticks for proteinuria and invisible haematuria are advisable if only to be alert to recurrent disease. ◆ A review of the medication, taking the opportunity of stopping drugs such as H2 antagonists, PPIs that have hung over from the early postoperative phase, the dose of the main immunosuppressive drugs based on trough levels, and white cell count. Is the patient on appropriate agents to reduce cardiovascular risk (such as a statin) and/or bone protection? Can the blood pressure medications be reduced or simplified?

◆

The blood pressure should be measured properly, that is, after the patient has been sitting quietly for 10 minutes. ◆ There should be a brief look at the skin and the organization of the annual dermatology review confirmed.

The assessment of the potential living kidney donor (See Chapter 277.) This is a strange consultation for a nephrologist because the patient is usually healthy and one’s task is to confirm this but one has to be sure not to miss a hitherto unrecognized reason not to proceed. One starts by satisfying oneself that the potential donor is not under any duress and is well informed about the procedure and its consequences. There are three issues: ◆ Is it safe for the recipient? This is essentially asking whether there are any chronic blood-borne viral infections or potential abnormalities of the kidney that may preclude transplantation, for example, vascular abnormalities or renal tumours. ◆ Is the kidney healthy enough to be transplanted—is the GFR satisfactory, is there proteinuria or any evidence of CKD? This is a concern for both recipient and donor. ◆ Is the donor healthy enough to undergo non-essential surgery and are the risks of being uni-nephric acceptable? Occasionally one has to rely on a 24-hour ambulatory blood pressure monitoring, genetic testing in ADPKD, or even renal biopsy if there is invisible haematuria.

CHAPTER 6

Urinalysis Walter P. Mutter

Urinalysis aids in the diagnosis of renal disease especially in cases when a renal biopsy is not immediately available or is contraindicated. It is most informative when done by the treating physician with knowledge of the clinical context (Fogazzi et al., 1998). Inspection is done by eye. Routine chemical analysis is done by dipstick but urine microscopy is essential for it may reveal abnormalities even when chemical evaluation is normal (Szwed and Schaust, 1982).

Osmolality is a measure of dissolved particles in solution with a normal range of 50–1200 mOsm/kg. Specific gravity is generally directly proportional to urine osmolality but may be disproportionately higher in the presence of larger molecules such as glucose and iodinated contrast when measured by refractometry. Urine dipsticks are usually used to estimate specific gravity as they are rapid and convenient. They do not detect non-ionic substances such as radiocontrast, urea, or glucose (Siemens, 2010). Alkaline urine may cause falsely low readings. Proteinuria in the range of 100–750 mg/dL may cause elevated readings (Siemens, 2010).

Inspection

Protein

Colour

(See Chapter 7 for further detail.) Proteinuria may be quantified by dipstick, protein to creatinine ratio, albumin to creatinine ratio, or by a 24-hour urine collection. Reagent strips are most sensitive to albumin. If non-albumin protein is suspected, analysis of total urine protein using turbidometric or dye-binding techniques is recommended (Fogazzi et al., 2008a). Sensitivity is in the range of 15–30 mg/dL albumin. Microalbuminuria, defined as urine albumin excretion between 30 and 300 mg/day or 20 and 200 mg/L, may not be detected by dipstick testing. Very alkaline urine and gross haematuria may yield false-positive results (Pugia et al., 1999).

Introduction

Normal urine colour varies from almost clear to dark yellow as a result of urobilin pigment. The depth of the colour depends on the urine concentration. Table 6.1 lists typical causes for abnormal urine colour.

Clarity Cloudy or turbid urine is most commonly caused by urinary tract infections but may be due to contamination from vaginal secretions, faecal material, gross haematuria, crystals, or lipids as in chyluria (Fogazzi et  al., 2008a). Larger particles including cells, crystals, and casts will usually sediment with centrifugation.

Odour Ammonia ions give urine its typical odour. A foul or feculent odour may signify urinary tract infection. Sweet or fruity smelling urine may be present in diabetes, ketosis, or maple syrup urine disease. Phenylketonuria gives off a musty odour, isovaleric academia a sweaty foot odour, while hypermethioninaemia gives off a rancid or fishy odour (Fogazzi et al., 2008a).

Chemical analysis Chemical analysis of the urine is typically performed using a chemical reagent dipstick composed of absorbent squares that change colour after application of urine. Colour change may be interpreted by eye or by a colorimeter. A standard dipstick will measure up to 10 urine parameters including protein, blood, leucocytes, nitrite, glucose, ketones, pH, specific gravity, bilirubin, and urobilinogen.

Urine concentration Urine concentration may be measured as specific gravity or urine osmolality.Specific gravity is the weight of a solution divided by the weight of water. The normal range for urine is 1.005–1.030.

Blood The urine dipstick detects haemoglobin by measuring its peroxidase activity. Haemoglobin catalyses the oxidation of a chromogen to produce a coloured product. Myoglobin also catalyses this reaction, resulting in a positive test. A positive dipstick in the absence of red blood cells (RBCs) in the urine sediment should prompt consideration of haemolysis or rhabdomyolysis. The urine dipstick is quite sensitive to the presence of blood in the urine detecting haemoglobin levels > 0.015 mg/dL and in excess of three RBCs/μL (Siemens, 2010). False positives are possible if microbial peroxidase is present. False negatives may be seen in patients taking ascorbic acid (vitamin C) (Brigden et al., 1992).

Glucose Small amounts of glucose, usually 10 leucocytes/μL (Siemens, 2010). Sensitivity may be decreased by concentrated urine that prevents cell lysis as well as cephalexin, glucose, oxalic acid, and tetracycline (Fogazzi et  al., 2008a; Siemens, 2010).

Urine microscopy Identification of casts, cells, and crystals relies on the experience of the observer. Not surprisingly there is significant variation in the interpretation of urinary sediment findings (Wald et  al., 2009). For example, there is no consensus among nephrologists as to how many granular or renal tubular cell casts are required to support a clinical diagnosis of acute tubular necrosis (ATN) (Chawla et al., 2008). Inter-observer variability among nephrologists of common urinary findings has been reported. Ten nephrologists in a single centre agreed on the presence of dysmorphic RBCs only 31% of the time (Wald et al., 2009). However, nephrologists may perform better than hospital-based technologists in the identification of some characteristics including renal tubular epithelial (RTE) cells, granular casts, RTE casts, and dysmorphic RBCs (Tsai et al., 2005). Urine chemistry including pH, specific gravity, blood, and protein should be obtained in advance of the microscopic examination. Interval re-examination of the urine sediment is advised in patients with changing clinical status or where the initial findings are discordant with the clinical presentation.

Microscope selection Phase contrast microscopy may have advantages for cellular identification and is recommended in some guidelines (European Confederation of Laboratory Medicine, 2000; Kouri et al., 2000). However, many nephrologists use bright field microscopy and this may be the only available option (Fogazzi and Grignani, 1998). Both methods have been used for research and either is acceptable. A polarizing filter is helpful for identifying lipids and certain crystals.

Urine collection Unfortunately, there is a lack of standardization in the way urine is collected and processed in clinical practice (Fogazzi and Grignani, 1998). The following approach, adapted from published guidelines, is recommended (Fogazzi et al., 1998, 2008a). Every patient presenting to the nephrology clinic should undergo at least one complete urinalysis (Fogazzi et  al., 2008a). Written instructions should be provided to the patient in advance of a clinic visit. Ideally urine should not be collected within 72 hours after strenuous exercise or during menstruation in women. Men should retract the foreskin when present and women should spread the labia. The use of cotton wipes for urethral opening in both genders is reasonable (Fogazzi et al., 2008a). Urinary catheter specimens should be avoided. Urine should be collected midstream in a sterile plastic container. Urine from the first morning void is generally more concentrated, increasing the sensitivity for detecting abnormalities. However, prolonged stasis in the bladder may lead to lysis or degradation of cells or casts (Fogazzi et al., 2008a). Analysis within 2–3 hours is recommended as urine pH and temperature change

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urinalysis

rapidly after voiding. Formed elements such as cells and casts may degrade, especially in alkaline or dilute urine. Refrigeration at 4ºC is reasonable for short-term storage when urine cannot be examined immediately, particularly if culture is to be performed. However, this may lead to precipitation of calcium or uric acid crystals.

Preparation of the sample for microscopy Once collected, 10 mL of urine is spun in a plastic tube in a table top centrifuge at 400 G (2000 RPM for 10 minutes) then 9.5 mL of urine is aspirated or decanted and the remaining 0.5 mL agitated gently using a pipette. One drop, about 50 µL, is applied to a glass slide and covered with a glass cover slip. Examination begins at low power (100 ×) looking for cells, casts, and crystals ideally including 20 microscopic fields. Prolonged examination of 50 low power fields is appropriate for urine with few formed elements or when looking for the presence or absence of RBC casts or dysmorphic RBCs (Fogazzi et al., 2008a). Increased magnification up to 400 × is used for more detailed examination and cell identification. Urine elements may be quantified using counting chambers containing a fixed volume of urine. In the alternative, the nephrologist reports the number of objects per HPF at 400 ×. When done carefully, this technique correlates reasonably well with counting chambers (Fogazzi et al., 1998).

Crystals Crystals are commonly observed in urine, occurring in approximately 8% of unselected samples (Fogazzi, 1996). They are likely to be clinically significant when associated with renal failure such as calcium oxalate crystals in ethylene glycol poisoning, uric acid crystals in tumour lysis syndrome, and various drug crystals such as aciclovir in drug-associated renal failure (Fogazzi et al., 1998). Studies of crystalluria for establishing the diagnosis in nephrolithiasis or monitoring treatment have yielded variable results because of the overlap with the normal condition (Fogazzi, 1996). Both high and low urine pH and a low temperature promote crystal formation (Fogazzi, 1996; Fogazzi et al., 2008a). Acid urine is associated with calcium oxalate, uric acid, and amorphous urate crystals. Alkaline urine is associated with calcium phosphate, triple phosphate, and amorphous phosphates (Fogazzi et al., 1998). Examination of urine for crystals should be performed on urine as close to 37ºC as possible. If amorphous crystals are present, acidification or alkalinization of the urine may be helpful to dissolve them, clearing the field of view to observe other urine sediment findings (Fogazzi and Cameron, 1996). Calcium oxalate and uric acid crystals are soluble in alkaline solutions, whereas triple phosphate and calcium phosphate crystals are soluble in acidic solutions (Fogazzi, 1996).

Fig. 6.1  Monohydrated (biconvex disc) and bihydrated (bipyramidal) calcium oxalate crystals.

may appear as joined semicircles with central indentation known as the ‘dumbbell’ form (Martinez-Giron, 2008). They may be seen in a broad pH range of < 5.4–6.7. Factors favouring formation include low pH, concentrated urine, and high oxalate ingestion or absorption.

Uric acid (Fig. 6.2) Uric acid may form several types of crystals in patients with uric acid nephrolithiasis, hyperuricosuria, and tumour lysis syndrome.

Calcium oxalate (Fig. 6.1) Calcium oxalate crystals may be present in normal urine but are frequently seen in patients with nephrolithiasis, on a high-oxalate diet, metabolic defects, vitamin C in high doses, and ethylene glycol poisoning. Calcium oxalate dihydrate usually forms square or rectangular shapes with characteristic diagonal highlights forming a bi-pyramidal shape known as the ‘envelope’ form. Ovoid shapes are also seen (Fogazzi, 1996). Calcium oxalate monohydrate crystals

Fig. 6.2  Uric acid crystals (lozenge variety). Inset: the same under polarized light.

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Flat diamond or rhomboidal ‘lemon’-shaped sheets are most common. Polarization reveals beautiful multicolour birefringence. Amorphous urates appear as granular debris in urine of low pH, < 5.4–5.8 (Fogazzi, 1996).

initiating appropriate therapy if renal failure is present (Perazella, 2003; Martinez-Giron, 2008).

Phosphates

Red blood cells (Figs 6.3a and 6.3b)

Phosphate crystals include calcium phosphate, triple phosphate, and amorphous phosphates. Triple phosphate crystals are rectangular prism-shaped crystals known as ‘coffin lids’. Calcium phosphate crystals may also appear as stars or needles. Amorphous phosphates appear as fine granular clumps.

Miscellaneous crystals Miscellaneous crystals include cholesterol crystals which are rectangular plates and cystine crystals which are hexagonal. Tyrosine and leucine crystals may be observed in liver disease.

Crystals in renal failure Tubular damage and obstruction by crystals may cause renal failure. Examples are acute uric acid nephropathy, calcium oxalate in ethylene glycol poisoning, and various drugs including aciclovir, triamterene, indinavir, methotrexate, and sulphonamides (Fogozzi, 1996; Perazella, 1999; Ting et al., 2000). Drug crystals should be suspected in subjects receiving a high-dose, intravenous infusion, and those with volume depletion or renal failure. Pleomorphic and atypical shapes may suggest drug-induced crystals. Commonly observed drug crystals include aciclovir (needles), protease inhibitors such as indinavir (plates and stars), and amoxicillin (needles, sheaves of wheat) (Fogazzi et al., 2008a). The presence of crystals in the urine is not diagnostic of crystal-induced renal failure but should prompt the clinician to consider altering therapy, removing the offending drug, or

Cells RBCs appear as translucent biconcave disks of between 4 and 7 μm in diameter. RBCs may be isomorphic (similar to circulating RBCs) or dysmorphic (abnormal in shape (poikilocytosis) or size (anisocytosis)) (Thal et al., 1986). Isomorphic RBCs are associated with non-glomerular haematuria from a genitourinary or external source. Urinary catheterization is a common cause in the hospitalized patient (Hockberger et al., 1987). RBCs may appear dysmorphic as a result of changes in pH, osmolality, and protein concentration (Wandel and Kohler, 1998). For example, shape transformation from discocyte (normal) to echinocyte (crenated) to stomatocyte (indented ball) by altering urine chemistry is rapid and reversible (Wandel and Kohler, 1998). These changes may not indicate renal disease. Forms of dysmorphia associated with renal disease include anulocytes, ghost cells, schizocytes, codocytes, and knizocytes (Kohler et al., 1991). The cause of dysmorphic RBCs is unknown but might arise from passage through the glomerular barrier or tubules (Schuetz et al., 1985; Kubota et al., 1988). Dysmorphic RBCs are associated with glomerular bleeding but may also be seen in non-glomerular and tubulointerstitial disease (Birch and Fairley, 1979; Fairley and Birch, 1982; Schramek et  al., 1989; Kohler et  al., 1991). A  review of five studies from 1982 to 1984 showed that dysmorphic RBCs were seen in 95% of patients with a glomerular lesion and only 8% of patients with a non-glomerular lesion (Thal et al., 1986). However, other data in patients with haematuria and proteinuria showed lower sensitivity

(a) (b) Fig. 6.3  (a) Isomorphic erythrocytes (dark cells have lost their haemoglobin content). (b) Different types of dysmorphic erythrocytes.

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and specificity (Favaro et al., 1997). Clinical application is hampered by the lack of a standard definition of dysmorphism (Thal et al., 1986). Acanthocytes (also known as G1 cells) are a unique form of dysmorphic RBC characterized by a round shape with one or two smaller, round protrusions, vesicles, or blebs attached (Kohler et al., 1991; Kitamoto et al., 1993; Lettgen et al., 1995). Crenated RBCs (echinocytes) which do not signify pathology have numerous, symmetric spiculations or spikes and may be seen in concentrated urine. They may be confused with acanthocytes but the appendages are smaller and more numerous. Although there is no standard cut-off for the number of dysmorphic RBCs or acanthocytes required to diagnose glomerular haematuria, the presence of acanthocyturia, defined as a urine containing ≥ 5% acanthocytes, may be useful. In one study acanthocyturia showed 98% specificity and 52% sensitivity for a glomerular lesion on biopsy. The sensitivity increased to 84% if four urine samples were examined (Kohler et  al., 1991; Kohler and Wandel, 1993). In patients with isolated microscopic haematuria, the presence of dysmorphic RBCs and acanthocytes is helpful. In a review of 16 patients (10 children and 6 adults) the sensitivity and specificity for the presence of ≥ 40% dysmorphic RBCs was 59% and 91% respectively for glomerular disease. Acanthocyturia ≥ 5% was 69% and 85% sensitive and specific (Fogazzi et al., 2008b). Acanthocyturia is particularly useful in identifying non-diabetic glomerular disease in diabetics where haematuria is common. In 68 patients with biopsy-proven diabetic nephropathy, haematuria was present in 62% but acanthocyturia was present in only 4%. In contrast, acanthocyturia was present in 40% of those with a superadded glomerulonephritis (Heine et al., 2004). A reasonable definition of glomerular haematuria is ≥ 40% dysmorphic RBCs or ≥ 5% acanthocytes (Fogazzi et al., 2008a).

Leucocytes (Fig. 6.4) Leucocyturia is a non-specific finding requiring clinical interpretation. Leucocytes may be seen in urinary tract infection, glomerulonephritis, tubulointerstitial nephritis, and renal transplant rejection (Fogazzi et al., 1998). Sterile pyuria, or leucocytes in the absence of bacterial urinary tract infection, is characteristic of renal tuberculosis but may also be seen in nephrolithiasis, urinary tract malignancy, analgesic nephropathy, and treated infections. Most nephrologists do not attempt to differentiate leucocytes in unstained urine. Neutrophils are the most common leucocyte observed in urine. They have a diameter of 7–13  μm with a granular cytoplasm and lobulated nucleus (Fogazzi et al., 2008a). Eosinophils may be identified by Wright’s or Hansel’s stain (Nolan et al., 1986; Nolan and Kelleher, 1988). Eosinophils have been reported in association with acute interstitial nephritis (AIN), cholesterol emboli, and glomerulonephritis (Nolan and Kelleher, 1988). Some authors have questioned the utility of looking for urine eosinophils in AIN given the low specificity (Nolan and Kelleher, 1988; Fogozzi et  al., 1998, 2008a). Macrophages are large cells 15 to > 100  μm in diameter and may be seen in various glomerular diseases or as oval fat bodies containing phagocytized lipids (Fogazzi et al., 2008a). Lymphocytes are seen with cellular rejection in renal transplant patients but require special staining (Fogazzi et al., 2008a).

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Fig. 6.4  Neutrophils with their lobulated nucleus and granular cytoplasm (phasecontrast microscopy, original magnification 400).

Squamous epithelial cells Squamous epithelial cells are large, flat, 45–65 μm in diameter, and with a single small nucleus. They may come from the bladder trigone, urethra, or genital skin. They are frequently observed, especially in women, and do not indicate pathology. When observed with bacteria and white cells, genital contamination of the urine should be suspected.

Renal tubular cells RTE cells are 11–15 μm in diameter and may be round or rectangular with a large nucleus (Fogazzi et al., 2008a). They are usually seen in severe renal disease such as ATN, glomerulonephritis, AIN, or cellular rejection of renal transplants (Mandal et al., 1985; Fogazzi et al., 1998). The presence of renal tubular cells in the absence of other pathological urinary findings is rare (Fogazzi et al., 1998). In nephrotic patients, RTE cells may contain fat droplets and appear as ‘oval fat bodies’ similar to macrophages.

Uroepithelial or transitional cells Uroepithelial cells come from the urothelial lining of the urinary tract from the bladder, ureter, or renal pelvis. They may be round, oval, or rectangular. Cells from the superficial layers are 20–40 μm in diameter with a large, clear cytoplasm while those from the deeper layers tend to be smaller, 13–20 μm in diameter, oval, with a small and granular cytoplasm (Fogazzi et al., 2008a). Transitional cells from the deeper layer are strongly associated with urologic disease including stones, cancer, and hydronephrosis (Fogazzi et al., 1995, 1998, 2008a).

Microorganisms and miscellaneous cells Bacteria are common and, of course, require culture for identification and antibiotic sensitivities. Yeasts are clear circular cells, smaller than

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RBCs, sometimes identified by their budding or dividing. The hyphal forms of yeast form long, branching tubular structures similar to tree branches. Trichomonas vaginalis may be observed as a flagellated, motile, single-cell protozoan (Mutter and Brown, 2011). Schistosoma haematobium eggs may be present and associated with non-glomerular haematuria (Fogozzi et  al., 2008a). Non-genitourinary cells need to be considered, including intestinal epithelia in patients with ileal bladders (Gai et al., 2007). Spermatozoa are also commonly seen in subjects with retrograde ejaculation. Unusual cells that are large or multinucleated might indicate a malignancy.

Oval fat bodies Lipid may be present in the urine and is common with heavy proteinuria as in nephrotic syndrome and diabetic nephropathy (Fairley and Birch, 1993). Lipid droplets are small, about 1–5  μm in diameter. When incorporated into tubular cells or macrophages they are referred to as ‘oval fat bodies’ (Fairley and Birch, 1993).

Casts Casts are linear tubular structures that form in the renal tubules, usually the loop of Henle, distal tubules, or collecting ducts. The presence of casts in the urine is defined as cylinduria. Tamm–Horsfall mucoprotein provides the scaffolding and cells present in kidney or leaked into the urine during filtration become incorporated to form a cast (Rutecki et al., 1971; Hoyer and Seiler, 1979). The presence of certain casts is strongly correlated with the presence of renal disease identified by histology. The number of casts may predict the severity of injury (Gyory et al., 1984). There are seven major types of casts: hyaline, epithelial, granular, fatty, broad, waxy, and red or white blood cells. Unusual casts such as bilirubin casts in liver failure and bacterial casts in pyelonephritis have also been observed (Lindner et al., 1980). In renal conditions that wax and wane over time such as lupus nephritis, vasculitis, and cryoglobin-associated renal disease, the presence of casts is used to monitor disease activity (Fogazzi and Leong, 1996; Fogazzi et al., 1998). For example, RBC and WBC casts may precede clinical relapse in lupus nephritis (Hebert et al., 1995).

Fig. 6.5  A hyaline cast with a ‘fluffy’ appearance due to the fibrillary substructure of Tamm–Horsfall glycoprotein.

Hyaline casts (Fig. 6.5) Hyaline casts are concretions of Tamm–Horsfall protein and may be seen in normal urine. Other proteins such as immunoglobulins, fibrin, and complement may also be present (Fairley et al., 1983). Numerous hyaline casts are seen in volume contraction, congestive heart failure, and also seen after exercise or administration of diuretics (Imhof et al., 1972).

Epithelial cell casts Epithelial cell casts may be seen in both ATN and glomerulonephritis.

Granular casts (Fig. 6.6) Granular casts consist of degenerated tubular cells and strongly suggest the presence of renal pathology (Rutecki et  al., 1971). Coarse granular casts contain partially degenerated cells while fine granular casts result from further cellular degeneration. Muddy brown casts are pigmented granular casts observed in ATN. Small

Fig. 6.6  A finely granular cast.

numbers of granular casts may be seen in any form of chronic kidney disease.

Fatty casts Fatty casts may be granular or hyaline in appearance with small circular lipid droplets. Polarization reveals characteristic Maltese crosses. Fatty casts may also contain epithelial cells.

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Broad casts Broad casts are wider than other varieties and are believed to form in the larger collecting tubules. They may appear granular, waxy, or epithelial.

Waxy casts Waxy casts are broad, well demarcated, with flat sides and ends resembling melted wax. Sometimes they may appear in a corkscrew form. They are thought to be formed from disintegrated tubular cells and are associated with advanced kidney disease.

Red blood cell casts (Fig. 6.7) RBC casts suggest intrarenal inflammation. They may be present in pyelonephritis, allergic interstitial nephritis, and particularly severe glomerulonephritis. They are relatively infrequent, appearing in 22–38% of patients with glomerulonephritis but when present are specific for inflammatory glomerular pathology (Fogazzi et al., 1998). They are particularly useful to distinguish a glomerular source of haematuria in those with isolated haematuria. In a review of 16 patients with isolated microscopic haematuria, RBC casts were 100% sensitive for glomerular disease (Fogazzi et al., 2008b).

Leucocyte/white blood cell casts WBC casts may be seen in pyelonephritis, interstitial nephritis, and glomerulonephritis. Other urinary findings help determine the clinical significance. The finding of WBC casts may be particularly helpful when AIN is suspected and there is no evidence of urinary tract infection.

Clinical interpretation Urine sediment findings need to be interpreted within the clinical context. A bland urinary sediment does not exclude the presence of

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significant renal disease. It is a common finding in vascular disease and may be consistent with a recovering renal injury (Goorno et al., 1967; Eiser et al., 1979; Fogazzi et al., 1998).

Tubulointerstitial disease Patients with tubulointerstitial disease may have a bland urine sediment. AIN may be characterized by pyuria and WBC casts in the absence of urinary tract infection. However, in a case series of 21 patients with biopsy-proven AIN, only 14% showed WBC casts. Interestingly 29% had RBC casts. WBCs were present in 57% and almost 50% had haematuria (Fogazzi et al., 2012). Eosinophiluria is generally not sensitive or specific enough to be clinically useful (Nolan and Kelleher, 1988; Fogazzi et al., 2008a).

The sediment in nephrotic syndrome The nephrotic syndrome is associated with oval fat bodies, fatty casts, and cholesterol crystals (Moriggi et al., 1995). Patients with nephrotic syndrome caused by diseases such as membranous nephropathy and focal and segmental glomerulosclerosis may have haematuria but RBC casts are rarely seen (Fogazzi et al., 2008a). Changes in the sediment such as the appearance of RBCs might indicate a clinical change such as renal vein thrombosis. In diabetes and minimal change disease, there is usually little to find on microscopy in spite of heavy proteinuria.

The sediment in nephritic syndrome Patients with diseases such as lupus nephritis, immunoglobulin A  nephropathy, and post-infectious glomerulonephritis usually have nephritic sediment characterized by dysmorphic RBCs including acanthocytes and RBC casts. RBC casts are suggestive of glomerular inflammation but may be seen in non-inflammatory conditions such as malignant hypertension (Moriggi et  al., 1995), embolic renal infarction (Blakely et  al., 1994), and pre-eclampsia (Leduc et  al., 1991). Changes in the urine sediment including the number of RBCs and the number and character of casts are sometimes used as a marker of disease activity. WBC casts, RTEs, and leucocytes are common, especially in necrotizing vasculitis. The number of RBC casts has been shown to correlate with disease activity in granulomatosis with polyangiitis (formerly called Wegener’s granulomatosis) (Fujita et al., 1998).

Urinary findings in acute tubular necrosis

Fig. 6.7  An erythrocyte cast. Inset: a haemoglobin cast.

ATN is generally associated with granular casts often referred to as ‘muddy brown casts’ thought to result from degraded renal tubular cells. RTE cells and casts are often present (Schentag et al., 1979; Mandal et al., 1985; Marcussen et al., 1995). However, the urine sediment may be normal in ATN. A  scoring system based on the number of casts in the urine has been proposed as a marker of ATN severity but is not in standard use (Chawla et  al., 2008). Other urinary findings can suggest the aetiology of ATN such as uric acid crystals (urate nephropathy) and oxalate crystals (ethylene glycol poisoning) and drug crystals such as those formed by aciclovir (Fogazzi et  al., 1998). Myoglobin casts are seen in rhabdomyolysis, while bilirubin casts may be seen in renal failure associated with hyperbilirubinaemia (van Slambrouck et al., 2013).

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Urinary tract infection Common findings include foul odour and cloudy urine. Bacteria, leucocytes, isomorphic erythrocytes, superficial transitional cells, and leucocyte casts may be present (Fogazzi et al., 1998).

References Birch, D. F. and Fairley, K. F. (1979). Haematuria: glomerular or non-glomerular? Lancet, 2, 845–6. Blakely, P., Cosby, R. L., and McDonald, B. R. (1994). Nephritic urinary sediment in embolic renal infarction. Clin Nephrol, 42, 401–3. Brigden, M. L., Edgell, D., McPherson, M., et al. (1992). High incidence of significant urinary ascorbic acid concentrations in a west coast population—implications for routine urinalysis. Clin Chem, 38, 426–31. Chawla, L. S., Dommu, A., Berger, A., et al. (2008). Urinary sediment cast scoring index for acute kidney injury: a pilot study. Nephron Clin Pract, 110, c145–50. Eiser, A. R., Katz, S. M., and Swartz, C. (1979). Clinically occult diffuse proliferative lupus nephritis. An age-related phenomenon. Arch Intern Med, 139, 1022–5. European Confederation of Laboratory Medicine (2000). European urinalysis guidelines. Scand J Clin Lab Invest Suppl, 231, 1–86. Fairley, J. K., Owen, J. E., and Birch, D. F. (1983). Protein composition of urinary casts from healthy subjects and patients with glomerulonephritis. Br Med J (Clin Res Ed), 287, 1838–40. Fairley, K. F. and Birch, D. F. (1982). Hematuria: a simple method for identifying glomerular bleeding. Kidney Int, 21, 105–8. Fairley, K. F. and Birch, D. F. (1993). Microscopic urinalysis in glomerulonephritis. Kidney Int Suppl, 42, S9–12. Favaro, S., Bonfante, L., D’Angelo, A., et al. (1997). Is the red cell morphology really useful to detect the source of hematuria? Am J Nephrol, 17, 172–5. Fogazzi, G. B. (1996). Crystalluria: a neglected aspect of urinary sediment analysis. Nephrol Dial Transplant, 11, 379–87. Fogazzi, G. B. and Cameron, J.S. (1996). Urinary microscopy from the seventeenth century to the present day. Kidney Int, 50, 1058–68. Fogazzi, G. B., Carboni, N., Pruneri, G., et al. (1995). The cells of the deep layers of the urothelium in the urine sediment: an overlooked marker of severe diseases of the excretory urinary system. Nephrol Dial Transplant, 10, 1918–19. Fogazzi, G. B., Edefonti, A., Garigali, G., et al. (2008b). Urine erythrocyte morphology in patients with microscopic haematuria caused by a glomerulopathy. Pediatr Nephrol, 23, 1093–100. Fogazzi, G. B., Ferrari, B., Garigali, G., et al. (2012). Urinary sediment findings in acute interstitial nephritis. Am J Kidney Dis, 60, 330–2. Fogazzi, G. B. and Grignani, S. (1998). Urine microscopic analysis—an art abandoned by nephrologists? Nephrol Dial Transplant, 13, 2485–7. Fogazzi, G. B. and Grignani, S., and Colucci, P. (1998). Urinary microscopy as seen by nephrologists. Clin Chem Lab Med, 36, 919–24. Fogazzi, G. B. and Leong, S. O. (1996). The erythrocyte cast. Nephrol Dial Transplant, 11, 1649–52. Fogazzi, G. B. and Passerini, P. (1994). Urinary sediment—its use in recognizing complications of the nephrotic syndrome. Nephrol Dial Transplant, 9, 70–1. Fogazzi, G. B., Verdesca, S., and Garigali, G. (2008a). Urinalysis: core curriculum 2008. Am J Kidney Dis, 51, 1052–67. Fujita, T., Ohi, H., Endo, M., et al. (1998). Level of red blood cells in the urinary sediment reflects the degree of renal activity in Wegener’s granulomatosis. Clin Nephrol, 50, 284–8. Gai, M., Motta, D., Giunti, S., et al. (2007). Urinalysis: do not forget this type of cell in renal transplantation. J Nephrol, 20, 94–8. Goorno, W., Ashworth, C. T., and Carter, N.W. (1967). Acute glomerulonephritis with absence of abnormal urinary findings. Diagnosis by light and electron microscopy. Ann Intern Med, 66, 345–53.

Gyory, A. Z., Hadfield, C., and Lauer, C. S. (1984). Value of urine microscopy in predicting histological changes in the kidney: double blind comparison. Br Med J (Clin Res Ed), 288, 819–22. Hebert, L. A., Dillon, J. J., Middendorf, D. F., et al. (1995). Relationship between appearance of urinary red blood cell/white blood cell casts and the onset of renal relapse in systemic lupus erythematosus. Am J Kidney Dis, 26, 432–8. Heine, G. H., Sester, U., Girndt, M., et al. (2004). Acanthocytes in the urine: useful tool to differentiate diabetic nephropathy from glomerulonephritis? Diabetes Care, 27, 190–4. Hockberger, R. S., Schwartz, B., and Connor, J. (1987). Hematuria induced by urethral catheterization. Ann Emerg Med, 16, 550–2. Hoyer, J. R. and Seiler, M. W. (1979). Pathophysiology of Tamm-Horsfall protein. Kidney Int, 16, 279–89. Imhof, P. R., Hushak, J., Schumann, G., et al. (1972). Excretion of urinary casts after the administration of diuretics. Br Med J, 2, 199–202. Kitamoto, Y., Tomita, M., Akamine, M., et al. (1993). Differentiation of hematuria using a uniquely shaped red cell. Nephron, 64, 32–6. Kohler, H. and Wandel, E. (1993). Acanthocyturia detects glomerular bleeding. Nephrol Dial Transplant, 8, 879. Kohler, H., Wandel, E., and Brunck, B. (1991). Acanthocyturia—a characteristic marker for glomerular bleeding. Kidney Int, 40, 115–20. Kouri, T. T., Gant, V. A., Fogazzi, G. B., et al. (2000). Towards European urinalysis guidelines. Introduction of a project under European Confederation of Laboratory Medicine. Clin Chim Acta, 297, 305–11. Kubota, H., Yamabe, H., Ozawa, K., et al. (1988). Mechanism of urinary erythrocyte deformity in patients with glomerular disease. Nephron, 48, 338–9. Leduc, L., Lederer, E., Lee, W., et al. (1991). Urinary sediment changes in severe preeclampsia. Obstet Gynecol, 77, 186–9. Lettgen, B. and Wohlmuth, A. (1995). Validity of G1-cells in the differentiation between glomerular and non-glomerular haematuria in children. Pediatr Nephrol, 9, 435–7. Lindner, L. E., Jones, R. N., and Haber, M. H. (1980). A specific urinary cast in acute pyelonephritis. Am J Clin Pathol, 73, 809–11. Mandal, A. K., Sklar, A. H., and Hudson, J. B. (1985). Transmission electron microscopy of urinary sediment in human acute renal failure. Kidney Int, 28, 58–63. Marcussen, N., Schumann, J., Campbell, P., et al. (1995). Cytodiagnostic urinalysis is very useful in the differential diagnosis of acute renal failure and can predict the severity. Ren Fail, 17, 721–9. Martinez-Giron, R. (2008). Crystal-like structure in urine sediment. Diagn Cytopathol, 36, 252. Moriggi, M., Vendramin, G., Borghi, M., et al. (1995). Nephritic urinary sediment: not only in proliferative glomerulonephritis but also in malignant hypertension. Nephron, 70, 131. Mutter, W. P. and Brown, R. S. (2011). Point-of-care photomicroscopy of urine. N Engl J Med, 364, 1880–1. Nolan, C. R., 3rd, Anger, M. S., and Kelleher, S. P. (1986). Eosinophiluria—a new method of detection and definition of the clinical spectrum. N Engl J Med, 315, 1516–19. Nolan, C. R., 3rd and Kelleher, S. P. (1988). Eosinophiluria. Clin Lab Med, 8, 555–65. Perazella, M. A. (1999). Crystal-induced acute renal failure. Am J Med, 106, 459–65. Perazella, M. A. (2003). Drug-induced renal failure: update on new medications and unique mechanisms of nephrotoxicity. Am J Med Sci, 325, 349–62. Pugia, M. J., Lott, J. A., Profitt, J. A., et al. (1999). High-sensitivity dye binding assay for albumin in urine. J Clin Lab Anal, 13, 180–7. Rutecki, G. J., Goldsmith, C., and Schreiner, G. E. (1971). Characterization of proteins in urinary casts. Fluorescent-antibody identification of Tamm-Horsfall mucoprotein in matrix and serum proteins in granules. N Engl J Med, 284, 1049–52.

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Schentag, J. J., Gengo, F. M., Plaut, M. E., et al. (1979). Urinary casts as an indicator of renal tubular damage in patients receiving aminoglycosides. Antimicrob Agents Chemother, 16, 468–74. Schramek, P., Schuster, F. X., Georgopoulos, M., et al. (1989). Value of urinary erythrocyte morphology in assessment of symptomless microhaematuria. Lancet, 2, 1316–19. Schuetz, E., Schaefer, R. M., Heidbreder, E., et al. (1985). Effect of diuresis on urinary erythrocyte morphology in glomerulonephritis. Klin Wochenschr, 63, 575–7. Siemens (2010). Multistix and Labstix Reagent Strips Product Guide. Tarytown, NY: Siemens Healthcare Diagnostics. Szwed, J. J. and Schaust, C. (1982). The importance of microscopic examination of the urinary sediment. Am J Med Technol, 48, 141–3. Thal, S. M., DeBellis, C. C., Iverson, S. A., et al. (1986). Comparison of dysmorphic erythrocytes with other urinary sediment parameters of renal bleeding. Am J Clin Pathol, 86, 784–7.

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Ting, S. M., Ching, I., Nair, H., et al. (2009). Early and late presentations of ethylene glycol poisoning. Am J Kidney Dis, 53, 1091–7. Tsai, J. J., Yeun, J. Y., Kumar, V. A., et al. (2005). Comparison and interpretation of urinalysis performed by a nephrologist versus a hospital-based clinical laboratory. Am J Kidney Dis, 46, 820–9. Van Slambrouck, C. M., Salem, F., Meehan, S. M., et al. (2013). Bile cast nephropathy is a common pathologic finding for kidney injury associated with severe liver dysfunction. Kidney Int, 84, 192–7. Wald, R., Bell, C. M., Nisenbaum, R., et al. (2009). Interobserver reliability of urine sediment interpretation. Clin J Am Soc Nephrol, 4, 567–71. Wandel, E. and Kohler, H. (1998). Acanthocytes in urinary sediment—a pathognomonic marker? Nephrol Dial Transplant, 13, 206–7.

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Assessment of renal function Marijn Speeckaert and Joris Delanghe Estimation of glomerular filtration rate Exogenous compounds Assessment of kidney function in the clinical laboratory is usually based on the measurement of the clearance of various substances by the kidneys. The renal clearance of a substance is defined as ‘the volume of plasma from which the substance is completely cleared by the kidneys per unit time’. The clearance of a substance S is given by: Cs = (Us × V)/Ps. In most circumstances, an estimation of the glomerular filtration rate (eGFR) based on formulas incorporating serum creatinine is sufficient for clinical decision-making. However, for patients in whom GFR estimates based on serum creatinine are likely to be inaccurate or may have adverse consequences, measured GFR (mGFR) may be helpful. In the following sections we describe clinical situations in general medicine and nephrology where measurement of GFR should be considered (Stevens and Levey, 2009a).

Exogenous markers GFR can be measured as the clearance of exogenous or endogenous filtration markers. Non-radioactive compounds used as exogenous markers to measure GFR include iohexol, inulin, iodoacetate, and diethylenetriaminepentaacetic acid (DTPA). Clearances based on these exogenous markers are very accurate but are expensive and rather impractical and therefore are mainly restricted to research use. (See Table 7.1.)

Inulin The reference method to determine GFR is the urinary clearance of the fructose polymer inulin during a continuous intravenous infusion. Alternatively, the plasma clearance of inulin can be determined, which does not require urine collection (Ferguson et al., 1950). 51Cr-EDTA

The chromium-51 ethylenediaminetetraacetic acid (51Cr-EDTA) marker is not commercially available in the United States, but there is an extensive European experience with this marker. The urinary clearance of 51Cr-EDTA consistently underestimates inulin clearance by 5–15% in most, although not all studies, suggesting a degree of tubular reabsorption (BrochnerMortensen, 1978).

DTPA DTPA, an analogue of EDTA is usually labelled with technetium-99m (99mTc). Advantages include a short half-life (6 hours), minimizing

radiation exposure, and a high counting efficiency of 99mTc. DTPA is thought to be freely filtered at the glomerulus, with minimal tubular reabsorption, but it may undergo extrarenal elimination. Its major limitation is the potential of dissociation of 99mTc from DTPA and binding to plasma proteins, leading to underestimation of the GFR. The extent of dissociation is not predictable, leading to imprecision and bias. In addition, chelating kits and Tc generators are not standardized, making comparisons of mGFR with DTPA 99mTc among different institutions difficult (Stevens and Levey, 2009b). The magnetic resonance imaging contrast agents, gadolinium-DTPA or gadolinium-tetraazacyclododecane tetraacetic acid (gadolinium-DOTA), have recently been introduced as novel exogenous filtration markers because of their wide availability and their low rate of allergic reactions (Schuhmann-Giampieri and Krestin, 1991). In addition, a highly sensitive, novel, immunoassay technique is easily performed in most clinical laboratories and needs very low doses (1/40th of dose used for contrast) (BioPAL, Inc., 2007). The problem of systemic nephrogenic fibrosis has diminished enthusiasm for using this agent in subjects with significantly reduced GFRs, but because of the very low dose needed this complication is very unlikely to occur (Swaminathan and Shah, 2007).

Iohexol and iothalamate Iohexol and iothalamate are radiographic contrast agents and can be used as exogenous GFR markers comparable to inulin and 51Cr-EDTA (Schwartz and Furth, 2007). Concern about radiation led to the use of the non-radioactive radiographic contrast agent, iohexol (Omnipaque®) (Schwartz et al., 2006). Following iohexol administration, its content in collected plasma and urine samples is measured by high-performance liquid chromatography (HPLC). A  stable molecule such as iohexol would enable quality assessment schemes to be developed for GFR analysis. Iohexol accurately measures GFR with a four-point plasma disappearance curve (10, 30, 120, and 300 minutes) or, in most cases, with a two-point disappearance time (120 and 300 minutes) (Brandt et al., 2006). Iothalamate is commonly administered as a radioactive iodine label, but can also be administered in its non-radioactive form and is measured with HPLC (Notghi et  al., 1986). In the radioactive form, it is most commonly administered as a bolus subcutaneous injection. Iodine-125 (125I)-iothalamate has been widely adopted for measurement of GFR. To block thyroidal uptake, cold iodine is administered at the time of 125I-iothalmate administration, thus precluding its use in people with known allergies to iodine. Most studies comparing urinary clearance of iothalamate to inulin

chapter 7 

Table 7.1  Strengths and limitations of some exogenous GFR markers (Stevens and Levey, 2009a) Marker

Advantages

Disadvantages

Inulin

Gold standard

Expensive

Cr-EDTA

Widely available in Europe

Probable tubular reabsorption

99Tc-DTPA

Widely available in the United States

Iohexol

Not radioactive

Possible tubular reabsorption or protein binding Expensive Not used in patients with allergies to iodine

Iothalamate

Inexpensive

Probable tubular secretion

131I-hippuran

30% protein binding

showed a small positive bias, probably because of tubular secretion of iothalamate.

Endogenous markers Creatinine and creatinine standardization Creatinine is by far the most commonly used biochemical marker (Fig. 7.1) of renal function. Creatinine originates from the non-enzymatic hydrolysis of creatine and phosphocreatine, two substances found almost exclusively in the muscle (Wyss and Kaddurah-Daouk, 2000). This reaction occurs at a constant rate. Creatinine is a small molecule (molecular mass of 113 Da) that does not bind plasma proteins and is mainly eliminated by the kidney in

assessment of renal function

individuals with normal renal function. Creatinine is freely filtered by the glomerulus and is not reabsorbed throughout the tubules. Therefore its clearance can be measured as an indicator of GFR. Its production rate is decreased in patients with hepatic diseases (Cocchetto et  al., 1983). Serum or plasma creatinine concentrations do not only depend on the GFR, but also on muscle mass and on dietary protein intake (Crim et al., 1975; Heymsfield et al., 1983; Lew and Bosch, 1991). Tubular secretion, intestinal exchange, and the method and analytical standards used are also important variables. In cases of severe renal dysfunction, the GFR based on creatinine clearance rate will be ‘overestimated’, because the active tubular secretion of creatinine will account for a larger fraction of the total creatinine cleared. Ketoacids, cimetidine, and trimethoprim reduce creatinine tubular secretion and therefore increase the accuracy of the GFR estimate, particularly in severe renal dysfunction. Creatinine is rather unstable and should be measured without delay. Serum or plasma creatinine concentrations only increase beyond the reference values when kidney function is reduced by > 50%. Below this threshold, there is the so-called creatinine blind range, in which a measured creatinine clearance may provide more diagnostic information. In order to convert from conventional to SI units, the following conversion factor can be used: 1 mg/dL corresponds to 88.5 µmol/L. Creatinine can be determined using various modifications of the Jaffe principle (alkaline picrate reaction) (Delanghe and Speeckaert, 2011), by enzymatic dry chemistry (Toffaletti et al., 1983)  or by enzymatic colorimetric methods. The most commonly used assay is based on the so-called Jaffe reaction (Jaffe, 1886). In his landmark paper, Jaffe discussed that the alkaline picrate reaction could also occur, be it to a much lesser extent with a number of organic compounds (e.g. acetone, glucose). (These compounds were later designated as pseudo-chromogens and are a source of unspecificity in the Jaffe reaction). In the earliest

Creatine + creatine phosphate pools (± 120 g)

c

c c

Anabolism

Serum creatinine

Catabolism Creatinine pool Glomerular filtration

Extrarenal creainine Tubular secretion Urinary creatinine load: ± 1700 mg/24 h × 1.73 m2

Fig. 7.1 

45

46

Section 1  

assessment of the patient with renal disease

methods, serum creatinine was assayed by reactions based on alkaline picrate after deproteinization or dialysis, which eliminated the pseudo-chromogen effect of proteins (Levey, 1990). Today, however, analysers use untreated serum or plasma, making creatinine assays using alkaline picrate reaction prone to the so-called protein error. On average, this effect produces a positive difference of 27  µmol/L creatinine in Jaffe assays (Wuyts et  al., 2003). Because urine contains relatively little or no protein, the protein error affects only creatinine determinations in serum or plasma resulting in underestimation of the creatinine clearance when creatinine methods affected by protein error are used. Since Jaffe only observed a complexation between picric acid and creatinine in alkaline environment and never described an analytical method, variation amongst Jaffe methodological ‘recipes’ is broad. Notwithstanding stricter regulations, between-laboratory variation of Jaffe-based methods has not decreased over the last decade, despite technical progress in laboratory automation (Delanghe et al., 2008). Despite the known limitations, methods based on the Jaffe reaction are still extensively used for measuring serum creatinine. Haemolysed sera induce falsely increased values and the Jaffe reaction is negatively interfered by bilirubin. The ‘true’ creatinine value can be determined enzymatically. So-called compensated creatinine methods allow similar results as obtained by enzymatic creatinine assays. These values are about 20% lower compared with the non-compensated creatinine values (Junge et al., 2004). Standardization of serum creatinine measurements is very important because of the central role of this biomarker for the calculation of creatinine clearance, and the use of creatinine values for estimation of GFR (Levey, 1990). Estimations of GFR can be obtained using equations that empirically combine all of the average effects from biological factors that affect serum creatinine concentrations in serum other than GFR (Stevens and Levey, 2004). Assays not calibrated in agreement with the kinetic alkaline picrate method, used in the eGFR formula, introduce a source of error into GFR estimates. Implementing traceability of serum creatinine assays to gas chromatography-isotope dilution mass spectrometry (GC-IDMS) or liquid chromatography-isotope dilution mass spectrometry (LC-IDMS) leads to changes in the clinical decision-making criteria currently used for serum creatinine concentrations and creatinine clearance since calibration to an IDMS reference produces a lowering of serum creatinine values by 10–30% for most methods. (See Tables 7.2 and 7.3.)

Creatinine in urine For measuring creatinine in urine, a timed collection is necessary for measuring creatinine clearance. In most cases, the preferred method is the 24-hour collected urine, compensating for the variation in creatinine excretion during the day. Reference ranges are 0.87–2.41 g/24 hours (7.7–21.3 mmol/24 hours) for men and 0.67–1.59 g/24 hours (5.9–14.0  mmol/24 hours) for women. In contrast to the serum values, reference values for urinary creatinine are relatively independent from the method used.

Creatinine: recent analytical issues The National Kidney Disease Education Program (NKDEP), the College of American Pathologists (CAP), and the National Institute for Standard and Technology (NIST) have collaborated

to prepare a serum-creatinine reference material (NIST 967) with demonstrated commutability with native clinical specimens in routine methods. These materials are value-assigned with the GC-IDMS and LC-IDMS reference measurement procedures (Dodder et al., 2007). Nearly all clinical laboratory methods are now expected to have calibration traceable to an IDMS reference measurement procedure. The MDRD Study equation used for estimating GFR in adults has been updated to use coefficients appropriate for such methods (Levey et al., 2007). Routine reporting of eGFR for adults based on an IDMS-traceable creatinine has been proposed as clinical standard for patient care (Kallner and Khatami 2008; Narva, 2009). Variability in creatinine results may, however, contribute to substantial uncertainty in estimating GFR (Delanghe et al., 2008). Implementing traceability of serum creatinine assays to GC- or LC-IDMS has led to changes in the clinical decision-making criteria currently used for serum creatinine concentrations and creatinine clearance (Delanghe et al., 2011). Completeness of a 24-hour urine collection can be estimated by determining the excreted amount of creatinine. In order to obtain reliable test results, precise instructions dealing with the precise scenario of the timed urine collection should be provided to the patient and the nursing staff. When creatinine clearance is measured in patients who have been administered 3  × 800 mg cimetidine (a blocker of tubular secretion of creatinine) the day preceding the blood sampling, the effect of tubular secretion can be corrected. The cimetidine protocol, with creatinine clearance derived from a 2-hour urine collection, allows estimating of glomerular filtration rate in a clinical setting (Hellerstein et al., 1998). However, it cannot be used on a wide scale.

Creatinine clearance A classical creatinine clearance determination requires a serum or plasma specimen and a timed urine collection. The clearance is considered to be the amount of liquid filtered out of the blood. It is based on a ‘normal’ adult body surface of 1.73 m2. A small quantity of creatinine that appears in the urine (7–10%) is due to tubular secretion. By consequence, creatinine clearance usually exceeds inulin GFR by a factor of 1.1–1.2 at clearances above 80–90 mL/ min. The adult reference range (values standardized to 1.73 m 2 body surface) is 66–143 mL/min for the enzymatic method and 71–151 mL/min for Jaffe-based compensated tests. From the age of 40 onwards, creatinine clearance drops by about 8.5 mL/min per decade. From 2009 onward, most commercially available creatinine assays have been standardized to the reference method IDMS. This standardization change led to a worldwide reduction of serum creatinine values (Ceriotti et al., 2008). In routine clinical practice, 24-hour urine collections are an important source of error which impairs reliable calculation of the creatinine clearance. Consequently, practical formulas have been developed, which allow an estimation of the creatinine clearance without timed urine collections. The classical creatinine clearance is calculated as follows: creatinine clearance = urine creatinine ( mg / dL ) × urine volume ( mL / 24 h ) plasma creatinine ( mg / dL ) × 1440



chapter 7 

assessment of renal function

Table 7.2  Paediatric reference intervals for creatinine concentrations in serum, calculated with a non-parametric (NP) method and using fractional polynomials (FP) (Schlebusch et al., 2002) NP percentile Age group

N

2.5th

97.5th

Cord sera

51

46

86

Preterm neonates 0–21 days

58

28

87

Term neonates 0–14 days

69

27

2 months to < 1 year

41

1 to < 3 years 3 to < 5 years

FP percentile Midpoint age

2.5th

97.5th

—

—

At birth

29

90

81

7 days

22

73

14

34

7 months

11

34

45

15

31

2 years

15

30

41

23

37

4 years

21

34

5 to < 7 years

43

25

42

6 years

26

40

7 to < 9 years

46

30

48

8 years

31

46

9 to < 11 years

47

28

57

10 years

35

53

11 to < 13 years

42

37

63

12 years

38

59

13 to < 15 years

38

40

72

14 years

41

65

Cord sera

51

0.52

0.97

—

—

Preterm neonates 0–21 days

58

0.32

0.98

At birth

0.33

1.01

Term neonates 0–14 days

69

0.31

0.92

7 days

0.26

0.81

2 months to < 1 year

41

0.16

0.39

7 months

0.15

0.37

1 to < 3 years

45

0.17

0.35

2 years

0.18

0.33

3 to < 5 years

41

0.26

0.42

4 years

0.24

0.39

5 to < 7 years

43

0.29

0.48

6 years

0.29

0.46

7 to < 9 years

46

0.34

0.55

8 years

0.34

0.53

9 to < 11 years

47

0.32

0.64

10 years

0.39

0.60

11 to < 13 years

42

0.42

0.71

12 years

0.43

0.67

13 to < 15 years

38

0.46

0.81

14 years

0.47

0.73

µmol/L

mg/dL

Table 7.3  Reference intervals for creatinine concentrations in serum in adults (ages 18–74 years) (Junge et al., 2004) Percentile (90% CI) N Men Women

120 120

2.5th

97.5th

µmol/L

64 (63–66)

104 (99–107)

mg/dL

0.72 (0.71–0.75)

1.18 (1.12–1.21)

µmol/L

49 (46–55)

90 (83–103)

mg/dL

0.55 (0.52–0.62)

1.02 (0.94–1.17)

Conversion factor: 1 mg/dL = 88.5 µmol/L.

creatinine clearance =

(140 − age) × body weight ( kg ) 72 × serum creatinine ( mg / dL )

In women: creatinine clearance =

0.85 × (140 − age ) × body weight ( kg ) 72 × serum creatinine ( mg / dL )

In children, the Schwartz formula is frequently used: creatinine clearance =

0.55 × length ( cm )

serum creatinine ( mg / dL )

In term newborns and infants during the first year of life: In adults, the Cockcroft and Gault formula is used for the estimation of the creatinine clearance, which only requires the serum creatinine value, patient’s age, and body weight:

creatinine clearance =

0.45 × length ( cm )

serum creatinine ( mg / dL )

47

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The mentioned coefficients of the Schwartz formula must not be used in combination with enzymatic or so-called compensated creatinine assays, because this will lead to a serious overestimation of the creatinine clearance. For enzymatic IDMS calibrated creatinine assays a coefficient value of 0.42 instead of 0.55 is used (Schwartz et al., 2009). In contrast, enzymatic and compensated Jaffe-methods are well adapted for calculations with the Cockcroft and Gault formula. The coefficients of the Cockcroft and Gault formula do not match non-compensated creatinine determinations.

Calculation of GFR Only when relative GFR loss exceeds 50%, will serum creatinine values exceed the upper reference range (so-called creatinine-blind range). The serum creatinine concentration also depends on age, gender, and muscle mass. The best parameter for the determination of renal function is the GFR. International scientific societies now recommend the application of a new formula for the estimation of GFR. This formula has been validated in the framework of the Modification of Diet in Renal Diseases Study Group Study (MDRD Study) (Levey et al., 1993). The original MDRD Study equation was based on six variables: age, sex, ethnicity, and serum levels of creatinine, urea, and albumin (Levey et al., 1999). Subsequently, a four-variable equation consisting of age, sex, ethnicity, and serum creatinine levels was proposed to simplify clinical use (Levey et al., 2000). This equation is now widely accepted, and many clinical laboratories are using it to report GFR estimates. Extensive evaluation of the MDRD Study equation shows good performance in populations with lower levels of GFR but variable performance in those with higher levels. Variability among clinical laboratories in calibration of serum creatinine assays introduces error in GFR estimates and may account in part for the poorer performance in this range. The MDRD Study equation has now been re-expressed for use with standardized serum creatinine assays, allowing GFR estimates to be reported by using standardized serum creatinine and overcoming this limitation to the current use of GFR estimating equations (Levey et al., 2009):

(

)

GFR mL / min / 1.73 m 2 = 175 (serum creatinine)

−1.154

×

(age) × (0.742 in women) × (1.21 in African Americans) −0.2 033



The calculation (using coefficients adapted to IDMS standardization) may only be used in combination with corresponding IDMS-traceable methods. The MDRD Study equation is superior to the Cockcroft and Gault formulation in predicting kidney function in most people (Poggio et al., 2005). In 2009, a new formula, named the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) was also introduced (Levy et al., 2009), which provides a more accurate GFR estimation for patients with GFR values between 60 and 90 mL/ min. In this equation, the values of the constants of a, b, and c vary on the basis of race, sex, and serum creatinine. GFR = a × (serum creatinine / b ) × ( 0.993) c

age

The variable a takes on the following values on the basis of race and sex: for African Americans: women = 166; men = 163. For white/

other, the coefficients were respectively 144 for women and 141 for men. The variable b takes on the following values on the basis of sex: women = 0.7 and men = 0.9. The variable c takes on the following values on the basis of sex and creatinine measurement: for women: if serum creatinine ≤ 0.7 mg/dL = −0.329; if serum creatinine > 0.7 mg/dL = −1.209; for men: if serum creatinine ≤ 0.9 mg/ dL = −0.411; if serum creatinine > 0.9 mg/dL = −1.209. The investigators compared the abilities to identify patients with various degrees of kidney disease of the CKD-EPI formula with the MDRD formula. Although both formulas performed similarly well, the cases in which subjects were misclassified in terms of category of kidney disease, the CKD-EPI formula was more often correct (P < 0.001). In the separate subset of patients designated to validate the new formula, the CKD-EPI formula demonstrated less bias and improved precision and greater accuracy than the MDRD formula (P < 0.001), particularly for those patients with an estimated GFR > 60 mL/min. The median difference between measured and estimated GFR was 2.5 mL/min for the CKD-EPI formula and 5.5 mL/ min for the MDRD formula (P < 0.001). Because of the reduced error of overestimation of kidney function, the CKD-EPI equation yielded a lower estimated prevalence of kidney disease in the National Health and Nutrition Examination Survey (NHANES) dataset (You et al., 2011). Where the MDRD equation estimated the prevalence of kidney disease at 13.1%, the CKD-EPI formula estimated it slightly lower at a prevalence of 11.5%. This limited extent to which the equation overestimates GFR reflects the major advantage of this newer formula. The determination of the creatinine clearance is commonly used in routine diagnostics to estimate the GFR, when serum creatinine value is still within reference limits. The Cockcroft and Gault formula must not be used in patients with acute kidney insufficiency and/or an unstable renal function, very obese patients or in presence of pronounced oedema (overestimation of clearance). Multiple conditions are characterized by increased GFR and kidney weight such as diabetes, pregnancy, unilateral nephrectomy, and high protein intake. In these conditions, an increase of GFR due to increase of effective renal plasma flow is the primary event that over a period of days is followed by kidney growth (Bak et al., 2000). In a recent study, data from cross-sectional studies that compared two or more creatinine-based GFR estimating equations to a reference GFR measurement were analysed. Studies from North America, Europe, and Australia showed a better performance of the CKD-EPI equation at higher GFRs (approximately > 60 mL/ min per 1.73 m2). However at lower GFRs, the MDRD Study equation was superior. As the CKD-EPI and MDRD Study equations were developed in North American and European populations that mainly consisted of black people and white people, the equations had to be modified in Asian and African populations to improve their performance by adding or removing a ‘race/ethnicity’ coefficient. Those coefficients are not useful beyond the local population, possibly because of differences in GFR measurement methods or differences in populations in addition to race or ethnicity. In conclusion, neither the CKD-EPI nor the MDRD Study equation is optimal across all populations and GFR ranges. New equations should focus on other filtration markers instead of or in addition to creatinine (Earley et al., 2012).

GFR and drug dose calculation Estimation of the creatinine clearance is a key element in the calculation of the correct dose of many drugs which are

chapter 7 

characterized by a narrow therapeutic index and a renal elimination pathway (Nagao et al., 2005). The implementation of eGFR reporting and standardization of creatinine measurements has created some uncertainty and confusion concerning the assessment of kidney function for drug dosing adjustment (Narva, 2009). Pharmaceutical companies have used for many years the Cockcroft and Gault equation (Cockcroft and Gault, 1976) to estimate creatinine clearance as the basis for drug dose adjustments. There is no modified Cockcroft and Gault equation available for use with the IDMS-traceable creatinine results. Consequently, creatinine clearance estimated from the Cockcroft and Gault equation will be increased upon restandardization. The original paper by Cockcroft and Gault does not contain detailed information regarding the creatinine assay used. The Cockcroft and Gault formula estimates creatinine clearance, which is not synonymous for GFR, since creatinine clearance is partly influenced by a tubular secretion of the compound. On the other hand, MDRD and CKDP-EPI formulas provide an estimate for GFR. Because the MDRD Study equation is relatively new, eGFR data is not part of drug safety information or package inserts. There is a concern about dosing toxic drugs with narrow therapeutic indices (e.g. carboplatin). Some clinicians request back-calculation to a non-standardized creatinine which can then be used in the Cockcroft and Gault equation. Back-calculation appears to be unnecessary (Stevens et  al., 2009). Concordance for drug dosing recommendations based on measured GFR for cleared drugs cleared by the kidneys was best for MDRD (88%) compared to the Cockcroft and Gault equation adjusted for ideal body weight (CGIBW) (82%) and the unadjusted Cockcroft and Gault (85%). Of the three estimating methods, the Cockcroft and Gault equation was most likely to generate higher recommended drug dosages, and CGIBW was most likely to generate lower recommended drug dosages. The MDRD produced recommendations that were lower than Cockcroft and Gault in 9% of the study population and higher in 10% when the CGIBW was used. The use of the MDRD equation for drug dosing adjustment in adults is encouraged (Narva 2009). Implementation of the CKD-EPI formula (Levey et al., 2009) in place of MDRD does not have much clinical impact on this specific application since drug dose adjustment is rarely needed for patients with GFR > 60 mL/min. The National Kidney Disease Education Program (NKDEP) (National Kidney Disease Educational Program, 2011) has published an educational advisory on estimating kidney function for drug dosing purposes. The advisory encourages the use of MDRD or Cockcroft and Gault estimating equations and, when there is concern that estimated kidney function is not adequate for patient safety or when there is a distinct difference in recommended dose between the two methods, suggests consideration of measured creatinine clearance or assessment of GFR by the clearance of exogenous compounds. Special attention should be paid to the elderly. MDRD has only been validated for patients aged between 18 and 70 years. MDRD overestimates renal function as age increases. While the optimized Cockcroft and Gault equation underestimates renal function, this was of a smaller magnitude, consistent across age, and thus better suited for dose calculation in the elderly (Roberts et al., 2009). Therefore, it is important not to extend the use of MDRD to the age groups for which the formula has not been validated. The new CKD-EPI formula has been validated over a wider range of ages and seems to be more accurate, also in the elderly patients (Aucella et al., 2010; Michels et al., 2010).

assessment of renal function

GFR estimation in paediatrics Creatinine restandardization should be a major concern in paediatrics due to the lower reference ranges for serum creatinine in infants and children (Delanghe, 2008). For calculating GFR, this systematic positive bias has been greatly compensated by the overestimation attributable to tubular secretion of creatinine, which is relatively more important in children. Although some Jaffe manufacturers try to correct for protein error through the use of a fixed compensation factor for the protein content in adults (Wuyts et  al., 2003), this procedure overcorrects with paediatric samples because of children’s low serum protein concentrations. Children, particularly younger children, have lower albumin and immunoglobulin (IgG) plasma concentrations than adults. In consequence, use of restandardized Jaffe-type assays results in overcompensation when used in children or infants. This overcorrection produces inaccurate GFR estimates, especially in neonates and children. Compensating calibration in Jaffe assays to IDMS standards results in an underestimation of serum or plasma creatinine due to the lower reference values for total protein in younger children. The enzymatic methods manage to measure serum creatinine more correctly (Delanghe et al., 2008). The IFCC Scientific Division recommends that more specific creatinine methods be adopted (Panteghini and IFCC Scientific Division, 2008). In enzymatic creatinine methods, analytical non-specificity is largely eliminated. The lower enzymatic creatinine result (when the result has not been adjusted to Jaffe-like results) leads to a marked increase of creatinine clearance and GFR estimations because of the increased effect of tubular secretion on test results. The use of enzymatic assays emphasizes the relative proportion of tubular secretion of creatinine which makes serum or plasma creatinine less suitable as a GFR marker in paediatric medicine, despite the analytical improvement (Delanghe, 2008). For estimating GFR in children and infants, the Schwartz (Schwartz et al., 1976, 1984; Schwartz and Gauthier, 1985) and the Counahan–Barratt equations (Counahan et  al., 1976)  are used. Both provide GFR estimates based on a constant value multiplied by the child’s height divided by the serum creatinine concentration. The values (SI units) for the constant used in both equations differ considerably: k = 38 (Counahan et al., 1976) and k = 48.7 (Schwartz et al., 1984; Delanghe et al., 2008). Since these formulas were validated 35 years ago, reassessment of classical formulas for estimating creatinine clearance and GFR using modern creatinine assays is necessary. As a result of restandardization, the original Schwartz-formula overestimates GFR. In the revised equation (Schwartz et al., 2009), with height measured in cm, using the calculation of 0.413 × (height/serum creatinine) (conventional units) or 46.67  × (height/serum creatinine; µmol/L), provides a good approximation of the estimated GFR formula. A newly developed CKiD equation uses serum creatinine, urea, and cystatin C (Cys C), plus height and sex, to estimate GFR, yielding the equation:

(

)

GFR mL / min per 1.73 m2 = 39.1 height ( m ) / Scr ( mg / dL ) [30 / BUN mg / dL)  0.169 1.099  male

0.516

× 1.8 / Cys C ( mg / L )  height ( m ) / 1.4]0.188 . 

( )

0.294



The new equation gives a better agreement with measured GFR than the updated Schwartz equation. There is also the Lund–Malmö

49

50

Section 1  

assessment of the patient with renal disease

equation which was validated for both adults and children (Nyman et  al., 2008). These formulas are only valid in combination with enzymatic methods with calibration traceable to an IDMS reference procedure. However, as the majority of the manufacturers of in vitro diagnostics will prefer general solutions based on a recalibration setup at the upper reference limit of an adult population, there is a risk of overcompensation because of the differences in serum matrix. So far, the updated Schwartz formula is the only paediatric GFR-estimating equation with coefficients suitable for use with IDMS-traceable creatinine methods. The Counahan equation was found to have nearly the same performance as the updated Schwartz equation (Miller, 2009). Because of the difficulties in adapting creatinine assays to the new calibrators in a uniform way, Cys C offers a promising alternative for calculating GFR in children. Compared to serum creatinine, low-molecular-weight proteins have a better diagnostic sensitivity for detecting an impaired GFR in children (Keevil et al., 1998). Although some caveats must be taken into account for a correct interpretation of the results, (Manetti et al., 2005; Hari et al., 2007), the protein-based GFR calculations only require serum values. The progress in the standardization of these protein assays will enable the wide-scale use of these methods.

Epidemiological studies As a result of creatinine restandardization, measured and calculated creatinine clearance values will increase, and the corresponding reference interval will be different. In an attempt to reduce late referral and to improve the care of patients with chronic kidney disease (CKD), different organizations have issued guidelines on the timely referral of patients to a nephrologist (National Kidney Foundation, 2005). Most suggest referral of patients with a GFR < 60 mL/min/1.73 m2, and mandate referral if the GFR is < 30 mL/min/1.73 m2. KDIGO published updated guidelines in 2013 (Kidney Disease:  Improving Global Outcomes (KDIGO) CKD Work Group, 2013). Important differences in classifications were obtained when different correction formulae for creatinine (the basis for GFR calculation) were applied.

Urea Urea was the very first marker of renal dysfunction, and renal failure is still referred to as uraemia. Urea is a small molecule (molecular mass: 60 Da) produced from the metabolism of amino acids. Its daily production varies largely with protein intake and is also dependent on protein degradation in the intestines as observed during gastrointestinal bleeding (Cottini et al., 1973; Maroni et al., 1985; Chalasani et  al., 1997). In the United States, often BUN (blood urea expressed as nitrogen) is used as a parameter, which corresponds to the urea-N-value. Urea value (mg/dL) is obtained by multiplying the BUN value by 2.14. More than 90% of urea is excreted through the kidneys. Urea is filtered freely by the glomeruli; 40–60% of the filtrated urea moves passively out of the renal tubules a process depending on urine flow rate. When water reabsorption is increased in renal tubules, or when intravascular volume is depleted, increased urea reabsorption leads to high serum urea concentration. Protein intake strongly correlates with serum urea concentration. In catabolic states, the loss of endogenous protein nitrogen results in urea production. Rapid muscle breakdown can also lead to a temporary increase in the serum urea concentration.

Because of these effects of protein metabolism upon urea concentration, serum creatinine is a more specific marker for renal function. The BUN/creatinine ratio may be helpful in the discrimination between prerenal and postrenal azotaemia. The reference for the BUN/creatinine ratio is between 10:1 and 20:1. Lower values usually denote acute tubular necrosis, low protein intake, starvation, or severe liver disease. High ratios with normal creatinine levels may be observed when there is tissue breakdown, prerenal azotaemia, high protein intake, or after gastrointestinal haemorrhage. High ratios associated with elevated creatinine concentration are found in either postrenal obstruction or prerenal azotaemia. Because urea is the major nitrogen compound produced by protein catabolism it allows an estimation of the amount of catabolized protein (1g N ~ 6.25 g protein). The reference range for urea in serum or plasma is between 20 and 50 mg/dL (3.3–8.3 mmol/L). For converting conventional into SI units, the conversion factor:  1 mg/ dL = 0.1667 mmol/L can be used. Urea clearance is a poor indicator of GFR, as its production rate is dependent on several non-renal factors, including diet. In addition, the variable level of back-diffusion will influence both plasma and urine urea concentrations. Urine urea excretion allows a good estimate of the protein content in the diet: 1 g of protein corresponds with ± 300 mg (50 mmol) of urea synthesis. The reference range for a daily urine urea excretion is between 1500 and 2600 mg/dL (250–433  mmol/L). Ammonia contamination (e.g. in urinary tract infection caused by Proteus spp.) leads to falsely increased values.

Cystatin C Serum concentrations of low-molecular weight marker proteins are primarily determined by GFR. An ideal marker has to have a constant production rate and should not vary in its concentration in situations with an acute-phase reaction. Cys C seems to share these ‘ideal’ properties. Cys C is a cysteine-proteinase inhibitor with a low molecular mass (13,300 Da), and is produced at a constant rate by all nucleated cells. Its small size and high pI (9.2) enable it to be freely filtered at the glomerulus. In normal conditions, serum Cys C is almost completely filtered by the renal glomerulus and largely catabolized by proximal tubular cells. Since serum Cys C concentration is closely correlated with the GFR, its determination has been introduced as a marker of renal function (Delanghe 2008). The Cys C concentration in serum is therefore determined by the GFR and in contrast to creatinine, Cys C concentrations are not affected by muscle mass and these values can also be used in children (Table 7.4). Unlike creatinine, serum Cys C reflects renal function independent of age, gender, and body composition. In the routine clinical laboratory, Cys C can be assayed using immunonephelometric or immunoturbidimetric tests. As the interindividual variation of serum Cys C-concentration is lower than that of creatinine, this protein marker has a higher diagnostic sensitivity and specificity in particular in patients with a moderate reduction of the GFR (between 40 and 80 mL/min). Cys C also offers a promising alternative for calculating GFR in children, as only a determination in serum or plasma is required. Especially in the so-called blind range of creatinine or in patients with low muscular mass, Cys C proves to be a superior marker compared with serum creatinine. Formulas have been developed

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Beta-trace protein

Table 7.4  Cystatin C reference values Age group

(mg/L)

< 1 month

1.37–1.89

1–12 months

0.73–1.17

Adult men

0.50–0.96

Adult women

0.57–0.96

allowing reliable estimation of GFR based on Cys C (Bökenkamp et  al., 1998). GFR-estimating equations based on Cys C are currently limited to the laboratory method that was used to derive the equation. Thyroid dysfunction affects serum Cys C concentration, possibly influencing the production rate of the protein (Manetti et al., 2005). Cys C is upregulated in certain tumours. Also pharmacological factors (e.g. glucocorticoid treatment) may influence the concentrations of serum Cys C (Delanghe 2008). In 2010, the IFCC Working Group for the Standardisation of Cystatin C (WG-SCC), in collaboration with the Institute for Reference Materials and Measurements (IRMM), announced the availability of the new certified reference material ERMDA471/IFCC. The new certified reference material will allow a global standardization of this analyte (Grubb et al., 2010). Because of its low individual variability, Cys C has fewer inherent limitations as a screening test for detecting a declining GFR than serum creatinine. However, serum creatinine is probably still the better assay for following sequential changes in an individual with confirmed renal disease (Keevil et al., 1998). Serum creatinine concentrations are lower in malnourished children and lead to overestimation of GFR, while Cys C levels are unaffected. Agreement between the Schwartz formula and gold standard GFR shows considerable bias, with a mean difference of 10.8% and a trend towards overestimation of the GFR by the Schwartz formula with lower GFRs. Cys C-based GFR estimates show significantly less bias and serves as a better estimate for GFR in children. Recently, combined GFR estimates based on both creatinine and Cys C have been presented (Stevens and Levey, 2009b).

Beta-trace protein (BTP) is a low-molecular-weight glycoprotein with a molecular weight of 23–29 kDa, depending on the degree of glycosylation. BTP (prostaglandin D synthase) belongs to the lipocalin protein family. It transports small lipophilic substances. BTP can be measured using immunonephelometry. Recently BTP has been introduced for the measurement of kidney function in the creatinine-blind range. BTP, like Cys C, is independent of age and gender. The upper reference limits of serum BTP and Cys C are 1.01 and 1.20 mg/L, respectively. Above 2 years, the reference range of BTP is constant at 0.43–1.04 mg/L (Filler et  al., 2002). These data confirm the potential of BTP as an endogenous GFR marker. International standardization of BTP is still lacking.

Beta-2 microglobulin Beta-2 microglobulin (B2M; 11.3 kDa) has been advocated as a predictor of GFR (Tables 7.5 and 7.6), but its serum concentration can increase as an acute-phase reactant in disorders such as lupus nephritis, that clearly require adequate assessment of GFR. B2M has the disadvantage of being increased in patients with several malignancies, particularly lymphoproliferative disorders including multiple myeloma. Like Cys C and BTP, B2M has the advantages of age and muscle mass independence (Filler et al., 2002).

Markers of acute kidney injury Recently, a number of biomarkers have been proposed for the early diagnosis, predicting the prognosis and differential diagnosis of acute kidney injury (AKI) (see Chapter 222) (Fig. 7.2).

Neutrophil gelatinase-associated lipocalin Neutrophil gelatinase-associated lipocalin (NGAL) or lipocalin 2 (LCN2) is a 25-kDa protein covalently bound to matrix metalloproteinase-9 from neutrophils (Devarajan 2010). It serves as a natural immunity protein with antibacterial properties. NGAL is also expressed by other cells among which are kidney epithelial and tubular cells (Devarajan, 2010; Ichino et al., 2009). NGAL expression is induced in injured cells, and both serum and urine NGAL is a promising biomarker of AKI (Devarajan 2010).

Table 7.5  Various methods for assessing GFR using endogenous markers (Delanghe, 2008) Marker (molecular weight, kDa)

Standardization

Remark

Creatinine (0.131)

NIST SRM 967

Partially determined by tubular secretion, dependent on muscle mass

Conventional creatinine clearance

NIST SRM 967

Biased due to urine collection problems. Overestimation of GFR due to tubular secretion

Paediatric estimation formulas (e.g. Schwartz, Counahan)

No longer compatible with new NIST SRM 967 creatinine standard. Partly depends on body constitution. Large variation for constant factor reported

Cystatin C (13)

International standardization is in progress

Ciclosporin and corticosteroids may interfere; some tumours upregulate cystatin C. Results may be affected by thyroid dysfunction

Beta trace protein (± 26)

Not available

Limited data

Beta 2 microglobulin (11.3)

Not available

Increased in some inflammatory conditions and some malignancies

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Table 7.6  Formulas for assessing GFR using endogenous markers Formula

Calculation

Remark

Cockcroft–Gault

eCCr = (140 – age) × mass (kg) × (0.85 if female)/72 × serum creatinine (mg/dL)

Estimates creatinine clearance

CCr-corrected = CCr × 1.73/BSA

A corrected formula for body surface area (BSA)

eGFR = 186 × serum creatinine−1.154 × age−0.203 ×

May only be used in subjects aged 18–70. Quantitative expression not valid when GFR vales > 60 mL/min

MDRD

(1.212 if black) × (0.742 if female) CKD-EPI

eGFR = 141 × min (SCr/k,1)a × max (SCr/k,1)−1.209 × 0.993age × (1.018 if female) × 1.159 if black)

Estimates GFR

Schwartz

eGFR = 0.413 × (height/serum creatinine) (conventional units)

Used in children

Counahan–Barratt

eGFR = 0 − 43 height (cm)/serum creatinine (mg/100 mL).

Used in children

Lund–Malmö formula

eGFR = eX − 0.00587 × age + 0.00977 × LBM

Cystatin C based, valid over a broad age range

Urinary NGAL can be assayed using commercial immunoassays (Bennett et al., 2008; Grenier et al., 2010). The cut-off value for urinary NGAL is 100 micrograms/L (Hirsch et al., 2007; Bennett et al., 2008). In urinary tract infections, increased urinary NGAL concentrations have been observed (Yilmaz et al., 2009). As urinary neutrophils can be regarded as a potential source of urinary NGAL, leucocyturia contributes to urinary NGAL concentrations and affects thus the interpretation of urinary NGAL results (Decavele et al., 2011). In urine specimens with a high white blood cell (WBC) count, (e.g. intensive care patients) (Bagshaw et al., 2010; Cruz et al., 2010), the use of a fixed cut-off value for interpreting urinary NGAL data may therefore lead to false-positive results. On average, one leucocyte corresponds with 12 pg of urine NGAL. Although the contribution of neutrophil NGAL to urinary NGAL is generally small, the effect may become important if there is a high leucocyte count and urinary NGAL levels may exceed 100 micrograms/L. A mathematical correction compensating neutrophil NGAL contribution may increase the diagnostic efficiency of NGAL in presence of increased NGAL levels and high urinary WBC counts: Corrected NGAL ( micrograms / L ) = NGAL − 0.12 urinary WBC coun t, 109 cells / L .

(

)

In tubular dysfunction, tubular impairment may also contribute to elevated urine NGAL concentrations (Decavele et al., 2011). In patients with CKD, an inverse correlation with estimated GFR has been described for both serum NGAL and urinary NGAL, suggesting that under these particular conditions this protein may also represent a surrogate index of residual renal function (Bolignano et al., 2009).

Kidney injury molecule-1 In animal models, kidney injury molecule-1 (KIM-1) is one of the most highly induced proteins in the kidney after AKI, and a proteolytically processed ectodomain of KIM-1 is easily detected in the urine soon after AKI. Enzyme-linked immunosorbent assay (ELISA)-based assays for KIM-1 have been developed, but are not yet commercially available. In a small human cross-sectional study,

KIM-1 was markedly induced in proximal tubules from kidney biopsies in patients with established AKI, and urinary KIM-1 distinguished ischaemic AKI from prerenal azotaemia and chronic renal disease. In a case–control study of children undergoing cardiac surgery, urinary KIM-1 levels were increased in subjects who developed AKI, with an AUC of 0.83 at the 12-hour time point (Han et al., 2008). In a larger prospective cohort study of hospitalized patients with established AKI, urinary KIM-1 was associated with adverse clinical outcomes, including dialysis requirement and death (Liangos et al., 2007). An advantage of KIM-1 as a urinary biomarker is that, in contrast to NGAL, its expression seems to be limited to the injured or diseased kidney since no systemic source of KIM-1 has been described. KIM-1 is induced in the kidney and upregulated in the urine by a large number of nephrotoxins, including ciclosporin, cisplatin, gentamicin, and heavy metals.

Liver fatty acid binding protein Liver fatty acid binding protein (L-FABP) is a 14-kDa protein normally expressed in the proximal convoluted and straight tubules of the kidney, and upregulated in animal models of AKI. An ELISA for this analyte is commercially available. It was used to demonstrate that urinary L-FABP levels were significantly increased prior to the increase in serum creatinine in those patients who developed AKI post contrast dye. In a recent prospective study of children undergoing cardiac surgery, urine L-FABP increased at 4 hours post-bypass, with an AUC of 0.810 for a cut-off value of 486 ng/mg creatinine (Portilla et al., 2008). However, increased urinary L-FABP levels are also found in patients with non-diabetic CKD, early diabetic nephropathy, idiopathic focal glomerulosclerosis and polycystic kidney disease. Additionally, L-FABP is also abundantly expressed in the liver, and urinary L-FABP may be influenced by serum L-FABP levels.

Proteinuria/differentiation of proteinuria Proteinuria Excessive urinary protein excretion is a cardinal sign of renal disease and a pathogenetically important factor in the progression of

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Tubular proteins Alpha-1 microglobulin (A1M) Beta-2 microglobulin (B2M)

New biomarkers

Retinol binding protein (RBP)

Neutrophil gelatinase-associated lipocalin (NGAL)

Fatty acid binding protein (FABP)

Kidney injury molecule-1 (KIM-1) Cystatin C

Biomarkers for diagnosis of acute kidney injury

Delayed biomarkers for kidney injury

Renal lysosomal enzymes

Creatinine

N-Acetyl-β-D-glucosaminidase (NAG)

Cystatin C

Fig. 7.2  Biomarkers for early diagnosis, predicting prognosis and differential diagnosis of acute kidney injury (AKI).

renal and cardiovascular disease. Further analysis of the type of proteinuria provides information about the possible underlying renal disease (e.g. glomerulonephritis, interstitial nephropathy). In the step-by-step diagnostic work-up of renal disease, knowledge about the type of proteinuria (low molecular/high molecular mass) is useful. Proteinuria may be due to prerenal (overflow proteinuria), renal (glomerular, tubular), or postrenal (bleeding or infection of the kidneys, ureter, bladder, or urethra) causes. A protein excretion exceeding approximately 200 mg/24 hours (based on a daily urinary volume of 1.5 L) is considered clinically relevant. The reference range for urinary protein excretion is lower: < 150 mg in 24 hours. Under physiological conditions, urinary protein excretion does not exceed 150 mg/day for adults and 140 mg/m2 of body surface for children. The daily physiological proteinuria contains mucoprotein (e.g. Tamm–Horsfall glycoprotein, 70 mg), blood group-related substances (35 mg), albumin (16 mg), immunoglobulins (6 mg), mucopolysaccharides (16 mg), and very small amounts of other proteins such as hormones and enzymes (King and Boyce, 1963). There are four categories of pathological proteinuria:  ‘glomerular’, ‘tubular’, ‘overload’, and ‘benign’ (Abuelo, 1983). Apart from this, proteinuria can also be factitious as a presentation of Munchausen’s syndrome. Glomerular proteinuria is due to an increased glomerular permeability to proteins and occurs in primary and secondary glomerulopathies. It can be further classified into selective and non-selective proteinuria. Selective proteinuria is characterized by the presence of predominantly anionic proteins with a molecular weight of 50–80 kDa (e.g. albumin). In the case of non-selective proteinuria, larger proteins with a molecular weight > 50–80 kDa are detected, in addition to albumin. This is indicative for a more severe damage of the glomeruli. Tubular proteinuria is due to a decreased tubular reabsorption of proteins

normally present in the glomerular filtrate. It is seen in tubular and interstitial disorders, including those which develop in the course of chronic glomerular diseases. Overload proteinuria is secondary to an increased production, or release of low-molecular weight proteins, which are usually reabsorbed by the proximal tubular cells, such as immunoglobulin light chains (which are increased in monoclonal gammopathies), lysozyme (which is increased in some leukaemias), or myoglobin (which is increased in rhabdomyolysis). Benign proteinuria includes functional proteinuria, as seen in fever or after exercise, idiopathic transient proteinuria, and orthostatic proteinuria. Factitious proteinuria may originate from addition of exogenous proteins to urine specimens. This is sometimes observed in Münchausen syndrome (Boelaert et al., 1984), psychiatric disorders known as factitious disorders wherein those affected feign disease, illness, or psychological trauma in order to draw attention or sympathy to themselves. Once pathological proteinuria has been detected, it must be quantified and submitted to qualitative analysis to discover to which of the above four categories it belongs.

Detection Qualitative evaluation Dipsticks See also ‘Dry chemistry’ and ‘Albuminuria’. Although dipstick urinalysis is characterized by a low cost, a wide availability, and a short procedure time, to date there is no consensus among the guidelines regarding the use of dipsticks as a front-line screening test for proteinuria in asymptomatic individuals (National Kidney Foundation 2002; Konta et al., 2007; Graziani et al., 2009; Lamb et al., 2009). Recent studies show a high negative predictive value of dipstick testing and a minimal risk of falsely

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negative cases of macroalbuminuria, whereas positive results require further confirmatory testing (White et al., 2011).

Precipitation methods Precipitation methods evaluate the turbidity occurring after proteins are precipitated by sulphosalicylic acid, trichloracetic acid, or by heat and acetic acid-sodium acetate buffer. Turbidimetric methods detect all urinary proteins, but alpha-1-acid glycoprotein and Tamm–Horsfall mucoprotein are not precipitated by sulphosalicylic acid (Shiba et al., 1985). They are very sensitive and may detect protein concentrations as low as 2.5 mg/L. With these methods, the protein concentration is expressed on a semiquantitative scale, from 0 to +++ or ++++. Many drugs can cause positive interference, for example, large amounts of penicillin or cephalosporin analogues, miconazole, tolbutamide, or sulphonamide metabolites. False-positive results can also be caused by the presence in the urine of radiocontrast media.

Quantitative evaluation Urine proteins can be quantified by several methods (Table 7.7), but as urine can contain a large range of proteins and also many interfering substances, all available methods are imperfect. Benzethonium chloride (Iwata and Nishikaze, 1979), trichloroacetic acid precipitation (Henry et al., 1956), dye binding methods with Brome-phenol blue (Hemmingsen, 1972), Coomassie brilliant blue (Bradford, 1976), Ponceau red (Pesce and Strande, 1973) and pyrogallol-red, nephelometry, and turbidimetry have been suggested. All these methods can be automated, in contrast to the biuret examination (Weicheselbaum, 1946). Determination of total protein is a compromise because no procedure detects all the proteins in urine. The upper limits of reference intervals in healthy individuals are method-dependent. Several intervals are cited:  pyrogallol red method:  180 mg/day (Orsonneau et  al., 1989), for pregnant women: 260 mg/day (Higby et al., 1994); biuret method: 150 mg/ day; turbidimetric method:  75 mg/day (Henry et  al., 1956), turbidimetric method:  protein/creatinine 8 g/mol (second morning urine), or 70 mg/g in conventional units. Turbidimetric methods Although these methods are among the most frequently used (e.g. benzethonium chloride in alkaline medium), they suffer from major drawbacks. Trichloroacetic acid is unaffected by protein composition and can detect low protein concentrations, if the temperature of urine ranges between 20 and 25°C. However, numerous drugs can interfere with this method (Lorentz and Weiss 1986). The

method based on benzethonium chloride gives a stable turbidity and is less dependent on temperature than the previous methods. However, it produces 11–31% less turbidity for gammaglobulins than for albumin, and gives unevenly dispersed aggregates in the presence of large protein concentrations, with consequent falsely low protein estimates (McElderry et al., 1982). Dye-binding techniques These are based on the interaction between proteins and a dye, which causes a shift in the absorption maxima (measured photometrically) of the dye. Coomassie brilliant blue G250 (Bradford, 1976) is very sensitive, but with diluted samples significant deviations from linearity are possible. Moreover, underestimation of tubular proteinuria and interference from various metabolites, drugs, and preservative compounds has been observed. The method is somewhat improved by the addition of sodium dodecylsulphate. Ponceau S (Pesce and Strande, 1973) is equally sensitive to albumin and globulins, but falsely low values can result from the loss of precipitate during the decantation step while falsely high values may derive from the contamination of the precipitate by unbound dye. Aminoglycoside antibiotics interfere positively. Pyrogallol red-molybdate method can easily be implemented on automated analysers. This method has been improved by the addition of sodium dodecylsulphate (Orsonneau, 1989). Biuret methods These methods are based on the interaction between copper ions and the carbamide group of proteins (Lorentz and Weiss, 1986). They have the same sensitivity for all proteins. The protein in urine must be concentrated before the biuret reaction occurs. Interference from drugs, radiographic contrast media, and coloured metabolites is minimal (Bradford, 1976; Lorentz and Weiss, 1986). A modification of the biuret method, which is now the reference method recommended by the American Association for Clinical Chemistry, utilizes gel filtration to exclude small interfering compounds.

The protein:creatinine ratio The 24-hour urine collection is considered as the ‘gold standard’ for the quantitative evaluation of proteinuria. However, it is time-consuming, subject to error, and is frequently associated with significant collection errors, which are mainly due to improper timing and missed samples. The calculation of the protein:creatinine ratio on spot urine samples, which corrects for variations in urinary concentration due to hydration, is an alternative to the 24-hour

Table 7.7  Some quantitative methods for proteinuria Method Turbidimetric Trichloroacetic Benzethonium chloride Dye binding Coomassie brilliant blue Ponceau Biuret

Analytical sensitivity (mg/L)

Linearity (mg/L)

Distinctive features

20 10 2.5 20

20–2400 10–1600 5–1500 100–1600

Same sensitivity for albumin and globulins, many drugs can interfere Albumin overestimation, underestimation of increased protein concentrations High sensitivity, underestimation of tubular proteins Same sensitivity for albumin and globulins, positive interference with aminoglycoside antibiotics

chapter 7 

collection (Ginsberg et al., 1983). The measurements obtained with this method correlated well with those obtained in the classical way. All patients with proteinuria of > 3.5 g/24 hours had a ratio of greater than 3.5 in single voided samples, and all patients with a proteinuria of < 0.2 g/24 hours had a ratio of < 0.2. The applicability and reliability of this method has been confirmed by several investigators (Rodby et al., 1995; Ruggenenti et al., 1998), and is now recommended by the American Guidelines (National Kidney Foundation, 2002). The protein:creatinine ratio is preferably calculated on the first morning urine. This avoids the possible variations caused by the circadian rhythm of protein excretion, which is maximal during the day and minimal during the night, compared to the urinary excretion of creatinine, which is fairly constant over the day (Koopman et al., 1989).

Further exploration and selectivity of proteinuria In the case of a positive qualitative protein test, the cause of proteinuria should be investigated. Urine protein electrophoresis allows a differentiation of proteinuria. Electrophoretic techniques (Fig. 7.3) are essential in detecting the clonality of serum paraproteins (M components) and monoclonal immunoglobulin light chains in urine (Bence Jones proteins) (Merlini et al., 2001). Today immunofixation of urine or serum is the gold standard technique in defining the isotype of the myeloma clone. However, it can occasionally give misleading ‘pseudo-oligoclonal’ patterns in urine specimens that contain polyclonal immunoglobulins (Harrison et al., 1992). Some authors recommend immunofixation electrophoresis of all specimens because of the sensitivity of the method. As an alternative to screening and in monitoring of proteinuria, turbidimetric quantitation of urinary kappa or lambda chain excretion is also suggested, combined with immunofixation in case of a positive finding with unknown clonality. Another way to distinguish the different types of proteinuria is based on the measurement of specific proteins (e.g. albumin, transferrin, alpha-1-microglobulin (A1M), alpha-2-macroglobulin (A2M)). Electrophoresis provides useful information in problem cases (e.g. Von Münchausen syndrome or Münchausen by proxy syndrome—falsification of laboratory results by adding substances to urine).

1

2

3

4

5

6

Fig. 7.3  Various types of proteinuria: serum protein pattern for comparison (lane 1), selective (lane 2) and non-selective (lane 3) glomerular proteinuria, tubular proteinuria (lane 4), Bence Jones proteinuria (lanes 5 and 6).

assessment of renal function

Agarose-electrophoresis after protein concentration and the use of a very sensitive staining (e.g. gold stains) is one of the most widely used methods. Size fractionation of urinary proteins on sodium dodecyl sulphate-acrylamide gel electrophoresis (SDS-PAGE) has been used to classify prerenal, renal, and post-renal proteinuria before quantitative measurements of specific proteins became widely available. SDS-PAGE is a laborious procedure and is therefore rarely used in the routine laboratory. There is a tendency towards using specific urinary proteins as marker proteins. In overflow proteinuria, excessive amounts of small plasma proteins exceeding the reabsorption capacity of the proximal tubulus result in a protein excretion in the final urine (overflow proteinuria). Typical examples are haemoglobin monomers, for example, intravascular haemolysis and immunoglobulin light chains (Bence Jones proteins) in a monoclonal gammopathy. Postrenal proteinuria occurs when proteins enter the urine because of a haemorrhage or an inflammation within the lower urinary tract or genital tract. As a marker protein in such situation, A2M is mainly determined, as it cannot be filtered by the glomerulus because of its 725-kDa molecular mass. The selectivity of glomerular proteinuria is classically evaluated by the ratio of the clearance of IgG (molecular weight 150 kDa) to the clearance of transferrin (molecular weight 80 kDa). When the ratio is < 0.1 the proteinuria is defined as selective (Cameron and Blandford, 1966). In children with nephrotic syndrome, this strongly suggests the presence of minimal change disease responsive to corticosteroids. Other approaches are based on the ratio of the clearance of IgM to that of albumin (Bakoush et al., 2001).

Albuminuria The excretion of small amounts of albumin in urine was historically called ‘microalbuminuria’. According to the American Diabetes Association (2014), low-grade albuminuria corresponds to an albumin excretion rate of approximately 30–300 mg/24 hours. However, albuminuria is characterized by a large intraindividual variability. Low-grade albuminuria is an excellent marker for assessing vascular damage and early renal disease. Generalized atherosclerosis leads to medullary hypoperfusion and hypoxia, which induces renal tubular dysfunction. An impaired reabsorption of albumin and other tubular proteins is observed. Increased albumin excretion is regarded as the earliest marker for detecting changes in the glomerular basal membrane. At this stage, the glomerulopathy can still be modified by taking appropriate measures (strict metabolic control in diabetics, blood pressure reduction in hypertensives by association of angiotensin-converting enzyme inhibitors or angiotensin II-receptor blockers), preventing progression towards renal insufficiency. False-low results may occur in extreme proteinuria because of an antigen excess. Screening is not indicated in the presence of urinary tract infections, following strenuous physical exercise, uncontrolled diabetes mellitus or hypertension, cardiac insufficiency, fever, and menses. These factors may induce a temporary increase in albumin excretion. In contrast to cases of overt proteinuria or macroalbuminuria, 80% of the urinary proteins in normo- and microalbuminuria are of non-albumin nature. Further research is required to clarify the importance of specific non-albumin proteins in urine (Methven et al., 2011). The 24-hour urine collection has been considered the ‘gold standard’ method for the measurement of albuminuria due to

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the circadian rhythm of urinary protein excretion (Hansen et al., 2002). However, the urine albumin:creatinine ratio (ACR) on random urine samples can also be used (Assadi, 2002), as it has been demonstrated that total urinary protein and albumin excretion are equally powerful predictors of all-cause mortality and renal outcomes in patients with CKD attending a general nephrology clinic (Methven et al., 2011). As the urinary albumin concentration depends on the hydration status, the constant urinary excretion of creatinine ensures that the urinary ACR adjusts for hydration changes (Heerspink et al., 2010). Moreover, this ratio incorporates the predictive value of poor health as reflected by low muscle mass and decreased creatinine excretion (Nangaku, 2006). The most common methods to measure albuminuria are nephelometric (Stamp 1988) and turbidimetric assays, but those measurements are expensive, have a delay in the provision of results and require a specialist biochemistry laboratory (Shukla et  al., 1988; White et al., 2011). Urinary albumin measurements are currently not standardized due to a lack of a reference method and reference (primary and secondary (matrix)) material. Multiple molecular forms of albumin in urine are identified (Fig. 7.4). Modification of albumin by proteolysis during passage through the urinary tract and chemical modification during specimen storage leads to the formation of albumin fragments. Multiple methods have been developed to quantify albuminuria and significant different results are reported dependent on the available assay. The current point of view of the National Kidney Disease Education Program—IFCC Working Group on Standardization of Albumin considers the immunoassay with polyclonal sera as the primary method of quantifying urine albumin. At present there are no hard arguments for measuring immunochemically non-reactive albumin in urine.

Immunoassays using polyclonal antisera for the detection of urinary albumin remain the gold standard. The development of a reference measurement procedure remains one of the challenges for the future (Speeckaert et al., 2011). Test strips are also available, but the use of those strips as a screening tool is controversial due to the limited evaluation of the diagnostic accuracy of dipsticks for the general population. In a large Australian population-based cohort study, a dipstick (Bayer Multistix 10 SG) reading of ≥ 1+ identified adults with ACR ≥ 30 mg/g with high specificity and was characterized by a strong negative predictive value and a tendency toward a higher false-positive rate. There was a minimal risk of missed cases of macroalbuminuria (the identification of ACR ≥ 300 mg/g had a false-negative rate of < 0.1%). However, a large variation in positive predictive values was observed across low- and high-risk subgroups according to albuminuria prevalence. The diagnostic accuracy was not significantly influenced by the presence of diabetes, hypertension, or age. An inferior test sensitivity was reported in women, which may be explained by the use of creatinine to correct for concentration in the measurement of the reference standard (White et al., 2011). There may be an important difference between reagent strips produced by different manufacturers in detecting the same threshold of albuminuria or in calibration to indicate a positive test result (trace, 1+, or > 1+) (Lamb et al., 2009). As a significant heterogeneity in diagnostic test performance across different populations has been observed, evaluation of the test performance in a large population group remains essential (Konta et al., 2007; White et al., 2011). It is recommended that a positive strip test is followed by a quantitative determination (American Diabetes Association, 1994). New studies should focus on the impact of treatment interventions in individuals

Renal tubule Albumin

Albumin fragments

Cubilin Megalin Amnionless Proximal tubular cell

Early endosome Lysosome/ late endosome hybrid

Late endosome Reabsorption of albumin

Degradation of albumin Blood vessel

Fig. 7.4  Overview of the albumin fragmentation process. Derived from Speeckaert et al., (2011).

Lysosome

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with low-grade albuminuria and update the cost-effectiveness of dipsticks as a screening tool (White et al., 2011).

Tubular proteins Tubular proteinuria occurs when glomerular function is normal, but the proximal tubules have a diminished capacity to reabsorb and to catabolize proteins, causing an increased urinary excretion of the low molecular mass proteins that normally pass through the glomerulus, such as retinol-binding protein, A1M, and B2M. Since these products are present in the plasma in low concentrations, urine protein excretion in tubular proteinuria is usually < 1.5 g/day. Nephrotoxins that damage the proximal tubule include heavy metals (e.g. cadmium and lead) and some drugs (e.g. aminoglycosides, ciclosporin, cytostatic agents, and analgesics). Tubular proteinuria is also associated with acute and chronic pyelonephritis, renal vascular diseases, kidney transplant rejection, Fanconi syndrome, and Balkan nephropathy.

Alpha-1-microglobulin A1M is a 27-kDa glycoprotein produced by the liver and has been given other names, for example, complex-forming glycoprotein, heterogeneous in charge, protein HC (Penders and Delanghe, 2004). In plasma, approximately 50% of A1M forms a complex with IgA. Free, monomeric A1M passes through the glomerulus out into the primary urine, from which it is reabsorbed by the proximal tubules where it is catabolized. The reference range of the urinary concentration is 1000 mg/g creatinine), does the A1M concentration tend to be elevated with increasing albuminuria (Hoffmann et al., 1995). A1M is a useful marker for assessing renal damage in urological diseases (e.g. obstructive uropathy). Assaying urinary A1M provides a diagnostic tool for the diagnosis and monitoring of urinary tract disorders. A1M excretion was related to the grade of vesicoureteral reflux. The urinary A1M/creatinine ratio was highly sensitive and specific and correlated with DMSA scintigraphy. In contrast to cystitis, elevated urinary A1M excretion is seen in acute pyelonephritis (Everaert et al., 1998). Determination of urinary A1M has also been used in intoxications with heavy metals (cadmium (Jung et  al., 1993; Fels et  al., 1994), lead (Lim et al., 2001), and mercury-induced renal dysfunction (Kobal et al., 2000) at an early stage). Urinary A1M as a tubular marker and albumin as a glomerular marker were successfully used as an early marker for Balkan nephropathy screening (Cvorisćec, 2000). Urinary A1M can allow an early diagnosis of tubular proteinuria. In idiopathic membranous nephropathy, excretion of

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IgG and A1M was associated with the extent of tubulointerstitial damage. A1M is also useful for the early detection of nephropathy in diabetic subjects. The proximal tubules are damaged early during the period of subclinical diabetic nephropathy (Marczewski et al., 1996).

Beta-2-microglobulin B2M (molecular weight:  11800)  has been used as a marker protein for tubulopathy (Guder et al., 1998, Delanghe, 2007). B2M is filtered by the glomerulus and almost completely reabsorbed and catabolized in the proximal tubules. B2M is a marker of tubular reabsorption. However, the protein is unstable at urinary pH values < 6. In tubulointerstitial diseases, the activity of urinary proteases may be increased, which may accelerate the breakdown of B2M. The reference range is < 0.3 mg/L or < 0.2 mg/g creatinine. B2M is a less reliable urine marker than other microproteins since various diseases (multiple myeloma, Waldenström’s disease, B-cell lymphomas, primary systemic amyloidosis, HIV-infection) raise its serum level, which in turn increases renal elimination. B2M can be measured using nephelometry or turbidimetry.

Retinol binding protein Because of its low molecular mass (~ 21 kD), retinol binding protein (RBP) is almost completely filtered by the glomerulus. In the proximal tubules > 99% is reabsorbed. Its reference range in urine is < 0.5 mg/L. In case of an impaired tubular protein resorption, increased RBP values are found in urine. The protein can be measured using immunonephelometry.

Alpha-2-macroglobulin For the evaluation of a significant haematuria (RBC > 25 cells/μL), another specific protein (A2M) can be used as a marker for postrenal bleeding. A2M is a high molecular weight (725 kDa) endoprotease inhibitor, present in plasma. The protein cannot pass the glomerular basement membrane, even in glomerular nephropathy. Since it is not related to or dependent of the urinary RBC-concentration, A2M can provide additional information in haematuria cases. The reference range of A2M/creatinine in urine is < 10 mg/g (or 1.13 mg/mmol). High values are observed in postrenal bleeding (Guder et al., 1998).

N-Acetyl-beta-D-glucosaminidase N-Acetyl-beta-D-glucosaminidase (ß-NAG) is a lysosomal enzyme showing a high activity in the proximal tubulus. Increased urinary ß-NAG activities are observed following toxic damage to the proximal tubular and interstitial cells (e.g. tubular dysfunction, nephritis, nephrotoxic substances (aminoglycosides, cytostatic drugs, contrast media)). ß-NAG is stable at room temperature for approximately 3 hours, at 4°C up to 50 days. When urine pH values exceed 8, enzyme activity drops. In the presence of high glucose concentrations, activity diminishes because of non-enzymatic glycation. The reference range is lower than < 6.3 U/L or < 5 U/g creatinine. One possible approach is based on the simultaneous measurement in the same specimen of different proteins both of glomerular (albumin and IgG) and tubular origin (A1M). The simultaneous measurement of IgG and A1M can be used to evaluate the course and response to treatment in glomerular diseases such as membranous nephropathy (Bazzi et al., 2001).

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Enzymuria Normal individuals excrete small amounts of enzymes located in the cells of the renal tubules in their urine (Pesce and First, 1979). Increased excretion of these enzymes is a sensitive index of renal damage (Marhun, 1979). These measurements have the advantage of being easily made and have a great sensitivity, but unfortunately are not specific for the disease processes and are not therefore very useful for diagnostic purposes. A number of brush-border enzymes have been studied in normal and pathological urines. Most data relate to NAG, a hydrolytic enzyme present in lysosomes (Sherman et  al., 1983). The excretion of tubular enzymes or NAG increases in a great variety conditions e.g. use of aminoglycoside antibiotics (Mondorf et al., 1984), cisplatin (Rojanasthien et al., 2001), contrast media (Erley et al., 1999), heavy metals and organic solvents (Tassi et al., 2000), glomerulonephritis (Valles et al., 2000), acute interstitial nephritis (Wolf et al., 1990), and extracorporeal shock-wave lithotripsy (Weichert-Jacobsen et al., 1998). The assay of urinary NAG or other tubular enzymes may detect minimal degrees of renal dysfunction, which is both an advantage and a disadvantage for clinical use. The problem in using these assays diagnostically is that, for example, modest renal insults, intercurrent urinary tract infections, increased urinary flow, and therapeutic doses of aminoglycosides will all increase urinary enzyme excretion. Thus, the clinical value of urinary NAG estimations in patients is limited.

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Marhun, D. (1979). Evaluation of urinary enzyme patterns in patients with kidney diseases and primary benign hypertension. Curr Probl Clin Biochem, 9, 135–9. Maroni, B. J., Steinman, T. I. and Mitch, W. E. (1985). A method for estimating nitrogen intake of patients with chronic renal failure. Kidney Int, 27, 58–65. McElderry, L. A., Tarbit, I. F. and Cassells-Smith, A. J. (1982). Six methods for urinary protein compared. Clin Chem, 28, 2294–6. Merlini, G., Marciano, S., Gasparro, C., et al. (2001). The Pavia approach to clinical protein analysis. Clin Chem Lab Med, 39, 1025–8. Methven, S., MacGregor, M. S., Traynor, J. P., et al. (2011). Comparison of urinary albumin and urinary total protein as predictors of patient outcomes in CKD. Am J Kidney Dis, 57, 21–8. Michels, W. M., Grootendorst, D. C., Verduijn, M., et al. (2010). Performance of the Cockcroft-Gault, MDRD, and new CKD-EPI formulas in relation to GFR, age, and body size. Clin J Am Soc Nephrol, 5, 1003–9. Miller, W. G. (2009). Estimating equations for glomerular filtration rate in children: laboratory considerations. Clin Chem, 55, 402–3. Mondorf, A. W., Scherberich, J. E., Falkenberg, F. W., et al. (1984). Brush border enzymes and drug nephrotoxicity. In K. Solez and A. Wellthon (eds.) Acute Renal Failure, pp. 281–98. New York: Dekker. Nagao, S., Fujiwara, K., Imafuku, N., et al. (2005). Difference of carboplatin clearance estimated by the Cockroft-Gault, Jelliffe, Modified-Jelliffe, Wright or Chatelut formula. Gynecol Oncol, 99, 327–33. Nangaku, M. (2006). Chronic hypoxia and tubulointerstitial injury: a final common pathway to end-stage renal failure. J Am Soc Nephrol, 17, 17–25. Narva, A. S. (2009). Assessment of kidney function for drug dosing. Clin Chem, 55, 1609–11. National Kidney Disease Education Program, http://nkdep.nih.gov/ resources/ckd-drug-dosing-508.pdf National Kidney Foundation (2002). K/DOQI clinical practice guidelines for chronic kidney disease: evaluation, classification, and stratification. Am J Kidney Dis Suppl, 39, S1–S266. National Kidney Foundation (2005). K/DOQI clinical practice guidelines for chronic kidney disease: evaluation, classification, and stratification. Part 1. Executive summary. Am J Kid Dis Suppl, 39, S17–S30. Notghi, A., Merrick, M. V., Ferrington, C., et al. (1986). A comparison of simplified and standard methods for the measurement of glomerular filtration rate and renal tubular function. British Journal of Radiology 59, 35–9. Nyman, U., Björk, J., Lindström, V., et al. (2008). The Lund-Malmö creatinine-based glomerular filtration rate prediction equation for adults also performs well in children. Scandinavian Journal of Clinical Laboratory Investigation, 68, 568–76. Orsonneau, J. L., Douet, P., Massoubre, C., et al. (1989). An improved pyrogallol red-molybdate method for determining total urinary protein. Clin Chem, 35, 2233–6. Panteghini, M. and IFCC Scientific Division. (2008). Enzymatic assays for creatinine: time for action. Clin Chem Lab Med, 46, 567–72. Payn, M. M., Webb, M. C., Lawrence, D., et al. (2002). Alpha1-microglobulin is stable in human urine ex vivo. Clin Chem, 48, 1136–8. Penders, J. and Delanghe, J. R. (2004). Alpha 1-microglobulin: clinical laboratory aspects and applications.Clin Chim Acta, 346, 107–18. Pesce, A. J. and First, R. M. (1979). Proteinuria. An Integrated Review. New York: Dekker. Pesce, M. A. and Strande, C. S. (1973). A new micromethod for determination of protein in cerebrospinal fluid and urine. Clin Chem, 19, 1265–7. Poggio, E. D., Wang, X., Greene, T., et al. (2005). Performance of the Modification of Diet in Renal Disease and Cockcroft-Gault equations in the estimation of GFR in health and in chronic kidney disease. J Am Soc Nephrol, 16, 459–66.

Portilla, D., Dent, C., Sugaya, T., et al. (2008). Liver fatty acid-binding protein as a biomarker of acute kidney injury after cardiac surgery. Kidney Int, 73, 465–72. Roberts, G. W., Ibsen, P. M. and Schiøler, C. T. (2009). Modified diet in renal disease method overestimates renal function in selected elderly patients. Age Ageing, 38, 698–703. Rodby, R. A., Rohde, R. D., Sharon, Z., et al. (1995). The urine protein to creatinine ratio as a predictor of 24-hour urine protein excretion in type 1 diabetic patients with nephropathy. The Collaborative Study Group. Am J Kidney Dis, 26, 904–9. Rojanasthien, N., Kumsorn, B., Atikachai, B., et al. (2001). Protective effects of fosfomycin on cisplatin-induced nephrotoxicity in patients with lung cancer. Int J Clin Pharmacol Ther, 39, 121–5. Ruggenenti, P., Gaspari, F., Perna, A., et al. (1998). Cross sectional longitudinal study of spot morning urine protein: creatinine ratio, 24 urine protein excretion rate, glomerular filtration rate, and end stage renal failure in chronic renal disease in patients without diabetes. Br Med J, 316, 504–9. Schlebusch, H., Liappis, N., Kalina, E., et al. (2002). High sensitive CRP and creatinine: reference intervals from infancy to childhood. J Lab Med, 26, 341–6. Schuhmann-Giampieri, G. and Krestin, G. (1991). Pharmacokinetics of Gd-DTPA in patients with chronic renal failure. Invest Radiol, 26, 975–79. Schwartz, G. F., Feld, L. G. and Langford, D. J. (1984). A simple estimate of glomerular filtration rate in full-term infants during the first year of life. J Pediatr, 104, 849–54. Schwartz, G. J. and Furth, S. L. (2007). Glomerular filtration rate measurement and estimation in chronic kidney disease. Pediatr Nephrol, 22, 1839–48. Schwartz, G. J., Furth, S., Cole, S. R., et al. (2006). Glomerular filtration rate via plasma iohexol disappearance: pilot study for chronic kidney disease in children. Kidney Int, 69, 2070–7. Schwartz, G. F. and Gauthier B. (1985). A simple estimate of glomerular filtration in adolescent boys. J Pediatr, 106, 522–26. Schwartz, G. J., Haycock, G. B., Edelmann, C. M. Jr, et al. (1976). A simple estimate of glomerular filtration rate in children derived from body length and plasma creatinine. Pediatrics, 58, 259–63. Schwartz, G. J., Muñoz, A., Schneider, M. F., et al. (2009). New equations to estimate GFR in children with CKD. J Am Soc Nephrol, 20, 629–37. Sherman, R. L., Drayer, D. E., Leyland-Jones, B. R., et al. (1983). N-acetyl-β-glucosaminidase and β2-microglobulin: their urinary excretion in patients with parenchymatous renal diseases. Arch Intern Med, 143, 1183–5. Shiba, K. S., Kanamori, K., Harada, T., et al. (1985). A cause of discrepancy between values for urinary protein as assayed by the Coomassie Brilliant Blue G-250 method and the sulphosalycilic acid method. Clin Chem, 31, 1215–8. Shukla, P. S., Endeman, H. J. and Kortlandt, W. (1988). Effects of five different antisera on the immunoturbidimetric determination of urinary albumin. Clin Chem, 34, 430. Speeckaert, M. M., Speeckaert, R., Van De Voorde, L., et al. (2011). Immunochemically unreactive albumin in urine: fiction or reality? Crit Rev Clin Lab Sci, 48, 87–96. Stamp, R. J. (1988). Measurement of albumin in urine by end-point immunonephelometry. Ann Clin Biochem, 245, 442–5. Stevens, L. A. and Levey A. S. (2004). Clinical implications for estimating equations for GFR. Ann Intern Med, 141, 959–61. Stevens, L. A. and Levey, A. S. (2009a). Measured GFR as a confirmatory test for estimated GFR. J Am Soc Nephrol, 20, 2305–13. Stevens, L. A. and Levey, A. S. (2009b). Current status and future perspectives for CKD testing. Am J Kid Dis, 53 (S3), S17–S26. Stevens, L. A., Nolin, T. D., Richardson, M. M., et al. (2009). Comparison of drug dosing recommendations based on measured GFR and kidney function estimating equations. Am J Kid Dis, 54, 33–42. Swaminathan, S. and Shah, S. V. (2007). New insights into nephrogenic systemic fibrosis. J Am Soc Nephrol, 18, 2636–43.

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Tassi, C., Abbritti, G., Mancuso, F., et al. (2000). Activity and isoenzyme profile of N-acetyl-beta-d-glucosaminidase in urine from workers exposed to cadmium. Clin Chim Acta, 299, 55–64. Toffaletti, J., Blosser, N., Hall, T., et al. (1983). An automated dry-slide enzymatic method evaluated for measurement of creatinine in serum. Clin Chem, 29, 684–7. Vaidya, V. S., Ferguson, M. A. and Bonventre, J. V. (2008). Biomarkers of acute kidney injury. Annu Rev Pharmacol Toxicol, 48, 463–93. Valles, P., Peralta, M., Carrizo, L., et al. (2000). Follow-up of steroid-resistant nephrotic syndrome: tubular proteinuria and enzymuria. Pediatr Nephrol, 15, 252–8. Weichert-Jacobsen, K., Stöckle, M., Loch, T., et al. (1998). Urinary leakage of tubular enzymes after shock wave lithotripsy. Eur Urol, 33, 104–10. Weicheselbaum, T. E. (1946). An accurate and rapid method for the determination of proteins in small amounts of blood serum and plasma. Am J Clin Pathol, 10, 40–9.

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White, S. L., Yu, R., Craig, J. C., et al. (2011). Diagnostic accuracy of urine dipsticks for detection of albuminuria in the general community. Am J Kidney Dis, 58, 19–28. Wolf, G., Scherberich, J. E., Nowack, A., et al. (1990). Urinary excretion of dipeptidyl aminopeptidase IV in patients with renal diseases. Clin Nephrol, 33, 136–42. Wuyts, B., Bernard, D., Van den Noortgate, N., et al. (2003). Reevaluation of formulas for predicting creatinine clearance in adults and children, using compensated creatinine methods. Clin Chem, 49, 1011–14. Wyss, M. and Kaddurah-Daouk, R. (2000). Creatine and creatinine metabolism. Physiol Rev, 80, 1107–213. Yilmaz, A., Sevketoglu, E., Gedikbasi, A., et al. (2009). Early prediction of urinary tract infection with urinary neutrophil gelatinase associated lipocalin. Pediatr Nephrol, 24, 2387–92. You, L., Zhu, X., Shrubsole, M. J., et al. (2011). Renal function, bisphenol A, and alkylphenols: results from the National Health and Nutrition Examination Survey (NHANES 2003–2006). Environ Health Perspect, 119, 527–33.

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Tubular function Marijn Speeckaert and Joris Delanghe Introduction Assessment of tubular function is more complicated than the measurement of the glomerular filtration rate (GFR), because tubular functions are more complex and differ along the nephron. Tubular functions cannot be studied as a whole, but must be divided according to the different segments of tubule. Most investigations of the tubular functions also necessitate an accurate measurement of the GFR.

Investigating proximal tubular functions The first postglomerular segment of the nephron is the proximal tubule. This segment reabsorbs about 60% of the filtered water and the solutes in a heterogeneous manner. Some functions are specific for the proximal tubule and their alterations indicate proximal tubule dysfunction. (See also Chapters 20 and 41).

Proximal tubule dysfunction Several rare syndromes are characterized by isolated renal phosphate wasting. A generalized impairment of the proximal tubule is known as the Fanconi syndrome, in which hypophosphataemia, renal glucosuria, hypouricaemia, aminoaciduria, and proximal renal tubular acidosis due to bicarbonate loss in the urine are observed (Clarke et al., 1995). The classic example is X-linked hypophosphataemic rickets, in which the defect in proximal tubular phosphate transport is due to a mutation in the PHEX gene.

Concentrating and diluting urine The kidney plays a central role in water homeostasis by its ability to concentrate or dilute urine according to water intake and extrarenal losses. Water is absorbed in the proximal tubule, the thin descending limb, and the collecting tubule. In the proximal tubule, water movement from lumen to blood is iso-osmotic and driven by active solute transports:  solutes accumulate at the external side of the basolateral cell membranes, generating a transepithelial osmotic pressure difference that drives water flow. Consequently, when an osmotically active solute is present in the tubular fluid and cannot be reabsorbed, water reabsorption in the proximal tubule is decreased leading to an osmotic diuresis. In the proximal tubule transepithelial water transport uses two routes: transcellular and paracellular. Water crosses proximal cell membranes through the type 1 aquaporin, which is constitutively expressed at the apical and basolateral sides of the cells. In the thick ascending limb and the distal convoluted tubule, no net water reabsorption occurs; only solutes are retrieved from the

urine. The loop of Henle creates the countercurrent multiplication process and the corticomedullary osmotic gradient (Morello and Bichet, 2001). In the distal convoluted tubule, the lumen osmolality decreases to 60 mOsm/kg H2O. The collecting duct has extremely low water permeability in the absence of the antidiuretic hormone vasopressin. Under this condition, no water transport is observed along the collecting duct and the final urine is maximally diluted. In the presence of vasopressin, transepithelial movements of water occur across the principal cells. Three types of aquaporins are involved in water transport in the principal cells. Aquaporin 3 and 4 expression are restricted to the basolateral plasma membrane and are not regulated by vasopressin. They provide the pathways that permit water to enter the interstitium. Aquaporin 2 is expressed at the apical plasma membrane exclusively in the presence of vasopressin, thus increasing water permeability. The transepithelial difference in osmolality drives water from the lumen to the interstitium. When the level of vasopressin in plasma is high enough, water reabsorption occurs along the entire collecting duct and the final urine is maximally concentrated (Nielsen et al., 2002).

Testing the concentrating ability The concentrating ability of the kidney depends not only on the action of vasopressin but also on the presence of an adequate osmotic corticomedullary gradient. This gradient is altered at the early stage of renal insufficiency, and is profoundly reduced in particular in pathology with medullary damage such as adult polycystic kidney disease, nephronophthisis, sickle cell disease, reflux nephropathy, tubular necrosis, or transplanted kidneys. The assessment of the concentrating capacity of the kidney is based on the measurement of the urine osmolality. Plasma osmolality must be measured simultaneously for a better interpretation of the results. Urine and plasma osmolalities are measured by the depression of the freezing point. The study of the maximal capacity of the kidney to concentrate urine is important whenever a form of diabetes insipidus is suspected. This defect can be of a genetic origin or can be acquired. It can be located in the kidney (nephrogenic diabetes insipidus) or due to the lack of vasopressin secretion. The maximal concentrating capacity of the kidney is determined by a fluid restriction test (Davis and Zenser, 1993). The patient has to stop all fluid and food intake from midnight the day prior to the investigation. The next morning, plasma and urine osmolalities are measured. In normal subjects, plasma osmolality ranges between 280 and 295 mOsm/kg H2O and urine osmolality is > 600 mOsm/ kg H2O. The maximal urine concentration, that usually exceeds 1000 mOsm/kg H2O, is achieved after water deprivation from 18 to 24 hours (Miles et al., 1954; Rowe et al., 1976). The maximum

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concentrating ability is lower in infants and elderly people (Rowe et  al., 1976; Monnens et  al., 1981; Aperia et  al., 1983; Davis and Davis, 1987). During the investigation, the urinary flow falls below 0.4 mL/min, blood pressure and urine osmolality are measured every hour and plasma osmolality every 2 hours. The test is usually stopped when urine osmolality exceeds 800 mOsm/kg H2O, which excludes major defects in the urine concentrating process, or when plasma osmolality is above the upper normal values (296 mOsm/kg H2O). This will distinguish between nephrogenic diabetes insipidus and central deficiency of vasopressin secretion. In central diabetes insipidus, DDAVP (desmopressin) increases urine osmolality above 750 mOsm/kg H2O within the first 2 hours following the injection (Radó et al., 1976; Curtis and Donovan, 1979; Moses et  al., 1985; Williams et  al,. 1986). In nephrogenic diabetes insipidus, DDAVP injection does not increase urine osmolality further. A blood sample is usually drawn before DDAVP injection to allow plasma vasopressin concentration measurement in case of central deficiency. This test can be used in infants and children (Monnens et al., 1981). In compulsive water drinkers, urine osmolality rises above 600 mOsm/kg H2O. Surreptitious water ingestions induce a fall in urine osmolality between two successive collections accompanied by an increase in urinary flow rate.

Testing the diluting capacity The diluting capacity of the kidney can also be tested. This investigation is useful in patients who present with unexplained hyponatraemia. The lowest urinary osmolality is obtained when vasopressin secretion is completely suppressed. Diuretics, by decreasing sodium reabsorption especially in the distal convoluted tubule (thiazide), impair the diluting capacity. The diluting capacity requires normal solute intake. At the beginning of the investigation, plasma and urinary osmolality are measured, then the patient drinks a water load of 20 mL/kg body weight in 20 minutes (DeRubertis et al., 1971; Davis and Zenser, 1993). In normal subjects, the plasma osmolality remains within the normal range after the water load and urine osmolality rapidly decreases below 100 mOsm/kg H2O within 2 hours and 80% of the ingested water is excreted within 4 hours. An alteration of the diluting capacity can be due to an incomplete inhibition of vasopressin secretion, or to adrenal insufficiency, hypothyroidism, potassium depletion, or liver disease.

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For urine, the specific gravity is a function of the number and weight of the dissolved solute particles However, in some conditions there is a remarkable divergence between osmolality and specific gravity. If larger-molecular-weight substances are present, there will be divergence between specific gravity and osmolality. It used to be measured with a hydrometer, commonly called a ‘urinometer’, a weighted float marked with a scale for specific gravities from 1.000 to 1.060. The urinometer is simple and quick to use, but has some drawbacks:  it requires a volume of urine not always available in spot samples. There may be difficulty in reading the meniscus and there may be a tendency for the device to cling to the side of the container. Moreover, specific gravity varies with temperature and urinometers may be inaccurately calibrated. Urine specific gravity should be corrected for protein and glucose, if either is present (Schrier, 2007). Proteins increase the specific gravity by 0.001 for each 0.4 g/dL and glucose by 0.001 for each 0.27 g/dL (Hofmann et al., 1995).

Refractometry Refractometry is the most reliable way to measure specific gravity. This method is based on measurement of the refractive index, which is related to the weight of solutes per unit volume of urine. Several types of refractometers are available, which are very simple to use and require only one drop of urine. In some automated test strip readers, a refractometer is integrated in the instrument.

Osmolality

Relative density

Osmolality is the measure of solute concentration, defined as the number of osmoles (Osm) of solute per litre (L)  of solution (osmol/L). Osmolality is a measure of the osmoles of solute per kilogram of solvent (osmol/kg or Osm/kg). Measurements of the osmotic concentration of urine are considered more valid than specific gravity measurements. Osmolality is a function of the number of particles in solution and is usually measured through the effect of colligative properties on the freezing point of the solution. Numerous manual or fully automated osmometers are available commercially. Measurement of osmolality offers definite advantages over other methods. These include (a) no temperature correction is necessary, (b) only small volumes of urine are required, and (c) there is no interference from proteins or other macromolecules. However, high glucose concentrations contribute significantly to the measured osmolality (10 g/L of glucose = 55.5 mOsm) (Hofmann et al., 1995). (See Table 8.1.)

Effects of dilution may be measured by four methods: specific gravity, refractometry, osmolality, and dry chemistry (dipsticks).

Dry chemistry

Assessment of renal concentration ability The capacity of the kidneys to conserve water can be assessed by measuring the solute concentration of urine. After fluid deprivation of 18 hours or more, urine osmolality exceeding 850 mOsm/kg is considered normal. In cases of a complete antidiuretic hormone (ADH) deficiency or in nephrogenic diabetes insipidus (where the normal response to ADH is lacking), urine osmolality rarely exceeds 300 mOsm/kg.

Specific gravity Specific gravity is a measure of the content of solids in a solution. The measurement of urine specific gravity is related to osmolality.

Alternatively, specific gravity can be measured as a urine dipstick test. After intravenous administration of iodine-containing contrast media, extraordinarily high values (> 1.040) may be temporarily obtained.

Urine conductivity The conductivity can be used as a substitute for osmolality (reference range: 8–32 mS/cm). The conductivity measurement is related to the concentration of electrolytes in the urine. In contrast to osmolality and specific gravity, uncharged particles (e.g. glucose, urea) do not contribute to conductivity. There is a good agreement between conductivity, urine density, and osmolality (r = 0.8 or higher) (Hofmann et al., 1995). Conductivity correlates better

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assessment of the patient with renal disease Table 8.1  Serum and urine osmolality Serum osmolality Ref. values: 280–296 mOsm/kg

Urine osmolality Ref. values: 500–800 mOsm/kg

Clinical significance

Normal or increased Decreased Normal Normal or increased

Increased Decreased Decreased Decreased (without increased fluid intake) Increased

Volume deficit Volume excess Increased fluid intake or diuretics use Kidneys unable to concentrate urine or lack of ADH (diabetes insipidus) SIADH (syndrome of inappropriate ADH secretion)

Decreased

with osmolality than the creatinine concentration (r = 0.84 and 0.68, respectively) and therefore might serve as a better indicator of hydration status ((Hofmann et al., 1995; Hannemann-Pohl and Kampf, 1999).

Osmolar clearance and free-water clearance The quantitative ability of the kidney to excrete or retain water can be estimated by the calculation of osmolar and free-water clearance. Conceptually, the rate of water excreted in urine can be divided into two components. One contains all the urinary solutes and the amount of water required to obtain a solution iso-osmotic to plasma. This defines the osmolar clearance. The other component is made of solute-free water and is called free water clearance: CH2O = V – Uosm × V/Posm. This theoretical flow is positive when urine is hypotonic to plasma (water has been ‘added in excess’ to osmoles in urine) and negative when final urine is hypertonic to plasma (water has been ‘substracted in excess’ to osmoles to concentrate urine). The contribution of urea to water excretion and the use of urea concentration to calculate osmolality are detailed in Soroka et al. (1997).

transporter (Moe et al., 2000). This transporter is also expressed in the apical membrane of the collecting duct principal cells together with the amiloride sensitive sodium channel (ENaC) (Kim et al., 1998). In this segment, sodium reabsorption is regulated by the mineralocorticoid hormone aldosterone and coupled to K+ secretion. A global measurement of sodium absorption by the tubules is obtained by measuring the fractional excretion of sodium (FENa) which is expressed as the ratio of the sodium excretion rate to the sodium filtered load: where is the urinary concentration of sodium, the urine flow rate. In various clinical conditions, physicians are interested in knowing what proportion of sodium is being reabsorbed in the proximal and distal parts of the nephron. The measurement of sodium absorption in the proximal tubule is based on the assumption that lithium is absorbed to the same extent as sodium in this segment but is not handled in the distal parts of the nephron (Thomsen, 1990). Measurement of the urine sodium concentration provides information on the integrity of tubular reabsorptive function. The following equation can be used to calculate the fractional excretion of sodium: FE Na = 100 × U Na × PCrn / PNa × U Crn .

Fractional excretion of sodium



Sodium reabsorption occurs along the entire nephron by various mechanisms. In the proximal tubule, 60% of the filtered sodium is recovered from the initial urine by active transcellular movements and passive paracellular pathways. In the first mechanism, sodium enters the apical pole of proximal cells coupled to organic (glucose, amino acids, etc.) or inorganic (phosphate) solutes via specific sodium-dependent carriers or is exchanged for H+ ions. In all tubule segments, sodium is actively pumped out of the cells via the basolateral Na+/K+-ATPase. The passive paracellular sodium flux is due to osmotic water flow from lumen to blood that entrains sodium through the lateral intercellular space (Moe et al., 2000). The thick ascending limb reabsorbs about 30% of the sodium filtered at the glomerulus. Sodium is transported across the apical membranes via the Na+/K+/Cl2−-cotransporter and the Na+/H+exchanger. The K+-recycling at the apical membrane generates a lumen positive transepithelial voltage driving additional paracellular sodium reabsorption. Furosemide and bumetanide inhibit the Na+/K +/Cl 2−cotransporter and sodium absorption in the thick ascending limb. The distal convoluted tubule reabsorbs about 10% of the filtered sodium load via the apical thiazide-sensitive sodium chloride

A FENa < 1% suggests a prerenal cause, because volume depletion or a decrease in the effective circulating volume induce an increase in sodium reabsorption. A value > 2–3% is indicative of acute tubular necrosis or injury. In renal tract obstruction, values may be either > or < 1%. The value is lower in early disease, but with renal damage from the obstruction, the value becomes higher (Steiner, 1984).

References Aperia, A., Broberger, O., Herin, P., et al. (1983). Postnatal control of water and electrolyte homeostasis in pre-term and full-term infants. Acta Paediatr Scand Suppl, 305, 61–5. Clarke, B. L., Wynne, A. G., Wilson, D. M., et al. (1995). Osteomalacia associated with adult Fanconi’s syndrome: clinical and diagnostic features. Clin Endocrinol, 43, 479–90. Curtis, J. R. and Donovan, B. A. (1979). Assessment of renal concentrating ability. Br Med J, 1, 304–5. Davis, B. B. and Zenser, T. V. (1993). Evaluation of renal concentrating and diluting ability. Clin Lab Med, 13, 131–4. Davis, P. J. and Davis, F. B. (1987). Water excretion in the elderly. Endocrinol Metab Clin North Am, 16, 867–75. DeRubertis, F. R., Michelis, M.F., Beck, N., et al. (1971). ‘Essential’ hypernatremia due to ineffective osmotic and intact volume regulation of vasopressin secretion. J Clin Invest, 50, 97–111.

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Hannemann-Pohl, K. and Kampf, S. G. (1999). Automation of urine sediment examination: a comparison of the Sysmex UF-100 automated flow cytometer with routine manual diagnosis (microscopy,test strips, and bacterial culture). Clin Chem Lab Med, 37, 753–4. Hofmann, W., Edel, H. and Guder, W. G. (1995). A mathematical equation to differentiate overload proteinuria from tubulo-interstitial involvement in glomerular diseases. Clin Nephrol, 44, 28–31. Kim, G. H., Masilamani, S., Turner, R., et al. (1998). The thiazide-sensitive Na-Cl cotransporter is an aldosterone-induced protein. Proc Natl Acad Sci U S A, 95, 14552–7. Miles, B. E., Paton, A., and De Wardener, H. E. (1954). Maximum urine concentration. Br Med J, 16, 901–5. Moe, O. W., Berry, C. A., and Rector, F. C. J. (2000). Renal transport of glucose, aminoacids, sodium, chloride and water. In B. M. Brenner (ed.) The Kidney, pp. 375–415. Philadelphia, PA: W. B. Saunders Company. Monnens, L., Smulders, Y., van Lier, H., et al. (1981). DDAVP test for assessment of renal concentrating capacity in infants and children. Nephron, 29, 151–4. Morello, J. P. and Bichet, D. G. (2001). Nephrogenic diabetes insipidus. Annu Rev Physiol, 63, 607–30. Moses, A. M., Scheinman, S. J., and Schroeder, E. T. (1985). Antidiuretic and PGE2 responses to AVP and dDAVP in subjects

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with central and nephrogenic diabetes insipidus. Am J Physiol, 248, F354–F359. Nielsen, S., Frøkiaer, J., Marples, D., et al. (2002). Aquaporins in the kidney: from molecules to medicine. Physiol Rev, 82, 205–44. Radó, J. P., Marosi, J., Szende, L., et al. (1976). The antidiuretic action of 1-deamino-8-D-arginine vasopressin (DDAVP) in man. Int J Clin Pharmacol Biopharm, 13, 199–209. Rowe, J. W. Shock, N. W. and DeFronzo, R. A. (1976). The influence of age on the renal response to water deprivation in man. Nephron, 17, 270–8. Schrier, G. (2007). Diseases of the Kidney and Urinary Tract. Philadelphia, PA: Lippincott Williams & Wilkins. Soroka, S. D., Chayaraks, S., Cheema-Dhadli, S., et al. (1997). Minimum urine flow rate during water deprivation: importance of the nonurea versus total osmolality in the inner medulla. J Am Soc Nephrol, 8, 880–6. Steiner, R. (1984). Interpreting the fractional excretion of sodium. Am J Med, 77, 699–702. Thomsen, K. (1990). Lithium clearance as a measure of sodium and water delivery from the proximal tubules. Kidney Int Suppl, 28, S10–S16. Williams, T. D., Dunger, D. B., Lyon, C. C., et al. (1986). Antidiuretic effect and pharmacokinetics of oral 1-desamino-8-D-arginine vasopressin. 1. Studies in adults and children. J Clin Endocrinol Metab, 63, 129–32.

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Renal radiology: overview Michael J. Weston Brief introduction to radiology chapters Imaging of form and function is an integral part of modern medical practice essential for diagnosis, treatment, and monitoring. Moreover, image-guided biopsies, drainage procedures, and ablations have circumvented the need for many invasive surgical procedures. There are a multitude of different techniques available so their diversity makes an understanding of radiological practice essential for the clinicians who rely upon them. Chapters  10–16 will highlight the uses of plain radiography, fluoroscopy, ultrasound, computed tomography (CT), magnetic resonance imaging, radionuclide studies, and image-guided intervention. All imaging studies work best if a specific question is asked. This helps to choose both the best modality and protocol to answer the question. The clinical information given will often assist the interpretation of the findings. The more vague the indication for a scan, the less likely that useful information will be provided. Both the requesting clinician and the radiologist need to be clear how the scan result will alter management. Performing imaging procedures that will not alter the outcome is wasteful and unkind to the patient. It is important to understand the adverse effects of imaging tests. Some, such as the effects of ionizing radiation, are well described but often poorly understood. A  surprisingly high proportion of doctors who are not radiologists have been found in surveys to be ignorant of which radiological techniques involve ionizing radiation and are unable to quantify the relative risks involved. Adverse effects of contrast medium administration are particularly pertinent to the nephrologist. Other adverse effects are less appreciated, for example, the burden imposed by incidental findings discovered on tests done for other reasons. It is not unknown for patients to embark on a series of tests requested to clarify incidental imaging findings, causing them to suffer both psychological and physical harm, without, in the end, any benefit to their health. The clinician should remember the need to treat the patient and not the scan. An understanding of the difference between form and function is essential. For instance, dilation of the renal collecting system does not equate to obstruction. Ultrasound of the kidneys in someone who has a urinary conduit will often show hydronephrosis because an ultrasound cannot distinguish obstruction from chronic reflux. Renal function will clearly have a role in deciding if further imaging is needed. Isotope renography or a conduitogram are both potential next imaging steps. Similarly, pregnant women often show

physiological renal upper tract dilation so finding hydronephrosis will not distinguish pregnancy-related dilation from stone disease in someone with flank pain. Magnetic resonance urography can solve this problem whilst avoiding ionizing radiation to the fetus. Many radiological tests do combine some measure of function along with a depiction of anatomy. CT with intravenous contrast medium enhancement will give a broad indication of the kidney’s ability to excrete. This is not as reliable as isotope renography in assessing relative function, though both tests are contraindicated or unhelpful in renal failure. Positron emission tomography (PET) in its commonest manifestation looks at glucose metabolism and when superimposed on or combined with a CT scan done at the same time can clearly highlight areas of high metabolism—most commonly looking for metastases or lymph nodes involved in post-transplant lymphoproliferative disorder. An important task for the referring clinician is preparation and patient consent for the radiological test. Iodinated contrast agents are nephrotoxic; it is not enough to assure the radiologist of appropriate prior patient hydration, the patient should also give informed consent. This is more than just understanding the risk of worsening renal failure but also considering alternate ways of investigating the problem. Clearly, patient consent is even more of an issue when an image-guided biopsy is proposed. Proper consent requires input from both the clinician requesting the biopsy and the radiologist who will perform the procedure. Incorrect patient preparation may also prevent a scan from being done and waste an appointment slot. For instance, having breakfast just before attending for a PET/CT scan will mean the scan will not be done (because of the effect on glucose metabolism). Finally, the imaging history needs to be kept in mind. The answer to a problem may not lie in asking for a new imaging test but in a reappraisal of the previous imaging. The following chapters will show how to choose the right test, to recognize their limitations and risks, and to understand what the findings mean. The patient needs to be properly involved in the decision-making process so that they are prepared both mentally and physically for the procedure and its purpose. It is important for clinicians to establish a good relationship with the radiology department and the radiologists to ensure that the best service can be delivered. In summary: 1. Only request investigations if the result can be interpreted and used to alter management.

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renal radiology: overview

2. Choose the right test by consulting the radiology colleague in advance with a description of the problem.

6. Be aware of potential adverse effects.

3. Always review previous imaging.

8. Ensure that the patient is properly prepared for the investigation, for example, bowel preparation, full bladder, fasted for PET scans, and hydrated for contrast administration.

4. Involve the patient in the discussion and take proper consent. 5. Do not confuse anatomical and functional findings.

7. Treat the patient and not the scan.

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Ionizing radiation and radiation protection Jeannette Kathrin Kraft and Peter Howells Overview Ionizing radiation is a natural phenomenon and we are constantly exposed to small amounts of radiation from natural and artificial sources. The largest amount of man-made radiation we receive as a population is from medical imaging. Radiation exposure in medical imaging can be expressed as effective dose measured in millisieverts (mSv). As effective dose is difficult to measure in practice, indicators of dose such as dose area product (DAP) and dose length product (DLP) are commonly used. It is usual to compare the amount of radiation patients receive through imaging with natural background radiation (2.4 mSv per year). Ionizing radiation has potential adverse effects on the human body. It may cause acute effects due to cell death called tissue reactions or deterministic effects. Damage to the cell’s DNA may lead to random or stochastic effects such as cancer induction. Available data on the risk of inducing cancer are extrapolated from the high-dose exposures of atomic bomb survivors or radiation accidents. It is difficult to estimate cancer risk accurately at the low radiation doses commonly used in medical imaging. Therefore models such as the linear non-threshold model are used to extrapolate risk at low-level radiation exposure. As medical imaging has the potential for harm, the risk to the patient has to be managed and controlled by all heath care professionals. The Ionising Radiation (Medical Exposure) Regulations 2000 (IR(ME)R 2000) impose obligations on hospital authorities to minimize radiation doses to patients in the United Kingdom (Department of Health, 2000). This can be achieved through careful imaging and technique selection, justification, and optimization using the ALARA principle in keeping the dose ‘as low as reasonably achievable’ or practicable. Repeat studies should be avoided and diagnostic reference levels (DRLs) regularly audited. Both healthcare professionals and patients need to be educated about the risks and benefits of imaging. Employees working with ionizing radiation are regularly monitored. Benefit from medically indicated imaging almost always outweighs the risk especially in emergency situations such as assessment of trauma. Radiation should not be feared.

Introduction In 1895, the German physicist Wilhelm Conrad Röntgen discovered X-rays when he noted that a fluorescent screen began to glow while he was experimenting with a vacuum tube. Soon after, in 1896, Becquerel noted that photographic plates stored with uranium were

fogged, thereby discovering natural occurring radiation. By the early 1900s, X-rays were used widely in medical imaging. However, soon after their discovery adverse health effects became apparent and many early radiologists died of radiation-induced sickness and cancer (Berry, 1986). In 1915, the British Röntgen Ray Society adopted radiation protection recommendations, probably representing the first joint effort at radiation protection. In the 1920s, film badges were introduced for routine monitoring of personnel and in 1925 the first exposure limits were set. In the 1950s, the study of biological effects continued with the long-term follow-up of the subjects exposed to radiation from the two atomic bombs in Japan in 1945 and is still ongoing today (Richardson et al., 2009; Ozasa et al., 2012). Radiation accidents such as Chernobyl in 1986 and Fukushima in 2011 increased public awareness. After the advent of computed tomography (CT) in the 1970s, the use of ionizing radiation in medical imaging has steadily increased. Approximately 62  million CT examinations are performed each year in the United States and approximately 4 billion radiological examinations are performed worldwide (Brenner and Hall, 2007; United Nations Scientific Committee on the Effects of Atomic Radiation (UNSCEAR), 2011). The first direct studies of cancer risk in children who have undergone CT scanning were published in 2012 and 2013 (Pearce et al., 2012; Mathews et al., 2013). Although the risk for any individual may be small, the increasing radiation exposure to the population may be a health risk in the future (Brenner and Hall, 2007). However, there is still a lack of awareness of radiation exposure and associated risks among medical practitioners (Shiralkar et al., 2003).

Definition of ionizing radiation Radiation is a form of energy transmitted through space. There are many types including heat, light, radio waves, microwaves, X-rays, and gamma rays. The frequency of the wave describes its position in the electromagnetic spectrum. Low-energy radio waves with low frequencies are at one end of the electromagnetic spectrum; high-frequency, high-energy waves such as X-rays or gamma rays are at the other end. High-frequency waves that carry a large amount of energy can penetrate material and transfer energy to an atom that may cause displacement of an electron from its orbit around the nucleus. This process is called ionization. Ionizing radiation can be electromagnetic radiation such as X-rays or gamma rays or particulate radiation such as alpha particles, beta particles,

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or neutrons. It can be produced in a generator such as an X-ray machine or come from radioactive material as used in nuclear medicine or radiotherapy.

ionizing radiation and radiation protection

Measuring radiation

constituents of the soil. The average annual effective dose from natural sources in the world is estimated to be 2.4 mSv (UNSCEAR, 2010). When describing radiation exposure from medical examinations we often compare the dose received from that examination to natural background radiation.

Absorbed dose

Man-made radiation

When ionizing radiation passes through tissue it deposits energy causing ionization and excitation of the matter. The amount of energy deposited divided by the mass of tissue exposed is called the ‘absorbed dose’. The unit of absorbed dose is joule per kilogram (J/ kg) but for convenience it has been given a special name: gray (Gy). This is an easy quantity to measure but is not a useful indicator of long-term risk.

Radiation exposure may be increased in several ways by human activity and technological processes, for example, air travel (increased levels of cosmic radiation), nuclear testing, and nuclear accidents such as Chernobyl in 1986, nuclear power production, occupational exposure, consumer products such as electronic equipment, fire alarms, and luminous numbers on watches and clocks. However, the largest amount of man-made radiation exposure is through medical imaging procedures.

Equivalent dose Different kinds of radiation cause different biological effects for the same amount of energy deposited. For instance, neutrons or alpha particles cause more damage per unit absorbed dose than X-rays or gamma rays. To allow for this, a quality factor for radiation was introduced leading to the expression ‘equivalent dose’. For X-rays and gamma rays the absorbed dose equals the equivalent dose. The unit of equivalent dose is also J/kg but to avoid confusion it has been given a different name: sievert (Sv).

Effective dose In addition, different tissues have variable sensitivity to radiation explained by cell turnover rate and tissue-specific characteristics. To account for this a tissue weighting factor was added to modify equivalent dose to ‘effective dose’. The unit for the effective dose stays the same: Sv. Doses in medical imaging and radiation protection are usually organ specific and refer to effective doses. As they are often small, most are expressed as millisievert (mSv).

Indicators of dose In practice, it is not possible to measure effective dose directly. Within a procedure other indicators of dose are generally used, such as the DAP in radiography and fluoroscopy or DLP in CT. DAP measures the dose in air multiplied by the X-ray beam area at a point close to the X-ray unit. This quantity is a constant for any distance from the X-ray tube. DLP measures the dose per rotation of the CT scanner multiplied by the length of the patient included in the scan. Effective dose can be inferred from each of these values but only with knowledge of other parameters.

Sources of ionizing radiation Radiation is a natural phenomenon which we are exposed to continuously from natural and artificial man-made sources. Low levels of radiation cannot be seen or felt, so people are not usually aware of it.

Natural background radiation Natural background radiation is ubiquitous and includes high-energy cosmic rays, radioactive nuclides from the earth’s crust such as uranium and thorium, and radon gas. The annual effective dose from natural background radiation varies substantially throughout the world due to variation in altitude and the

Medical exposures Medical exposures account for 98% of the contribution from all artificial sources and are now the second largest contributor to the population dose worldwide, representing approximately 20% of the total from all sources. It is estimated that from 1997 to 2007, approximately 3.6 billion medical radiation procedures were performed annually worldwide, an increase of 40% on the last decade (UNSCEAR, 2010). This increase is demonstrated particularly for high-dose examinations, such as CT scanning, the use of which is increasing rapidly. Medical radiation procedures were 65 times more frequent in developed countries compared to underdeveloped countries reflecting an imbalance of radiation exposure and healthcare provision. In several countries medical radiation exposure is now larger than exposure from natural sources (UNSCEAR, 2010).

Potential health effects of ionizing radiation Biological effects As previously described, high-energy ionizing radiation causes displacement of an electron from its orbit around the nucleus. This electron can cause direct damage when it hits a strand of DNA or indirect damage when the electron reacts with surrounding water molecules producing free radicals or ions. A free radical is a molecule with unpaired electrons that in turn avidly undergoes chemical reactions with surrounding molecules such as DNA. Damage to a cell’s DNA may have several consequences. Single-strand breaks may simply be repaired without detrimental effects to the cell. However, a double-strand break can be repaired incorrectly which may have hereditary effects if it occurs in gonadal cells, or produce malignancy through activation of oncogenes (UNSCEAR, 2011). The amount of damage to an individual or organ depends on the type and amount of radiation and therefore the energy transferred to the tissues, the duration of exposure, the distance from the radiation source, and whether the exposure was continuous or intermittent. The probability of a tissue or organ suffering effects from ionizing radiation is called radiosensitivity. Tissues with high rates of mitosis and undifferentiated cells are the most sensitive. Therefore, bone marrow cells, germ cells, lens cells, and bowel mucosal cells are highly sensitive whereas bone and neural cells are relatively less radiosensitive.

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Tissue reactions (deterministic effects) Ionizing radiation may cause acute effects which are a result of cell death. They require a threshold dose of manifestation which is not the same for each individual or tissue. These are also known as deterministic or non-stochastic effects and are usually seen at high radiation doses such radiotherapy or radiation accidents. Because a threshold can be identified, radiation protection measures and occupational dose limits can eliminate such effects for workers. Deterministic effects include skin erythema, ulceration and burns, bone marrow depression, cataracts, acute radiation sickness, sterility, and fetal death (Mettler and Upton, 2008b). Healing may occur but permanent damage to tissues may result in fibrosis or necrosis. Although relatively rare, such effects have been observed from medical imaging such as CT scanning and interventional procedures (Shope, 1996; De Olazo Banaag and Carter, 2005; Bogdanich, 2010).

Random (stochastic) effects Damage to a cell’s DNA increases the probability of cancer induction. If germ cells are affected, hereditary effects may be observed. These effects are random and long term and are also known as ‘stochastic’ or ‘non-deterministic effects’. There are two important studies that investigated the direct risk of cancer induction from medical imaging. These have shown a small increased risk of leukaemia and brain cancer after cranial CT in children (Pearce et al., 2012; Mathews et al., 2013). Lifespan studies on atomic bomb survivors indicate increased risk for development of leukaemia and solid cancers including stomach, lung, liver, colon, breast, gallbladder, oesophagus, bladder, and ovary after exposure to ionizing radiation (Richardson et al., 2009; Ozasa et al., 2012).

Estimating risk from medical examinations Cancer risk At the low radiation doses (effective doses < 100–200 mSv) commonly used in medical imaging, statistical limitations make it difficult to estimate cancer risk accurately. Very large numbers of patients would need to be followed up over a lifetime to quantify risk at low doses of radiation exposure (Brenner et al., 2003). Most of the available data on cancer induction was therefore extrapolated from high radiation doses received over a short time interval following the survivors of the detonation of the atomic bombs in 1945 and radiation accidents. The assumption that radiation risk continues in a linear fashion when extrapolated from high to low doses is termed the ‘linear non-threshold model’ (Brenner et al., 2003; National Academy of Sciences, 2006). At present this model is the most widely accepted theory to describe radiation effects at low doses. It assumes that no level of radiation exposure is safe and that even the smallest dose of radiation has a potential to cause harm. Data from atomic bomb survivors and radiation accidents have been used to develop risk models and theories. Such models predict that for a radiation exposure of 0.1 Sv above background radiation, 1 person in 100 would be expected to develop cancer. Similarly, for a radiation exposure of 10 mSv, which is approximately the effective dose that would be received from a CT scan of the abdomen, 1 in 1000 people could be expected to develop cancer (National Academy of Sciences, 2006). At present it is not possible to determine whether a cancer was induced by radiation or any other cause.

In a population, radiation effects especially from low-dose radiation can be obscured by the large number of non-radiation cancers occurring. One in three people will develop cancer from natural causes and one in six people are predicted to die of such cancer (National Academy of Sciences 2006). There is also a considerable latent period between radiation exposure and the induction of cancer. The mean latent period for leukaemia is 7–10  years, 10–15  years for bone tumours, and approximately 20 years for most solid tumours (Mettler and Upton, 2008a). This time lag of cancer induction explains the higher risk for younger patients and the relatively low risk for elderly patients.

Hereditary defects Excess hereditary effects due to radiation exposure have not been demonstrated in humans and were not detected in atomic bomb survivors of Hiroshima and Nagasaki (National Academy of Sciences, 2006; UNSCEAR, 2011). Minor effects are likely to be so small that current studies are unable to differentiate them from naturally occurring mutational effects. However, radiation-induced mutations can theoretically occur in reproductive cells (egg and sperm) resulting in hereditable diseases. Such effects have been previously demonstrated in animal studies, mainly mice (UNSCEAR, 2001).

Risks to the fetus In utero radiation of the fetus causes great anxiety. Children and the fetus are more sensitive to ionizing radiation than adults because of the larger number of dividing cells and the longer lifespan available to develop cancer. Radiation exposure very early on in a pre-implantation stage could lead to an all-or-nothing phenomenon of spontaneous abortion or normal pregnancy. High doses delivered during organogenesis may cause central nervous system abnormalities inducing microcephaly or mental retardation. Malformations are thought to have a threshold of 100–200 mGy absorbed dose which is rarely reached in medical diagnostic imaging. During the third trimester, the organs develop and the risk of congenital malformations is low but there is still a small risk of inducing cancer or leukaemia. However, diagnostic radiological procedures should not normally exceed 50–100 mGy and with such an exposure there is a 99% chance that the child will not develop cancer or leukaemia (Brent, 1984; International Commission on Radiological Protection, 2000).

Absolute risk to a patient An estimation of the subject’s risk from a medical procedure can be made by applying the average effective dose of the procedure and taking the age at the time of exposure, the organ irradiated, and the sex into account (Wall et al., 2011). As risk from radiation exposure is cumulative, radiation doses from several procedures are added to calculate risk. Because of the benefits of imaging in diagnosing illness and injury, a medically indicated imaging study will almost always outweigh the risk associated with it. This is especially apparent when evaluating risks associated with common daily activities such as driving a car. Those activities carry a much higher risk of dying than the cancer risk of 1:1000 associated with a single CT scan of abdomen delivering a dose of 10 mSv (Health Protection Agency, 2001; National Academy of Sciences, 2006; National Safety Council, 2010). Therefore, when assessing radiation exposure at

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Table 10.1  Typical effective doses to adults from selected radiological procedures (National Radiological Protection Board, 1994; Hart et al., 2012) Radiograph

3rd quartile DAP per radiograph (Gy × cm2)

Effective dose estimated from NRPB coefficients (mSv)

Chest PA

0.3

0.03

Abdomen AP

4.4

1.1

IVU

14

2.5

AP = anterior–posterior; DAP = dose area product; NRPB = National Radiological Protection Board; PA = posterior–anterior.

very low radiation doses such as, for instance, for single exposure of a chest radiograph (0.01 mSv), the risk of radiation-induced cancer persists but in practice becomes negligible (Health Protection Agency, 2001).

Quantifying radiation in medical imaging When describing radiation doses received by patients the comparison is with natural background radiation (2.4 mSv per year). Effective doses for most radiological examinations have been published (Mettler et al., 2008). Plain film radiographic examinations vary by a large factor from 0.001 mSv to 10 mSv. CT examinations have relatively high average effective doses between 2 and 20 mSv and interventional procedures can give effective doses between 5 and 70 mSv (Mettler et  al., 2008). For comparison, a round-trip flight from London to New  York will result in a radiation exposure from increased cosmic radiation of about 0.1 mSv (Brenner et al., 2003). From time to time, the Health Protection Agency publishes the results of national patient dose surveys performed in the United Kingdom, for radiological examinations and for CT (Shrimpton et  al., 2005; Hart et  al., 2012). The doses are typically shown as entrance dose, DAP, or DLP for CT, but effective doses can be estimated from tables published by the National Radiation Protection Board (1994). Doses of selected examinations are shown in Tables 10.1 and 10.2.

Managing risk to patients Even though uncertainties remain about the absolute risk of low-dose radiation, the risk is not entirely negligible. Concerns Table 10.2  Typical effective doses to adults from selected CT procedures (Shrimpton et al., 2005; Hart et al., 2012) CT scan

Reference DLP per scan (Gy × cm)

Effective dose estimated from NRPB coefficients (mSv)

Chest (high resolution)

170

2.4

Abdomen

470

7.1

Chest, abdomen, and pelvis

940

14.1

DLP = dose length product; NRPB = National Radiological Protection Board.

ionizing radiation and radiation protection

about radiation and induction of cancer are likely to be in the public focus for the foreseeable future. It is therefore the responsibility of all healthcare professionals to manage and control the risk to protect patients. This point is reinforced by government authority through legislation.

Ionising Radiation (Medical Exposure) Regulations 2000 The IR(ME)R 2000 impose obligations on hospital authorities and clinicians to minimize radiation doses to patients (Department of Health, 2000). The three principles governing patient protection in medical imaging are justification, optimization, and DLRs.

Imaging selection Not all imaging modalities use harmful radiation. The same information might be acquired with an imaging modality that does not use ionizing radiation such as sonography or magnetic resonance imaging (MRI). Sometimes it can be beneficial just to observe the patient for a time period for symptoms to unmask. This is part of a decision process to justify the examination.

Justification For each examination performed there should be careful risk/benefit analysis. It is the responsibility of the clinician as the ‘Referrer’ (term under IRMER) to supply sufficient information for the radiology professional as the ‘Practitioner’ to decide whether the examination is justified. The iRefer: Making the Best Use of Clinical Radiology publication by The Royal College of Radiologists can help clinicians to select appropriate imaging (The Royal College of Radiologists, 2012). For most examinations, the radiographer or ‘Operator’ working within departmental protocols will decide whether to proceed and perform the examination. This is not unique to the risk assessment associated with ionizing radiation. Similar consideration are made, for instance, when considering the risk of bleeding or infection from a diagnostic biopsy, the risk of general anaesthesia, or simply whether it is safe for the patient to be transferred to a different department.

Technique selection and optimization If imaging using ionizing radiation is necessary, the radiology professional will aim to use the lowest possible radiation dose giving an accurate diagnosis. This attempt at optimization refers to the ALARA principle of reducing the radiation dose to ‘as much as reasonably achievable or practicable’. This includes only scanning the area indicated (e.g. renal area rather than the whole abdomen), working under local protocols, adjusting scanning parameters for children, avoiding multiple phase scanning (e.g. non-contrast, arterial, and portal venous phase CT scans), and controlling scan time in fluoroscopy. Optimization also includes quality assurance programmes, regular audits, and planned replacement of elderly or outdated equipment.

Repeat studies Repeat studies can be avoided. Patients with chronic or long-term problems such as young patients, for example, with nephrolithiasis or kidney transplants should be carefully managed as radiation dose is thought to be cumulative over time. These patients should not be repeatedly subjected to CT scans each time they present.

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Perhaps the clinical question can be answered with an ultrasound scan, reserving CT scanning for problem-solving. Discussion with the radiology professional should be encouraged.

Diagnostic reference levels Under IR(ME)R 2000, DRLs should be established for each standard examination and type of equipment at each imaging department. These levels indicate the radiation dose a typical patient is likely to receive from a standard radiological examination. However, DRLs are not strictly dose limits as the radiation dose received by a patient depends on factors such as their size, the technique selected, and, for fluoroscopic examinations, the screening time. DRLs may be seen as action thresholds and are regularly audited. If they are regularly exceeded, further optimization of technique or particular justification is needed. Radiation doses from interventional procedures can be high, particularly to the skin where the same area is persistently irradiated during an extended procedure. For such procedures, similar action thresholds, in terms of DAP or total screening time, can be used to inform the radiology professional of the possibility of tissue reactions and possible injury that would be likely to result if the same body area is continuously irradiated.

Communication and education Communication between the clinician and the radiology professional is essential and will help to tailor a radiological investigation to the patient’s needs, reducing the amount of radiation. Educating clinicians and patients about the risk of radiation exposure from medical procedures is the aim of campaigns such as ‘Image Gently’ for children and ‘Image Wisely’ for adults (Alliance for Radiation Safety in Pediatric Image Imaging, n.d.; American College of Radiology, n.d.).

Minimizing risk to staff Principles Any measures that reduce patient dose also reduce potential occupational dose to staff. During fluoroscopy, dose can be reduced by reducing the beam size through collimation, using pulsed fluoroscopy and last image hold instead of continuous screening, and by keeping exposure times as short as possible. Although the operator may stay away from the primary beam directed at the patient, a significant dose from scattered radiation is delivered to the operator when performing fluoroscopic examinations in theatre. The inverse square law demonstrates that doubling the distance to the source reduces the dose by a factor of four. Therefore stepping back during exposure or using a remote control will significantly reduce staff doses. Structural shielding and protective clothing such as aprons, thyroid shields, and goggles reduce the dose received by staff. Local rules and procedures are in place to protect staff and should be regularly reinforced through continuous education and training.

Dose monitoring Usually, employees working with ionizing radiation are regularly monitored. The International Commission on Radiological Protection has recommended dose limits for staff working with ionizing radiation and these are reiterated in radiation protection

legislation in the United Kingdom (UK Government, 1999). For employees over 18 years of age the annual limit on effective dose is 20 mSv, for trainees and employees under 18 years 6 mSv, and 1 mSv per year for the general public (UK Government ,1999; Heron et al., 2010). Additional annual limits on dose are set at 500 mSv equivalent dose for extremities, such as the fingers and hands of those involved in interventions, and for each 1  cm2 of skin. An annual limit on dose to the lens of the eye is set at 150 mSv, but is under review at the time of writing, with the expectation that it will be reduced to 20 mSv. In practice, occupational doses are constrained below these annual limits with a requirement to ‘classify’ radiation workers whose doses are expected, following risk assessment of their work, to exceed 3/10 of any annual limit.

Conclusion Medical imaging using ionizing radiation is invaluable in evaluating the sick patient. Radiation should not be feared, especially in emergency situations when there is a clear benefit from scanning. Radiation cannot be entirely avoided but it can be managed by following the ALARA principle. Medical exposures should be justified, duplication of scans should be avoided, and the lowest radiation dose consistent with the diagnostic objective should always be used.

References Alliance for Radiation Safety in Pediatric Image Imaging (n.d.). Image Gently. [Online] American College of Radiology (n.d.). Image Wisely. [Online] . Berry, R. J. (1986). The radiologist as guinea pig: radiation hazards to man as demonstrated in early radiologists and their patients. J R Soc Med, 79, 506–9. Bogdanich, W. (2010). After stroke scans, patients face serious health risks. N Y Times, 31 July. Brenner, D. J., Doll, R., Goodhead, D. T., et al. (2003). Cancer risks attributable to low doses of ionising radiation: assessing what we really know. Proc Natl Acad Sci USA, 100(24), 13761–6. Brenner, D. J. and Hall, E. J. (2007). Computed tomography—an increasing source of radiation exposure. N Engl J Med, 357, 2277–84. Brent, R. L. (1984). The effects of embryonic and fetal exposure to x-ray, microwave and ultrasound. Clin Obstet Gynecol, 26(2), 484–510. De Olazo Banaag, L. and Carter, M. L. (2005). Radionecrosis induced by cardiac imaging procedures: a case study of a 66-year-old diabetic male with several comorbidities. J Invasive Cardiol, 20(8), E2336. Department of Health (2000). The Ionising Radiation (Medical Exposure) Regulations 2000 (together with notes on good practice). London: Department of Health. Hart, D., Hillier, M. C., and Shrimpton, P. C. (2012). Doses to Patients from Radiographic and Fluoroscopic X-Ray Imaging Procedures in the UK-2010 Review. HPA-CRCE-034. Chilton: Health Protection Agency.

Health Protection Agency (2001). National Radiation Protection Board. X-rays How safe are they? [Patient information leaflet] Heron, J. L., Padovani, R., Smith, I., et al. (2010). Radiation protection of medical staff. Eur J Radiol, 76, 20–3. International Commission on Radiological Protection (2000). Pregnancy and medical radiation. ICRP Publication 84. Ann ICRP, 30(1). Mathews, J. D., Forsythe, A. V., Brady, Z., et al. (2013). Cancer risk in 680,000 people exposed to computed tomography scans in childhood

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or adolescence: data linkage study of 11 million Australians. BMJ, 346, f2360. Mettler, F. A., Huda, W., Yoshizumi, T. T., et al. (2008). Effective doses in radiology and diagnostic nuclear medicine: a catalogue. Radiology, 248(1), 254–3. Mettler, F. A. and Upton, A. C. (2008a). Cancer induction and dose-response models. In Medical Effects of Ionising Radiation (3rd ed.), pp. 71–116. Philadelphia, PA: Saunders Elsevier. Mettler, F. A. and Upton, A. C. (2008b). Deterministic effects of radiation. In Medical effects of Ionising Radiation (3rd ed.), pp. 285–388. Philadelphia, PA: Saunders Elsevier. National Academy of Sciences. (2006). Health Risk from Exposure to Low Levels of Ionising Radiation: BEIR VII Phase 2. [Free Executive Summary] [Online] National Radiological Protection Board (1994). Estimation of Effective Dose in Diagnostic Radiology from Entrance Surface Dose and Dose-Area Product Measurements. NRPB-R262. London: National Radiological Protection Board. National Safety Council (2010). Injury Fact. Lifetime Odds of Death for Selected Causes, United States 2006. [Online] Ozasa, K., Shimizu, Y., Suyama, A., et al. (2012). Studies of the mortality of atomic bomb survivors, report14, 1950–2003: an overview of cancer and non-cancer diseases. Radiat Res, 177, 229–3. Pearce, M. S., Salotti, J. A., Little, M. P., et al. (2012). Radiation exposure from CT scans in childhood and subsequent risk of leukaemia and brain tumours: a retrospective cohort study. Lancet, 380(9840), 499–505. Richardson, D., Sugiyama, H., Nishi, N., et al. (2009). Ionizing radiation and leukemia mortality among Japanese atomic bomb survivors, 1950–2000. Radiat Res, 172, 368–82.

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Shiralkar, S., Rennie, A., Snow, M., et al. (2003). Doctors’ knowledge of radiation exposure: questionnaire study. BMJ, 327(7411), 371–2. Shope, T. B. (1996). Radiation-induced skin injury from fluoroscopy. RadioGraphics, 16, 1195–9. Shrimpton, P. C., Hillier, M. C., Lewis, M. A., et al. (2005). Doses from Computed Tomography (CT) Examinations in the UK—2003 Review. NRPB-W67. Chilton: National Radiological Protection Board. The Royal College of Radiologists (2012). iRefer: Making the Best Use of Clinical Radiology. London: The Royal College of Radiologists.

UK Government (1999). The Ionising Radiations Regulations 1999. SI 1999 No. 3232. London: Her Majesty’s Stationery Office.

United Nations Scientific Committee on the Effects of Atomic Radiation 2001 (2001). Report: Hereditary Effects of Radiation, Annex. New York: UNSCEAR. United Nations Scientific Committee on the Effects of Atomic Radiation 2008 (2010). Report: Sources of Ionising Radiation, Annex A: Medical Radiation Exposures. New York: UNSCEAR. United Nations Scientific Committee on the Effects of Atomic Radiation 2010 (2011). Report: Summery of Low Dose Radiation Effects on Health. New York: UNSCEAR. Wall, B. F., Haylock, R., Jansen, J. T. M., et al. (2011). Radiation Risks from Medical X-Ray Examinations as a Function of Age and Sex of the Patient. HPA-CRCE-028. Chilton: Health Protection Agency.

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Plain radiography, excretion radiography, and contrast radiography Akira Kawashima and Andrew J. LeRoy Introduction Radiology is an essential component in the evaluation of the urinary tract. For decades, excretion radiography following intravenous (IV) administration of iodinated contrast media (also referred to as intravenous urography (IVU), intravenous pyelography (IVP), and excretory urography (EXU)), has been the primary imaging modality of the urinary tracts. In recent years, other imaging modalities such as ultrasonography (US), computed tomography (CT), and magnetic resonance imaging (MRI) have superseded excretion radiography. However, these cross-sectioned imaging examinations also have their limitations. Plain radiography, contrast radiography, and EXU still remain important in the diagnosis of some urinary tract diseases (Table 11.1). Plain radiography is both a primary abdominal examination technique and an essential initial component of subsequent excretion and contrast radiographic studies. Contrast radiography is obtained following direct injection of contrast material into the urinary tracts in an antegrade or retrograde fashion. The urographic imaging sequence with IV contrast material or direct contrast injection is designed to optimize depiction of specific portions of the urinary tract during maximal contrast opacification, and a tailored urographic study may provide diagnostic detail beyond the current capabilities of other imaging modalities (Hattery et al., 1988; Banner, 2001; Dyer et al., 2001) This chapter will outline basic uroradiologic imaging with each subsection providing information on technique, indications, complications, and normal anatomical findings.

image prior to IV or direct contrast injection for conventional urography. This scout radiograph helps establish appropriate radiographic exposure technique factors, evaluate the patient’s positioning, and confirms that residual oral contrast material from previous studies has been eliminated from the gut. The indications for the examinations and patient risk factors should be reviewed carefully. A  menstrual history should be obtained in women of childbearing age. Radiation exposure should always be limited appropriately. The presence of a fetal skeleton on the scout image justifies postponing the planned contrast study. A  scout image is imperative prior to contrast studies because stones and calcifications can be obscured by contrast material in the urinary tract on subsequent images.

Anatomical landmarks Several abdominal and pelvic organs can be identified on plain radiographic images. Organs such as the liver, spleen, kidneys, and bladder can be outlined on the image based on the contrasting densities of the organs themselves compared to surrounding retroperitoneal or mesenteric fat. The outline of the kidneys is helpful for assessing renal size, focal cortical scarring or mass, and for delineating intrarenal calcifications. The psoas muscle margins usually can be seen on the plain radiograph (see Fig. 11.1), but may be obscured by retroperitoneal fluid or mass. Plain films can demonstrate abnormalities of the bowel gas pattern (e.g. ileus, bowel obstruction, and faecal impaction) and also allow for evaluation of skeletal anomalies, such as sacral agenesis and spinal dysraphism.

Plain radiography

Excretion radiography

Plain radiography of the abdomen and pelvis is most often referred to as a KUB (Kidneys, Ureters, and Bladder) film or an abdominal ‘flat plate’ (Fig. 11.1), and can provide important information about radiopaque urinary calculi (Figs 11.2 and 11.3), calcifications, renal masses (Fig. 11.2), and renal size and contour (Dyer et  al., 1998; Barbaric and Pollack, 2000). A  combination of projection radiographs and non-contrast coronal tomography is valuable in outlining the kidneys and visualizing calculi. An anteroposterior (AP) projection radiograph of the abdomen in supine position may also be referred to as a scout (preliminary)

Excretion radiography is frequently used as a non-invasive screening procedure for the entire urinary tract because it provides both anatomic and functional information (Friedenberg and Harris, 2000).

Patient preparation A thorough pre-examination ‘bowel prep’ with a mild laxative (e.g. 10 ounces (28 g) of magnesium citrate solution) the night before the procedure is advisable to rid the colon of stool, which may interfere with clear visualization of the urinary tract. Withholding of fluid

Table 11.1  Glossary of terminology in uroradiography Terms (abbreviation)

Definitions

Plain abdominal radiography, KUB, plain film, conventional radiograph, flat plate, scout

Film-screen or digital projection radiograph without use of iodinated contrast medium

Excretion radiography, excretory urography (EXU), intravenous Imaging of the kidneys and urinary tracts before and after the IV administration of iodinated urography (IVU), intravenous pyelography (IVP) contrast medium Contrast radiography Retrograde pyelography

Imaging of the upper urinary tracts before and after the direct retrograde injection of contrast medium

Antegrade pyelography

Imaging of the upper urinary tracts before and after the direct antegrade injection of contrast media with percutaneous needle puncture of a calyx

Cystography Static cystography

Imaging of the bladder before, during and after the direct contrast administration either in a retrograde fashion through a transurethral Foley catheter or in an antegrade manner through a suprapubic tube

Voiding cystourethrography (VCUG)

Imaging of the bladder and urethra during micturition after the direct administration of contrast medium into the bladder. The procedure is monitored under fluoroscopy and recorded with spot films or video recording.

Retrograde urethrography (RUG) Loopography ileal conduit study

Opacification of the ileal loop before, during and after the direct injection of contrast medium with reflux of into the upper urinary tracts. The procedure is monitored under fluoroscopy and recorded with spot and overhead films.

Pouchgraphy

(A)

(B)

(C)

Fig. 11.1  Plain radiography and excretion radiography. (A) Plain radiograph of the abdomen and pelvis reveals normal contour of the kidneys and the medial margin of the psoas muscles (arrows). L = liver; LK = left kidney; RK = right kidney; UB = urinary bladder; UT = uterus. (B) Magnified view of the left kidney with abdominal compression 8 minutes after IV contrast material administration. Calyx (white arrows), infundibulum (I), renal pelvis (RP), proximal ureter (U). The calyx which project posteriorly is seen en face (white arrowhead). Normal fold of the ureter at the ureteropelvic junction (black arrow). (C) Abdominal radiograph after release of ureteral compression reveals normal ureters and urinary bladder (UB). The left ureter at the ureteropelvic junction (arrow) is better distended and straight. UT = uterus.

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assessment of the patient with renal disease demonstrate the calyces, pelves, and proximal ureters (pyelographic phase) (Fig. 11.1). The ureters are generally well visualized on the 10-minute radiograph after release of external compression. Films of the bladder (often including a post-void film) conclude the examination.

Diuretic excretion radiography

Fig. 11.2  Patient with renal stones and renal cell carcinoma. Plain radiograph demonstrates bilateral renal calculi (short arrows) with the largest over the left renal hilum (arrowhead). A round mass is projected over the lower pole of the left kidney (long arrow).

and solid food overnight improves renal concentration and excretion of the contrast medium. An empty stomach also decreases the possibility of aspiration of solid food by a patient vomiting after IV contrast administration.

Filming procedure A plain abdominal radiograph prior to the administration of IV contrast media is essential. After contrast medium is injected, lower abdominal compression is applied anteriorly. Approximately 2–3 minutes after the contrast material has been administered, nephrotomograms (usually three) are obtained to visualize the renal parenchyma (nephrographic phase) (Hattery et  al., 1988). Radiographs obtained 8–10 minutes after the contrast injection

(A)

(B)

This modification of excretion radiography is reserved for those patients suspected of having volume-dependent hydronephrosis in whom the initial (dehydrated) excretion radiography revealed no obstruction. With a brisk diuresis after furosemide injection (Lasix®, 20 mg IV), a borderline ureteropelvic junction or ureteral narrowing may become inadequate to allow free flow of the increased urine volume and may reveal in hydronephrosis and even reproduce or worsen the pain associated with obstruction.

Contrast media IV iodinated contrast material is excreted by glomerular filtration with little or no tubular secretion, with subsequent concentration in the tubules and collecting ducts and then progressive opacification of the urinary tract. Urine concentrations are determined by the dose administered, the glomerular filtration rate, and the renal tubular function. As the urine is concentrated in the renal collecting ducts, the relative concentration of the contrast media is enhanced 50–100-fold.

Contrast dosage Contrast medium can be administered either as a bolus injection or as a drip infusion. The bolus injection allows for a rapid and dense nephrogram when images are obtained 2–3 minutes after the injection. In the average-size adult, 60–100 mL of contrast medium is usually administered.

(C)

Fig. 11.3  Patient with renal stones and transitional cell carcinoma. (A) Magnified view of plain radiograph demonstrates two adjacent stones projected over the lower pole of the left kidney. (B) Excretory radiograph shows the left renal stones in the lower intrarenal collecting system. Note irregular narrowing of the left renal pelvis (arrow) with associated caliectasis. Kidneys are congenitally malrotated bilaterally. (C) Magnified view of antegrade pyelogram following a percutaneous needle puncture of a lower pole calyx shows a large filling defect in the mid to upper left renal pelvis with urothelial irregularity (arrows), characteristics of urothelial tumour. The two stones (arrowheads) lie in the lower left collecting system.

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Adverse effects The majority of adverse effects of iodinated contrast media are mild or moderate and only observation, reassurance, and support are required. However, the more severe adverse effects are often preceded by mild or moderate symptoms or prodrome. Reactions to contrast agents usually occur within 20 minutes following IV injection. The most threatening contrast reactions are those unanticipated events that result in sudden compromise of critical body functions, particularly systemic anaphylactic reactions or profound cardiovascular collapse. The widely used newer non-ionic, low-osmolar contrast media (LOCM) offer a slightly more comfortable, safer examination to patients who are sensitive to the older high-osmolar contrast media (HOCM) and produce fewer adverse effects. The risk of rare life-threatening reactions is not, however, eliminated by the use of these LOCM (Palmer, 1988; Katayama et  al., 1990). Appropriate training and vigilance by healthcare workers are therefore necessary in clinical areas where contrast media are administered. Anaphylactoid (idiosyncratic) reactions are generally classified as (1) mild, (2) moderate, and (3) severe (Cohan et al., 2010). Minor side effects, do not usually require therapy. Nausea and vomiting are the most common adverse reactions. Other minor reactions include rash, itching, hives (less than four), swelling, headache, dizziness, shaking, nasal congestion, pallor, flushing, chills, sweats, and anxiety. These symptoms are usually self-limiting, but observation is required to confirm resolution and lack of progression. Only supportive treatment is necessary. Patient reassurance is usually helpful. Moderate reactions include tachycardia, bradycardia, hypertension, mild hypotension, vasovagal reactions, dyspnoea, bronchospasm, wheezing, and pronounced cutaneous reaction such as extensive hives (four or more) or diffuse erythema. The observed clinical signs and symptoms of moderate reactions should be considered as indications for immediate treatment (Table 11.2). These situations require close, careful observation of possible progression into a life-threatening event. Patients with moderate symptoms such as systemic urticaria or facial oedema are treated with antihistamines and subcutaneous epinephrine. Severe types of reactions include laryngeal oedema, and/or, severe hypotension, require small doses of 1:10,000 dilution of epinephrine intramuscularly. The most severe reactions include convulsions, arrhythmias, unresponsiveness, and cardiopulmonary arrest. Prophylactic corticosteroids are strongly recommended for ‘at-risk’ patients who require contrast media (Lasser, 1988; Greenberger and Patterson, 1991). They should also receive LOCM. However, no regimen has eliminated repeat reactions completely. Pre-testing is not predictive, and may itself be dangerous, so is not recommended (Yamaguchi et al., 1991). Alternative means of imaging to obtain the required information should be considered before embarking on this potentially dangerous path.

Contrast-induced acute kidney injury (See Section 11.) IV contrast media may aggravate pre-existing renal dysfunction in a small percentage of patients. The mechanism is not known. There is no standard definition for reporting contrast media-induced nephrotoxicity. The definition of ‘significant’ changes varies considerably among studies. Acute contrast-induced nephrotoxicity has been defined as an increase in the baseline creatinine values of 20– 50% or an absolute increase from 0.5 mg/dL to 1.0 mg/dL (Katzberg,

plain, excretion, and contrast radiography

1997). Most cases of contrast media-induced nephrotoxicity are unrecognized because the serum creatinine concentration is not systematically checked after procedures. This nephrotoxicity is usually self-limiting. Serum creatinine usually peaks within 3–5 days, and usually returns to baseline within 10–14 days (Katzberg, 1997). Nephrotoxicity is extremely rare when kidney function is normal. Patients with chronic kidney disease caused by diabetic nephropathy appear to be the most at risk of developing contrast-induced nephropathy. They are more susceptible when dehydrated and when exposed to relatively large volumes of contrast agents. Therefore in high-risk patients, hydration is advised. Because treatment options are limited once oliguric renal failure has developed, most clinical effort has been aimed at prevention of contrast-induced acute kidney injury. Alternatively, CT urography with oral or IV hydration and with reduced contrast dose could be performed. In addition to active hydration, several studies have suggested various pharmacologic agent administrative regimens may be of benefit in preventing contrast-induced acute kidney injury, including N-acetylcysteine (Tepel et al., 2000) and bicarbonate. In patients with underlying chronic renal failure who undergo regular dialysis, contrast media is readily cleared by dialysis because contrast media molecules are not protein bound. Unless there is significant underlying cardiac dysfunction, or very large volumes of contrast media have been administered, there is no need for urgent dialysis. Patients with multiple myeloma may occasionally develop acute kidney injury following IV contrast administration. Dehydration should always be avoided in patients with myeloma. In diabetic patients with normal renal function taking the oral agent metformin (Glucophage®), discontinuing metformin is advised for at least 48 hours after administration of IV contrast material to minimize the risk of developing metformin-associated lactic acidosis. Inadvertent subcutaneous extravasation of IV administered contrast medium is not uncommon. Such extravasation may result in local pain, erythema, and swelling. These symptoms usually resolve with local therapy including both warm and cold compression and elevation of the affected extremity (Cohan et  al., 2010). An immediate surgical consultation is indicated when patients develop increased swelling or pain after 2–4 hours, altered tissue perfusion, changes in sensation in the affected extremity, or skin ulceration.

The normal excretion urogram The renal parenchyma is best assessed during the nephrographic phase of urography. The normal kidney may range from 9 to 13 cm in cephalocaudal dimension depending on sex and age. The left kidney is frequently larger that the right kidney by approximately 0.5  cm. The kidneys have sharp margins and a smooth contour. Congenital fetal lobulation, a common normal variant, can be differentiated from scars, prior renal infarction or inflammation by their smooth contour and regular spacing and relationship to normal calyces and is often bilateral. Renal parenchyma measures 3–3.5 cm in thickness in the polar regions and 2–2.5 cm in the interpolar regions. The number of the calyces varies considerably. Each calyx (minor calyx) is deeply cupped and surrounds one papilla. The peripheral portion of the calyx is called the fornix. A group of calyces, termed compound calyces, drains two to four papillae and is frequently seen in the polar regions. Two or more infundibula (major calyces), each leading to single or multiple calyces, arise

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Table 11.2  Reactions to iodinated contrast material and treatment Type of reactions

Treatment

Hives

Contrast injection discontinued if not completed. Minimal (less than four): observation If diffuse or symptomatic, diphenhydramine PO/IV 25–50 mg If severe, epinephrine IV (1:10,000) 0.1 mL/kg slow push over 2–5 min

Angio-oedema

Closely monitor the airway, O2 by mask Isotonic IV fluids Epinephrine SQ: (1:1000) 0.1–0.3 mL(0.1–0.3 mg) If severe or associated with hypotension or airway compromise, epinephrine IV (1:10,000) 0.1 mL/kg slow push over 2–5 min, up to 3 mL/dose. Repeat in 5–30 min as needed

Diffuse erythema

Isotonic IV fluid, 0.9% normal saline or lactated Ringer’s 1–2 L rapidly If hypotensive: epinephrine IV (1:10,000), 0.1 mg (1 mL) slowly. Repeat up to 1 mg Hydrocortisone (Solu-Cortef®) IV 200–300 mg push over 1–2 min

Bronchospasm

O2 10 L/min by mask Isotonic IV fluids Beta-agonist inhalers, metaproterenol (Alupent®) and albuterol (Proventil®), 2 puffs and inhale. Repeat as necessary Epinephrine SQ (1:1000), up to 0.3 mL. Repeat up to 1 mg total. If hypotensive, add epinephrine (1:10,000), slowly IV 0.1 mg (1 mL) slowly Repeat up to 1 mg total

Laryngeal oedema

O2 6–10 L/min by mask Isotonic IV fluids Epinephrine IV (1:10,000), 0.1 mg (1 mL) slowly. Repeat up to 1 mg total

Pulmonary oedema

Elevate torso O2 10 L/min by mask Furosemide (Lasix®) 10–40 mg IV over 2 min Morphine 1–3 mg IV Hydrocortisone (Solu-Cortef®) IV 200–300 mg push over 1–2 min Transfer to intensive care unit or emergency department

Hypotension with tachycardia (anaphylactic shock)

Legs elevated 60º or more, or Trendelenburg position O2 by mask Isotonic Ringer’s lactate or normal saline IV fluids Epinephrine IV (1:10,000) 0.1 mg (1 mL) slowly. Repeat up to 1 mg total Transfer to intensive care unit or emergency department

Hypotension with bradycardia (vagal reaction)

Legs elevated 60º or more, or Trendelenburg position O2 by mask Isotonic lactated Ringer’s or normal saline IV fluids, 1–2 L rapidly Atropine 0.5–1.0mg IV push slowly. Repeat up to 3 mg

Hypertensive crisis

O2 10 L/min by mask Secure IV access Furosemide (Lasix®) 40 mg slowly over 2 min. Nitroglycerine 0.4 mg tablet sublingual. Repeat every 5–10 min Transfer to intensive care unit or emergency department

Seizures/convulsions

O2 6–10 L/min by mask Lorazepan (Ativan®) 2–4 mg IV Transfer to emergency room for further evaluation and workup

Modified from Cohan et al. (2010).

separately from the renal pelvis. Conventionally, all branches from the renal pelvis, whether single or multiple, are termed infundibula. Normal infundibula are straight without bowing or displacement. The renal pelvis sometimes appears to be outside of the confines of

the kidneys, where it often has a distended appearance (the extrarenal pelvis). The upper ureter usually begins as a smooth extension from the renal pelvis and descends lateral to the transverse processes of

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the upper lumbar vertebrae. The middle third of the ureter is usually superimposed on the transverse processes of the lower lumbar vertebrae. The ureter crosses anterior to the iliac vessels at a slightly higher position on the right than the left. The distal ureter courses posterolaterally and then anteromedially to enter the bladder. Peristaltic activity may change the size and shape of the calyces, pelvis, and ureter from image to image. When the bladder is progressively distended, it is smoothly marginated and appears roughly spherical. On a post-void image, the mucosal pattern of the bladder is frequently identified.

CT urography Cross-sectional imaging studies including US, CT, and MRI are now used more often to assess the renal parenchyma because it has been shown that the sensitivity of detection of small renal masses with excretory urography is greatly decreased. With introduction of helical CT, CT urography has been increasingly used as a more definitive study for the evaluation of the urinary tract. The renal parenchyma is evaluated with CT scans before and after IV contrast administration, and then the collecting system is visualized by reformatted images generated from thin-section multidetector helical CT images obtained during excretory phase of contrast enhancement (Chow and Sommer, 2001; Caoili et al., 2002; Kawashima et  al., 2004a). CT urography has replaced standard excretion radiography in the majority of patient evaluations with urologic indications.

Contrast radiography Retrograde pyelography Retrograde pyelography is the opacification of the ureter and pelvicaliceal system by the direct retrograde injection of contrast media at the time of cystoscopy (Fig. 11.4) (Imray et al., 2000). The examination may be done in a cystoscopic suite, or the ureter may be cannulated and the patient may be subsequently brought to the radiology department for the examination. The examination is best performed with fluoroscopy and appropriate spot and overhead images. When a urothelial lesion is suspected, subsequent endoscopy with brushing or biopsy of the lesion for a histological diagnosis is performed under fluoroscopic control. Retrograde pyelography is performed to investigate lesions of the ureter and renal collecting system that cannot be defined adequately by less invasive imaging or, to visualize the collecting systems and ureters when iodinated contrast media is contraindicated. The procedure is performed with sterile technique and is contraindicated in a patient with a urinary tract infection. Retrograde pyelography cannot be performed when patients cannot or should not undergo cystoscopy (e.g. patients recovering from recent bladder or urethral surgery). The procedure may be impossible or incomplete when it is difficult to cannulate the ureter (e.g. patients with very large prostates). Delayed images can be obtained after retrograde pyelography to evaluate drainage of the collecting system. If significant obstruction is identified during retrograde pyelography then ureteral stent placement should be considered to avoid the risk of bacterial spread into the upper tract above the obstruction. Other complications of retrograde pyelography include ureteral perforation and contrast reaction. The most common ureteral injury during retrograde pyelography is perforation, occurring

Fig. 11.4  Retrograde pyelogram reveals normal contrast filling of the right ureter and intrarenal collecting system.

during advancement of the catheter or guide wire. These injuries are usually managed with either observation or stent placement depending upon the extent of the injury. Up to 10–15% of contrast media can be absorbed during retrograde pyelography. Caution is therefore advised in patients with a known contrast allergy. Fluoroscopic monitoring of retrograde pyelography is helpful to avoid excess contrast volume injection, reducing the amount of extravasation from the distended upper collecting system.

Antegrade pyelography Antegrade pyelography is performed to visualize the upper tracts and to delineate the site or nature of upper urinary tract obstruction when excretion radiography is unsatisfactory, retrograde pyelography cannot be performed (e.g. ureteral diversion), or alternative imaging techniques (US, CT, and MRI) are not definitive. This technique is indicated to determine whether a dilated collecting system is obstructed or not when there is renal dysfunction after kidney transplantation. Pyelography is an essential component of upper urinary tract urodynamic testing (Whitaker test). Antegrade pyelography is contraindicated in patients with uncorrectable bleeding diatheses, diffuse skin infection over the puncture site, or anatomic anomalies which preclude safe renal puncture. Non-dilated collecting systems are not a contraindication, but it is much more difficult to puncture a non-dilated collecting system percutaneously. A peripheral calyx or the renal pelvis is punctured percutaneously with a 21-guage, thin-walled needle from a posterior or posterolateral approach (Fig. 11.3). Renal localization is provided by means of contrast excreted after an IV injection or, in the event of a non-visualizing kidney, with US guidance. The procedure is carried out under fluoroscopic control and spot images are obtained.

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The side effect of this procedure is inadvertent puncture of adjacent intra-abdominal structures. Although puncture of the renal vein, kidney parenchyma, liver, spleen, or colon is possible, few complications ensue because of the small size of the needle.

Cystography Static cystography Static cystography provides information on bladder volume, contour, position, and integrity. Static cystography is performed to assess suspected bladder rupture, to demonstrate bladder diverticula, delineate vesicoenteric fistulae, and to assess postoperative healing following bladder or distal ureteral surgery. The normal cystogram The distended bladder is a smooth-walled organ with either a round or an oval shape. The oval-shaped bladder is often aligned vertically in the female and horizontally in the male. In the newborn the bladder lies above the symphysis pubis and descends as the child grows. In the older child and young adult the bladder lies at or below the level of the symphysis pubis.

Voiding cystourethrography Voiding cystourethrography (VCUG) provides anatomic information about the lower urinary tract during the physiologic act of micturition. VCUG is performed to diagnose vesicoureteral reflux, to assess bladder emptying, and to evaluate the urethra for posterior urethral valves in the infant male, urethral stricture disease and urethral diverticula in female patients. VCUG is useful in assessing certain types of voiding dysfunction (e.g. detrusor-external sphincter dyssynergia and neuropathic bladder) and demonstrating reflux into an ectopic ureter which inserts into the urethra. The bladder is filled with contrast material using a transurethral catheter as for a static cystogram. Once filled, the older child or adult patient is asked to void in the upright position. The procedure is monitored with videofluoroscopy and recorded with either spot images or video recording. In male patients, the voiding images should be obtained with the pelvis in a 45º oblique position similar to retrograde urethrography (RUG), so that the entire length of urethra is better demonstrated (Fig. 11.5.)

Fig. 11.5  Voiding cystourethrogram demonstrates normal bladder and female urethra (arrow). No vesicoureteral reflux.

or small bowel segment (pouch) is the most common method of establishing permanent urinary diversion (Spring and Deshon, 2000; Banner, 2001). The isolated but otherwise intact bowel loop serves as a simple conduit for urinary flow, transporting urine outward toward the stoma in a continuous, rhythmic, isoperistaltic manner. The detubularized pouch, on the other hand, lacks the contractivity to propel urine to the outside, thus becoming a reservoir (e.g. neobladder, continent diversion) rather than a conduit.

Retrograde urethrography RUG provides detailed visualization of the anterior urethra in the male. Unlike a voiding cystourethrogram, RUG often incompletely visualizes the posterior urethra because of the resistance to retrograde flow provided by the external urethral sphincter. Complete evaluation of the entire urethra often requires both procedures, which may be performed at separate intervals. RUG is rarely indicated in female patients. RUG is most frequently indicated to assess suspected or known urethral stricture disease, suspected urethral trauma, and to demonstrate urethral diverticula, fistulae, and neoplasms (Kawashima et al., 2004b). Retrograde urethrograms should be performed in all patients with pelvic trauma prior to cystography in order to minimize further urethral injury with planned bladder drainage catheter insertion.

Loopography and pouchography After cystectomy, anastomoses of the ureters to an isolated intact segment (loop) of ileum, transverse colon, or a detubularized large

Fig. 11.6  Loopogram shows contrast filling of the ileal conduit (IC) with reflux into the distal ureters bilaterally. A large filling defect in the right lower intrarenal collecting system with urothelial irregularity (arrow) is characteristics of urothelial tumour.

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Radiographic examination of loops or pouches is referred to as loopography or pouchgraphy, respectively. A loopogram is performed to visualize the bowel conduit and subsequently the upper urinary tracts by reflux of injected contrast media (Fig. 11.6). This procedure is utilized in postoperative patients with progressive renal failure to assess conduit integrity and to exclude ureteral obstruction. In such patients, the absence of reflux may indicate ureteral anastomotic obstruction. Alternatively, neobladder and continent diversion ureteral anastomoses are often created with antirefluxing techniques. Therefore, the absence of reflux into the ureters on pouchograms may not indicate urinary obstruction. Loopography is essential to delineate the anatomy of a urinary diversion and probable site of ureteral implantation prior to any planned endourologic or interventional procedures on the diverted ureters.

References Banner, M. P. (2001). Diagnostic uroradiology. In P. Hanno, S. B. Malkowicz, and A. J. Wein (eds.) Clinical Manual of Urology, pp. 87–133. New York: McGraw-Hill. Barbaric, Z. L. and Pollack, H. M. (2000). Abdominal plain radiography. In H. M. Pollack, B. L. McClennon (ed.) Clinical Urography (Vol. 1), pp. 67–146. Philadelphia, PA: W. B. Sanders. Caoili, E. M., Cohan, R. H., Korobkin, M., et al. (2002). Urinary tract abnormalities: initial experience with multi-detector row CT urography. Radiology, 222, 353–60. Chow, L. C., Sommer, F. G. (2001). Multidetector CT urography with abdominal compression and three-dimensional reconstruction. AJR Am J Roentgenol, 177, 849–55. Cohan, R. H., Jafri, S., Choyke, L.P., et al. (2010). Manual on Contrast Media, Version 7. Reston, VA: American College of Radiology. Dyer, R. B., Chen, M. Y., and Zagoria, R. J. (1998). Abnormal calcifications in the urinary tract. Radiographics, 18, 1405–24. Dyer, R. B., Chen, M. Y., and Zagoria, R. J. (2001). Intravenous urography: technique and interpretation. Radiographics, 21, 799–821.

plain, excretion, and contrast radiography

Friedenberg, R. M. and Harris, R. D. (2000). Excretory urography. In H. M. Pollack and B. C. McClennon (eds.) Clinical Urography (Vol. 1), pp. 147–281. Philadelphia, PA: W.B. Sanders. Greenberger, P. A. and Patterson, R. (1991). The prevention of immediate generalized reactions to radiocontrast media in high-risk patients. J Allergy Clin Immunol, 87, 867–72. Hattery, R. R., Williamson, B., Jr., Hartman, G. W., et al. (1988). Intravenous urographic technique. Radiology, 167, 593–9. Imray, T. J., Lieberman, R. P., and Pollack, H. M. (2000). Retrograde pyelography. In H. M. Pollack, B. L. McClennon (eds.) Clinical Urography (Vol. 1), pp. 282–302. Philadelphia, PA: W. B. Sanders. Katayama, H., Yamaguchi, K., Kozuka, T., et al. (1990). Adverse reactions to ionic and nonionic contrast media. A report from the Japanese Committee on the Safety of Contrast Media. Radiology, 175, 621–8. Katzberg, R. W. (1997). Urography into the 21st century: new contrast media, renal handling, imaging characteristics, and nephrotoxicity. Radiology, 204, 297–312. Kawashima, A., Vrtiska, T. J., LeRoy, A. J., et al. (2004a). CT urography. Radiographics, 24(Suppl. 1), S35–54 Kawashima, A., Sandler, C. M., Wasserman, N. F., et al. (2004b). Imaging of urethral disease: a pictorial review. Radiographics, 24(Suppl. 1), S195–216. Lasser, E. C. (1988). Pretreatment with corticosteroids to prevent reactions to i.v. contrast material: overview and implications. AJR Am J Roentgenol, 150, 257–9. Palmer, F. J. (1988). The RACR survey of intravenous contrast media reactions. Final report. Australas Radiol, 32, 426–8. Spring, D. B. and Deshon, G. E., Jr. (2000). Radiology of vesical and supravesical urinary diversions and orthotopic bladder replacements. In H. M. Pollack and B. L. McClennon (eds.) Clinical Urography (Vol. 1), pp. 357–77. Philadelphia, PA: W. B. Sanders. Tepel, M., van der Giet, M., Schwarzfeld, C., et al. (2000). Prevention of radiographic-contrast-agent-induced reductions in renal function by acetylcysteine. N Engl J Med, 343, 180–4. Yamaguchi, K., Katayama, H., Takashima, T., et al. (1991). Prediction of severe adverse reactions to ionic and nonionic contrast media in Japan: evaluation of pretesting. A report from the Japanese Committee on the Safety of Contrast Media. Radiology, 178, 363–7.

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CHAPTER 12

Intervention Steven Kennish Introduction Imaging technology and interventional techniques are constantly being improved and refined. The percutaneous insertion of a nephrostomy drain was the first of these and remains the most commonly performed and useful intervention. Natural extensions of this technique include both percutaneous stone removal and antegrade ureteric stenting. Nephrologists must be aware of what these fundamental procedures entail, their indications, the techniques involved in performing them, and their limitations and complications.

Percutaneous nephrostomy Introduction Radiologically guided percutaneous drainage of the kidney was first described in 1955, and has become the key procedure in the management of renal tract obstruction (Goodwin et al., 1955).

Indications Indications vary depending on local expertise and preferences. The main reasons for inserting a percutaneous nephrostomy (PCN) are given in Box 12.1. The nephrologist is most likely to become involved in organizing a PCN for a patient initially referred with renal failure in whom subsequent investigations have demonstrated an obstructive ‘postrenal’ cause. This may be congenital, developmental, stone related, malignant, or iatrogenic. The acutely obstructed kidney becomes ischaemic, and a PCN is a quick and effective method of decompressing the collecting system, draining infected urine and relieving pressure on the nephrons (Rickards et al., 1993; Heyman et al., 1997). A PCN is often performed before an exact diagnosis has been made. Bilateral renal obstruction, obstruction of a single functioning kidney, systemic features of sepsis, or refractory hyperkalaemia necessitate urgent intervention. Nephrologists will, however, do as much as they can to improve the patient’s condition to make the procedure safe and easier. This will include dialysis, blood transfusion, and correction of acidosis. There is little evidence that a PCN is superior to a retrograde stent as the primary treatment, even for an infected obstructed collecting system, but it does avoid a general anaesthetic and ureteric manipulation (Türk et al., 2012). A urology opinion should be sought as to the most appropriate management. In practice, a PCN is usually preferred, especially in the sick patient. Malignant ureteric obstruction may be secondary to infiltrative primary tumour (often of the bladder or prostate), malignant nodal enlargement (including lymphoma), metastatic disease, or

retroperitoneal fibrosis. PCN drainage of the kidneys preserves nephrons and can prevent the development of urosepsis, Iatrogenic ureteric occlusion or injury is an infrequent complication of major laparoscopic surgery (Palaniappa et al., 2012). Percutaneous urinary diversion allows healing of a leak at a site of injury, or a defective ureteroileal anastomosis. Fistulating disease within the pelvis, incontinence, and irritative bladder symptoms post radiotherapy are also recognized indications for urinary diversion necessitating bilateral PCN insertion. The transplant kidney may require a PCN in the early postoperative period if the ureteric stent has become occluded, or if there is urine extravasation.

Techniques PCN insertion techniques are very much tailored by individual radiologists. The basic principles of the procedure are outlined in Box 12.2 The risks quoted for haemorrhage, infection, and procedural failure should be based on individual operator audit data (see Table 12.1). Pre-procedural analgesia is recommended, and the patient’s coagulation profile should be normal. Antibiotics are administered if there is a clinical suspicion of infection or there are risk factors such as stone disease, recent instrumentation or an indwelling foreign body such as a ureteric stent. PCN insertion is usually performed with ultrasound and fluoroscopic guidance with the patient in either the prone or supine/ oblique position. A  posterior axillary line approach allows a puncture along the avascular plane of Brödel, a natural watershed territory between the dorsal and ventral branches of the renal arteries, so as to minimize vascular injury (Dyer et  al., 2002). The transplant kidney should be approached from a skin puncture lateral to the iliac fossa incision, to try to avoid crossing the peritoneum. Ultrasound allows for the visualization and avoidance of nearby intra-abdominal structures. The dark anechoic fluid-filled hydronephrotic pelvicalyceal system is very clearly demonstrated against the echo-bright renal sinus fat (Fig. 12.1). A subcostal approach avoids crossing the pleura, which is painful and runs the risk of complications. Most radiologists use ultrasound alone to guide the initial puncture. Typically a thin 22-G trocar needle or a larger 19-G needle with a 15-G outer sheath can be used for calyceal puncture. In expert hands the hydronephrotic pelvicalyceal system is usually accessed safely by a single puncture while the subject holds their breath. The calyces of the non-hydronephrotic kidney are much more of a challenge.

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Box 12.1  PCN indications ◆ Relief of ureteric obstruction

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Table 12.1  Complication rates associated with PCN insertion (Lewis and Patel, 2004; Wah et al., 2004)

◆ Drainage of infected urine

Complication

Rate

◆ Pain relief in ureteric colic

Haemorrhage

< 4%

Urosepsis

< 4%

◆ Urinary diversion to allow the healing of leaks or fistulae ◆ Preparation for percutaneous renal tract surgery.

Limitations Spinal deformities and ectopic kidneys make the procedure difficult, but it is uncommon for overlying pleura or bowel to preclude a percutaneous approach completely. The only absolute contraindication to PCN is an uncorrected bleeding diathesis, and whilst a haematology opinion is useful, retrograde stents may be more appropriate. Uraemic and anaemic patients have poor platelet function so these problems should be dealt with before proceeding. Extra-long puncture needles have been developed to deal with the morbidly obese patient. Ultrasound transducer-mounted needle guides are helpful in securing a trajectory predicted by an on-screen pre-calibrated tramline target (Fig. 12.1). A single-step, 8-Fr needle trocar-mounted drain insertion may be appropriate in the grossly hydronephrotic infected kidney, reducing manipulation and therefore the risk of bacteraemia. A 0.038-inch J-tipped guidewire can be adequately visualized with ultrasound alone, if fluoroscopy is not immediately available, or undesirable, such as in the pregnant patient (Fig. 12.2). Often a retrograde ureteric stent is preferable. Children require general anaesthesia.

Complications PCN is a safe procedure. Nevertheless, as with any intervention, complications can occur. PCN insertion performed at night and by radiologists who do not regularly perform the procedure have higher complication rates (Lewis and Patel, 2004; Wah et al., 2004). Frank haematuria is normal after nephrostomy insertion. It usually clears within 24–48 hours, but often, reassuringly, before the patient leaves the radiology suite.

Renal pelvic injury

< 1%

PCN dislodgement

< 15%

Persistent or worsening haematuria is an indication for a nephrostogram. If the nephrostomy tube has migrated and drain sideholes lie within the vascular renal parenchyma, then a replacement tube to tamponade the track is needed. Pseudoaneurysms and arteriovenous fistulae are uncommon, and the need for embolization for bleeding post PCN is exceptionally rare (Farrell and Hicks, 1997). Despite best efforts to avoid manipulation within, and overdistension of an infected collecting system, systemic urosepsis can arise due to pyelovenous backflow. Inadvertent enteric injury is rare and minimized by the use of ultrasound (Zagoria and Dyer, 1999). A permanent external urine drainage bag is never popular with a patient, unless there is no other option, and the PCN is understood to be preserving or improving quality of life. Internalized drainage with a ureteric stent is always a more attractive option psychologically, but may not provide adequate drainage, or may cause side effects. There is little data on the consequences of a PCN drain/s on quality of life, but the chances of preserving renal function and prolonging life in the terminally ill patient is something that always requires careful consideration. Ultimately the decision should be taken by the patient.

Percutaneous nephrolithotomy Introduction Fernström and Johansen first described what is now known as percutaneous nephrolithotomy (PCNL) in 1976 (Fernström and Johansson, 1976).

Box 12.2  Principles of PCN insertion technique 1. Aseptic skin preparation and draping 2. Infiltration of local anaesthetic down to the renal capsule 3. Ultrasound-guided puncture of a target calyx 4. Aspiration of urine 5. Careful contrast administration under fluoroscopy guidance 6. Guidewire placement through the puncture sheath in to the pelvicalyceal system 7. Dilatation of a percutaneous track 8. PCN drain insertion and deployment in the pelvicalyceal system 9. Fixation of the PCN drain with dressings and connection to a drainage bag.

Fig. 12.1  The dark anechoic hydronephrotic pelvicalyceal system contrasts with bright hyperechoic renal sinus fat. The pre-calibrated needle guide tramlines can be used to assist renal puncture.

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assessment of the patient with renal disease Table 12.2  Approach to pelvicalyceal stone burden

Fig. 12.2  The 0.038-inch wire can be easily visualized with ultrasound alone to confirm position.

PCNL has replaced open stone surgery, offering similar stone clearance rates with reduced morbidity. PCNL remains a major undertaking, not least because extracorporeal shock wave lithotripsy (ESWL) and modern endourological techniques are pushing the PCNL case mix exclusively towards the most complex cases.

Indications The main indications for PCNL are stones > 2.5 cm in diameter, obese patients with a poor response to ESWL, stones associated with distal renal tract obstruction, lower-pole stones, and stones associated with indwelling foreign bodies, such as encrusted stents. Failed primary or repeated ESWL and/or ureteroscopy treatment are also common reasons for trying this technique.

Techniques Typically both an expert interventional radiologist and urologist are needed. Pre-procedural imaging should be carefully evaluated in a combined endourology meeting. A planning computer tomography (CT) scan should be used to evaluate all but the simplest of stones. An unenhanced phase demonstrates the stone burden and a delayed urographic phase details the pelvicalyceal anatomy, essential for planning access. Three-dimensional reconstructions are particularly helpful. Aberrant pelvicalyceal anatomy can be identified, and the pleural reflections and position of the colon can be taken into account when considering the approach. Because stones are colonized with bacteria (especially Proteus spp.), administration of antibiotics pre-procedure is recommended. Either a prone or supine approach is possible. A supine approach with the patient in the lithotomy position allows for combined percutaneous and retrograde intrarenal stone surgery. Supine techniques have been shown by some to be more time efficient (Hoznek et  al., 2012; Kumar et  al., 2012). The urologist usually places a retrograde ureteric catheter up to the renal pelvis to allow for pelvicalyceal opacification as preparation for the percutaneous approach. The expert radiologist interprets and cross references the preoperative CT and on-table fluoroscopic and ultrasound imaging to plan the safest and best access to the collecting system. The usual approach is to target a calyx, which is in line with the long axis of the stone. The aim is to create as few tracks as possible to clear the maximum stone volume (see Table 12.2). Although

Stone location

Track

Isolated lower pole

Direct lower pole puncture

Multiple lower pole

Upper pole puncture

Isolated or multiple upper pole

Lower pole puncture

Staghorn

Lower pole ± further tracks

flexible instruments and laser lithotripsy can be invaluable, an optimally placed track will allow for much more effective rigid ultrasonic or pneumatic lithotripsy. Occasionally large staghorn stones require multiple tracks. Tracks of between 24 and 30 Fr diameter can be created by telescopic metal dilators or serial fascial dilators, but there is a risk of kinking the heavy-duty access wire. Although expensive, the balloon dilatation system is generally preferred despite the PCNL audit in the United Kingdom demonstrating a non-significant trend towards a higher blood transfusion rate with this method (Tomaszewski et al., 2010; Armitage et al., 2012). (See Fig. 12.3.) Occasionally intraoperative bleeding will halt progress. A large-bore nephrostomy tube can be placed, and a second-look procedure can be arranged after the bleeding has settled. A post PCNL nephrostomy is not always necessary (Table 12.3).

Limitations and complications Retrograde catheter placement by the urologist is not always possible in patients with complex lower tract anatomy or urinary diversions. If the stone burden is large enough to warrant PCNL, a direct ultrasound-guided puncture on to a calyx or calyceal stone is usually still possible. There is little agreement in the literature about how to assess the presence of residual post-PCNL fragments, and what constitutes a ‘stone-free’ state (Armitage et al., 2012). Comparison studies are

Fig. 12.3  A 30-Fr access sheath, safety wire, and retrograde catheter are required for percutaneous management of this staghorn stone.

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Table 12.3  Advantages of post-PCNL nephrostomy placement versus ‘tubeless PCNL’ Post-PCNL nephrostomy

Tubeless

Allows drainage

Reduced post-op pain

Allows post-op nephrostogram

Reduced post-op hospital stay

Tamponades track

No increased risk of bleeding

limited by uncertainty over stratification of case mix and complexity, but the Guy’s Stone Score has been shown to predict post PCNL stone free rate accurately (Thomas et al., 2011). Complications are usually related to haemorrhage, urosepsis, or urine leak and have been classified by severity on the basis of the nature of further treatment required, using a modified Clavien grading system (Tefekli et al., 2008) (Table 12.4). If there is clinical concern for significant postoperative bleeding, an urgent CT angiogram should be arranged after appropriate resuscitation. Post-PCNL imaging follow-up should be tailored to the individual patient, with rapid staghorn formers and those with underlying metabolic disorders (e.g. cystinuria) requiring a more intense schedule. Repeat unenhanced CT is a cumulative radiation hazard, and the combination of plain film and ultrasound is usually sufficient if recurrent stones are to be treated with ESWL in the first instance. Less traumatic, specially designed miniaturized dilators, sheaths, and instruments are preferred in the paediatric population but are associated with longer operative times in the adult (Wah et al., 2013). Fluid balance and prevention of hypothermia are particular concerns in the anaesthetized child. The PCNL population is a high-risk group for further stone formation and intervention. Longer-term renal impairment and hypertension cannot confidently be attributed to post-PCNL parenchymal scarring, although there is no control group for comparison.

intervention

Table 12.5  Causes of ureteric obstruction Non-malignant

Malignant

Stone disease

Bladder

Tuberculosis

Prostate

Post radiotherapy

Cervical

Retroperitoneal fibrosis

Breast

Pelviureteric junction obstruction

Retroperitoneal

Hydronephrosis of pregnancy

Nodes and lymphomas (move to right)

Indications Ureteric stents drain the pelvicalyceal system in cases of ureteric obstruction (Table 12.5) and allow fistulae and leaks to resolve by providing a frame around which ureteral epithelialization is facilitated. Ureteric stents used to protect the transplant kidney ureteric anastomosis are placed intraoperatively. An antegrade (as opposed to a retrograde) approach to stenting offers a greater success rate, especially for distal ureteric strictures. Even if a contrast injection demonstrates complete ureteric occlusion, it is often still possible to negotiate the stricture in an antegrade fashion with a torque-controlled hydrophilic wire (Fig. 12.4). Antegrade ureteric stent placement requires percutaneous renal access, which can be secured in advance with a PCN, or gained at the same sitting.

Techniques The basic outline of an antegrade ureteric stent insertion is described in Box 12.3. An upper pole or interpolar calyceal track provides the best angle of approach towards the pelviureteric junction if there is the luxury of planning the original PCN. A mature access track of 8 Fr aids the

Antegrade ureteric stent insertion Introduction The placement of a ureteric stent in an antegrade fashion is a natural extension of the PCN technique. Most patients find internal drainage more convenient and cosmetically acceptable (Banner, 1998). A ureteric stent is less likely to be inadvertently displaced than a PCN drain, and usually has a lower morbidity (Holmes et al., 1993).

Table 12.4  A UK PCNL audit based on 1028 procedures documents complication rates (Armitage et al., 2012) Complication

Rate

Haemorrhage requiring transfusion

2.5%

Haemorrhage requiring embolization

0.4%

Post procedural fever

16%

Post procedural sepsis

2.4%

Visceral injury

0.4%

Fig. 12.4  Even very tight distal ureteric strictures can be easily crossed with a guidewire and catheter.

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Box 12.3  Antegrade ureteric stent insertion ◆ Patient consent ◆ Preparation including antibiotics and sedation with analgesia ◆ Aseptic skin preparation and draping ◆ Local anaesthesia ◆ Creation of a percutaneous track or exchange of a pre-existing nephrostomy over a guidewire for an angiographic catheter ◆ Nephro-ureterogram to demonstrate the anatomy ◆ Guidewire and catheter manipulation through the obstruction ◆ Exchange of hydrophilic wire for stiff wire and removal of catheter ◆ Stent insertion and deployment ◆ Placement of a covering non-locking loop drain.

procedure significantly, but single-step percutaneous puncture and stent deployment can be undertaken. Gentle continuous rotation of the hydrophilic wire is often all that is needed to allow passage through very tight strictures. An acute sense of urinary urgency is often felt by the patient, which may be eased by distending the bladder with a saline-contrast mix to distance the guidewire tip from the trigone. The hydrophilic wire is exchanged for a 0.038-inch working wire via a catheter. Occasionally a stricture cannot be crossed by the softer stent material, which may buckle. The stent then needs to be carefully withdrawn to allow balloon dilatation if appropriate. The tip of the stiff wire is left within the pelvicalyceal system when the proximal stent pigtail is deployed, so as to facilitate the placement of a covering PCN tube. This can be used if there is early stent failure, which can occur if a percutaneous track has been created at the same procedure, due to fresh occlusive blood clots. Covering PCN tubes can be capped off to allow renal function monitoring. The use of non-locking tubes is advised as locking sutures can become entangled with the proximal stent pigtail. After several days, non-locking PCN drains can be removed on the ward, or under fluoroscopic guidance once a nephroureterogram confirms satisfactory drainage.

Limitations and complications Advances in the development of polyurethane and silicone copolymers have led to the creation of soft stents, which are relatively resistant to occlusion, fracture, and migration, and are well tolerated. Nevertheless these factors remain problematic. Stent encrustation is a particular problem in a subset of patients whose risk factors include a long indwelling time, urinary sepsis, history of stone disease, and metabolic abnormalities (Holmes et al., 2010). Ureteric stents should not be placed in patients with an active urinary tract infection. Co-polymer stent patency rates have been demonstrated to fall from 95% at 3 months post insertion, to 54% at 6 months (Lu et al., 1994). Full-length metal stents provide increased patency rates for up to 12 months in patients with malignant ureteric strictures. Nevertheless, both co-polymer and metallic stents remain vulnerable to failure from extrinsic malignant compression of the ureter. Nephrostograms demonstrate that most flow occurs around, rather

than through a ureteric stent. If problematic, vesicorenal reflux can be minimized by bladder catheterization. Irritative bladder symptoms due to stimulation of the trigone by the distal stent pigtails can be minimized by careful stent positioning. Short, thermo-expandable titanium-nickel stents, which do not contact the trigone, are an option in the patient with a short malignant stricture. In the United Kingdom, registering a stent insertion with the British Association of Urological Surgeons (BAUS) national stent register has become best practice. The clinician is alerted by email when stent exchange is required; avoiding the forgotten severely encrusted stent, which often requires both an endourological and percutaneous approach to retrieval (British Association of Urological Surgeons, n.d.). Co-polymer ureteric stents should be changed by the urologists in a retrograde manner at cystoscopy initially after 6 months, and if uneventful, annually thereafter. Individual patient circumstances and risk factors for encrustation should however be considered and stent changes timed to pre-empt complications such as encrustation or occlusion.

Renal biopsy (See also Chapter 14.)

Introduction Real-time imaging is essential for a safe needle biopsy of the kidney. Ultrasound or CT can be utilized. Biopsies are performed both by radiologists and nephrologists. The former are called upon in challenging cases, for example, obese subjects.

Indications Non-focal renal parenchymal biopsy is utilized to diagnose a diffuse nephropathy or to exclude renal transplant rejection. Targeted renal biopsy is being increasingly utilized for assessing the indeterminate, small, solid renal lesion, although its use remains somewhat controversial. With modern advancements in histological techniques, however, the targeted biopsy of small renal lesions is gaining popularity as a way of avoiding the unnecessary morbidity of either surgery or ablation for benign lesions (Beland et al., 2007).

Technique Written informed consent and a normal patient clotting profile are again mandatory prerequisites. A posterior approach to the native kidney avoids crossing the peritoneum. Ultrasound is preferred as it offers continuous real-time imaging and the flexibility to alter the planned trajectory at any time. An angled approach with the needle allows ribs, pleura, and bowel to be avoided. Aseptic technique and draping are employed. A  sterile ultrasound probe cover is utilized. Infiltration of local anaesthesia can be guided by ultrasound to anaesthetize a track down to the renal cortex. A small skin incision to allow the insertion of a spring-loaded core biopsy needle is then made. Suspended patient respiration stabilizes the target kidney. The goal of non-focal renal biopsy is to maximize the amount of cortex obtained whilst avoiding damage to the renal hilum. The fewest cores possible should be obtained to minimize bleeding complications. Immediate assessment with microscopy is useful to ensure enough material is obtained for light, immune-, and electron microscopy.

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Patients should have bed rest and regular assessment of blood pressure and heart rate for at least 8 hours post procedure. If there are any concerns for significant bleeding an unenhanced and postcontrast arterial phase CT should be arranged in tandem with resuscitative measures. Renal angiography and embolization is the gold standard treatment for active bleeding (Fig. 12.5). The needle track for iliac fossa transplant kidneys should pass lateral to the incision scar to avoid crossing the peritoneum.

Limitations and complications Ninety-five per cent of patients undergoing non-focal renal biopsy should have an adequate diagnostic sample taken at one visit. Targeted renal biopsy has the greatest sensitivity (97%) and negative predictive value (89%) for larger lesions of between 4 and 6 cm in size (Rybicki et al., 2003; Uppot et al., 2010). Renal biopsy is a safe procedure but minor bleeding is very common. Small perinephric haematomas are seen in approximately 50% of patients and up to a third have frank haematuria post procedure. Severe bleeding is uncommon, however, with transfusion rates of approximately 1% and embolization required in far fewer. Iatrogenic arteriovenous fistulae tend to be self-limiting if small. Tumour seeding of the renal biopsy needle track is very rare with only a handful of case reports in the literature.

Drainage of collections Introduction Percutaneous imaging-guided drainage offers a safe and rapid way of dealing with troublesome fluid collections. It is well tolerated by the patient, and may avoid the need for further surgery.

Indications A renal or perirenal abscess may present acutely as a source of systemic sepsis. Specific post-transplant collections include urinomas, haematomas, and lymphocoele. If a fluid collection is believed to be infected, painful or otherwise detrimental to the patients’ progress it is appropriate to request drainage as long as it is deemed to be safely accessible by the radiologist. Decisions about the management of a postoperative collection must be taken by the surgical team.

Fig. 12.5  A urinoma consequent to a ureteroileal anastomotic defect is drained percutaneously with CT guidance.

Damage to adjacent organs is also a possibility. Pre-procedural antibiotics and a normal clotting profile are required. Antiseptic skin preparation, draping, and local anaesthesia are the next steps. Local anaesthetic infiltration can be imaged with ultrasound or CT to ensure that sensitive structures such as the peritoneum are anaesthetized adequately, and that the needle trajectory is appropriate. Long 20-G needles can be used to infiltrate local anaesthetic and act as pathfinders down to the collection to guide subsequent trocar mounted drain placement. A tandem technique can be used which allows for single step track dilatation. An 8-Fr drain is inserted alongside the long needle (Uppot, 2011). Final positioning is confirmed with CT before advancing the drain over the inner trocar and aspirating. A modified Seldinger technique is usual practice. A  15-G sheath mounted on a 19-G needle is inserted in to the collection.

Techniques Radiologists often favour either ultrasound or CT based on expertise and familiarity. There are, however, specific factors which would favour using one modality over the other. It is obviously vital to avoid iatrogenic injury to important overlying or adjacent structures such as large blood vessels and bowel. If a safe percutaneous route is not available in an axial plane then ultrasound is favoured over CT as the modality to guide drainage. The ultrasound probe can be angled steeply and used to negotiate overlying structures. Ultrasound also allows safe continuous real-time imaging where the progress of the advancing percutaneous needle tip can be followed. CT on the other hand involves ionizing radiation and even with CT fluoroscopy the radiologist has to advance the needle, pause, and re-image. (See Fig. 12.6.) CT more accurately depicts a gas- and liquid-filled collection from gas- and liquid-filled adjacent bowel. It is often difficult to make this distinction with ultrasound as highly echogenic gas bubbles prevent the ultrasound beam from progressing to (and therefore imaging) deeper tissues. Discomfort, bleeding, and subsequent drain complications such as blockage and migration should be included in patient consent.

Fig. 12.6  Embolization coils are placed in to the feeding artery for a post-nephrectomy bleeding pseudoaneurysm.

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Withdrawing the inner needle allows fluid aspiration prior to inserting a 0.038 J-tipped wire. Positioning can be confirmed with ultrasound or CT prior to serial fascial dilatation and subsequently drain placement over the wire. The initial aspirate can be sent for analysis. Special drain dressings can be supplemented with sutures for security. Tension on the drain should be avoided. Migration and eventually drain displacement leaves a partially drained collection, which is much more difficult to target for re-drainage. Irrigation of an infected collection is not routinely advocated, as it has a tendency to promote bacteraemia. Once several doses of appropriate antibiotics have been administered the drain can be flushed if required.

Limitations and complications Haemorrhage, adjacent organ injury, sepsis, and infecting a previously sterile fluid collection are recognized complications. Drains can be painful and may kink, block, and be pulled out. It is important when draining a urinoma that urinary diversion away from the leak is also considered.

References Ahallal, Y., Khallouk, A., and El Fassi, M. J. (2010). Risk factor analysis and management of ureteral double-J stent complications. Rev Urol, 12(2–3), 147–51. Armitage, J. N., Irving, S. O., and Burgess, N. A. (2012). Percutaneous nephrolithotomy in the United Kingdom: results of a prospective data registry. Eur Urol, 61, 1188–93. Banner, M. P. (1998). Antegrade and retrograde ureteral stent placement. In M. P. Banner (ed.) Radiologic Interventions: Uroradiology, pp, 96–127. Baltimore, MD: Williams and Wilkins. Beland, M. D., Mayo-Smith, W. W., Dupuy, D. E., et al. (2007). Diagnostic yield of 58 consecutive imaging-guided biopsies of solid renal masses: should we biopsy all that are indeterminate? Am J Roentgenol AJR, 188, 792–97. British Association of Urological Surgeons (BAUS). Stent Registry. [Online]

Dyer, R. B., Regan, J. D., Kavanagh, P. V., et al. (2002). Percutaneous nephrostomy with extensions of the technique: step by step. Radiographics, 22, 503–25. Farrell, T. A. and Hicks, M. E. (1997). A review of radiologically guided percutaneous nephrostomies in 303 patients. J Vasc Interv Radiol, 8(5), 769–74. Fernström, I. and Johansson, B. (1976). Percutaneous pyelolithotomy. a new extraction technique. Scand J Urol Nephrol, 10(3), 257–9. Goodwin, W. E., Casey, W. C., and Woolf, W. (1955). Percutaneous trocar (needle) nephrostomy in hydronephrosis. JAMA, 157, 891–4.

Heyman, S. N., Fuchs, S., Jaffe, R., et al. (1997). Renal microcirculation and tissue damage during acute ureteral obstruction in the rat: effect of saline infusion, indomethacin and radiocontrast. Kidney Int, 51, 653–63. Holmes, S. A., Christmas, T. J., and Rickards, D. (1993). Ureteric stents. In D. Rickards, S. Jones, K. R. Thomson, et al. (eds.) Practical Interventional Radiology, pp. 53–65. London: Edward Arnold. Hoznek, A., Rode, J., Ouzaid, I., et al. (2012). Modified supine percutaneous nephrolithotomy for large kidney and ureteral stones: technique and results. Eur Urol, 61(1), 164–70. Kumar, P., Bach, C., Kachrillas, S., et al. (2012). Supine percutaneous nephrolithotomy (PCNL): ‘in vogue’ but in which position? BJU Int, 110(11), 1118–21. Lewis, S. and Patel, U. (2004). Major complications after percutaneous nephrostomy—lessons from a department audit. Clin Rad, 59, 171–9. Lu, D. S., Papanicolaou, N., and Girard, M. (1994). Percutaneous internal ureteral stent placement: review of technical issues and solutions in 50 consecutive cases. Clin Rad, 49(4), 256–61. Palaniappa, N. C., Telem, D. A., Ranasinghe, N. E., et al. (2012). Incidence of iatrogenic ureteral injury after laparoscopic colectomy. Arch Surg, 147(3), 267–71. Rickards, D., Jones, S., and Kellett, M. (1993). Percutaneous nephrostomy. In D. Rickards, S. Jones, K. R. Thomson, et al. (eds.) Practical Interventional Radiology, pp. 23–33. London: Edward Arnold. Rybicki, F. J., Shu, K. M., Cibas, E. S., et al. (2003). Percutaneous biopsy of renal masses: sensitivity and negative predictive value stratified by clinical setting and size of masses. Am J Roentgenol AJR, 180, 1281–7. Tefekli, A., Ali Karadag, M., Tepeler, K., et al. (2008). Classification of percutaneous nephrolithotomy complications using the modified clavien grading system: looking for a standard. Eur Urol, 53(1), 184–90. Thomas, K., Smith, N. C., and Hegarty, N. (2011). The Guy’s stone score-grading the complexity of percutaneous nephrolithotomy procedures. Urology, 78(2), 277–81. Tomaszewski, J. J., Smaldone, M. C., Schuster, T., et al. (2010). Factors affecting blood loss during percutaneous nephrolithotomy using balloon dilatation in a large contemporary series. J Endourol, 24(2), 207–11. Türk, C., Knoll, T., Petrik, A. et al. (2012). Guidelines on Urolithiasis. [Online] European Association of Urology. Uppot, R. N. (2011). Nonvascular interventional radiology procedures. In E. Quaia (ed.) Radiological Imaging of the Kidney, pp. 271–87. Heidelberg: Springer. Uppot, R. N., Harisinghani, M. G., and Gervais, D. A. (2010). Imaging-guided percutaneous renal biopsy: rationale and approach. Am J Roentgenol AJR, 194, 1443–9. Wah, T. M., Kidger, L., Kennish, S., et al. (2013). MINI PCNL in a Pediatric Population. Cardiovasc Intervent Radiol, 36(1), 249–54. Wah, T. M., Weston, M. J., and Irving, H. C. (2004). Percutaneous nephrostomy insertion: outcome data from a prospective multioperator study at a UK training centre. Clin Rad, 59, 255–61. Zagoria, R. J. and Dyer, R. B. (1999). Do’s and don’t’s of percutaneous nephrostomy. Acad Radiol, 6(6), 370–7.

CHAPTER 13

Ultrasound Toby Wells and Simon J. Freeman Ultrasound physics Greyscale ultrasound Ultrasound creates an image using short-duration pulses of high-frequency sound waves generated by electrically stimulating a piezoelectric crystal in a handheld ultrasound transducer. The pressure wave generated is transmitted to the patient using a coupling gel between the transducer and skin surface. Returning echoes hit the same crystal and the vibrations induced are reconverted to an electrical signal which is then processed to create the image. Different tissues reflect the waves to varying degrees when exposed to the ultrasound beam. This behaviour is largely determined by tissue density and is expressed as the ‘acoustic impedance’. At the boundaries between soft tissues of different acoustic impedance the sound wave is partially reflected. If the interface is close to perpendicular with the ultrasound beam (more than 60°) the reflected echo will return to the transducer and be detected and its strength determines the brightness of the interface on the display. At soft tissue/air or soft tissue/bone interfaces the difference in acoustic impedance is so great that almost all the sound is reflected and none left to image deeper structures. The inability of ultrasound to penetrate gas-filled organs (such as bowel or lung) and bone can cause difficulty in obtaining a useful diagnostic ultrasound study. Other echoes are derived from tiny tissue structures that are similar in size to the ultrasound wavelength because instead of reflecting the ultrasound beam they cause it to be scattered in all directions. This phenomenon is of particular importance in generating a Doppler signal from moving red blood cells. Ultrasound is assumed to have a constant velocity in soft tissue (about 1540 m/s), so the time interval between transmission of the ultrasound pulse and reception of the returning echo can be used to calculate the distance of the reflecting interface from the transducer. The crystals within the transducer are fired sequentially and the returning reflected and backscattered echoes analysed for their strength and delay to construct a real-time two-dimensional image, referred to as the greyscale or B (brightness)-mode. Higher-frequency sound produces images of higher resolution, but the sound is attenuated more quickly and so tissue penetration is reduced. This should be taken into account when selecting the appropriate transducer for an ultrasound study. A high-frequency transducer (7–11 MHz) gives better resolution and is ideal for imaging superficial structures such as the testes, or for endocavity use such as in transrectal ultrasound. A lower-frequency transducer (3.5–5 MHz) is usually required to visualize abdominal structures such as the native kidneys. Selecting the appropriate

transducer is therefore a trade-off between image quality and depth penetration. Resolution is optimized by ensuring that the focus of the beam is adjusted to the level of the region of interest. High-frequency transducers usually have a flat surface (linear array) producing a parallel ultrasound beam of the width of the transducer. Lower-frequency abdominal and endocavity transducers usually have a curved surface producing a diverging ultrasound beam and a wide ultrasound sector. Harmonic imaging and spatial compounding are techniques that are now routinely employed by most manufacturers to produce images with fewer artefacts and greater resolution or clarity.

Doppler ultrasound The Doppler effect is the change in frequency that occurs when a sound source or detector are moving relative to one another. This is a familiar phenomenon to us in our everyday lives; for example, the change in tone of a siren as an emergency vehicle passes a static observer. The same principle can be employed in diagnostic ultrasound to demonstrate and measure blood flow. The frequency shift of an ultrasound wave that has been scattered by cells in moving blood is proportional to the velocity of the scattering object. Blood flowing towards the transducer will increase the frequency; conversely, flow away from the transducer will decrease the frequency. The frequency shift that results is small in comparison with the transmitted frequency; usually in the audible range. The Doppler frequency shift also depends on the transmitted ultrasound frequency and the cosine of the angle between the ultrasound beam and the direction of blood flow. The greatest Doppler frequency shift is generated when blood is flowing directly towards or away from the transducer, so there is no Doppler signal if the vessel is parallel to the transducer. The ultrasound practitioner must therefore optimize probe positioning and, where possible, utilize beam steering to ensure a satisfactory angle. If measurement of flow velocity is required, the angle of blood flow relative to the ultrasound beam must be manually selected using angle correction which permits the flow velocity to be calculated from the Doppler frequency shift. In this situation the angle should be 60° or less. Doppler information may be displayed as either a plot of velocity against time on a pulse wave (spectral) graph, or direction of flow and velocities and may be translated into different colours on a colour flow map. Power Doppler is another mode that assigns colour to blood flow. It uses the power of the Doppler signal rather than the frequency shift to display blood flow and is more sensitive to low flow than colour Doppler but gives no information about velocity or direction of flow.

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Fig. 13.1  (A) Colour flow Doppler allows identification of an interlobar artery in a renal transplant. The spectral trace obtained is normal—note the forward flow in diastole with a trace likened to a ‘ski jump’, and resistive index (RI) of 0.65. (B) The spectral Doppler trace from this renal transplant demonstrates reversed diastolic flow, in this case due to renal vein thrombosis.

The normal spectral arterial renal Doppler trace has high velocity flow in systole but also persistent forward flow throughout diastole, with a trace likened to a ‘ski slope’ (Fig. 13.1). Various parameters have been described to quantify this pattern. One of these, the resistance index (RI), is described in the renal transplant ultrasound section of this chapter.

Ultrasound contrast imaging Ultrasound contrast agents are increasingly being used in many different areas of ultrasound practice. The most recently available agents are based on stabilized microbubbles of perfluorocarbon gas in lipid shells. These have an excellent safety profile and no renal toxicity (Correas et al., 2001). Ultrasound takes advantage of the resonant properties of the microbubbles in the sound beam, enhancing visualization of blood flow. Contrast-enhanced ultrasound (CEUS)

studies are performed at very low power settings using harmonic techniques. This requires specific software which is now available on most mid- and high-range systems. There are many potential applications of ultrasound contrast agents, including characterization of complex renal cysts, renal vascular disorders, infection, transplant assessment, differentiation between complex renal cysts and solid lesions, and between renal pseudomasses and tumours (McArthur and Baxter, 2012). There are further applications for CEUS in scrotal, bladder, and prostatic ultrasound (Piscaglia et al., 2012) (Fig. 13.2).

Safety Ultrasound is a safe imaging technique which does not involve ionizing radiation. Despite the enormous numbers of ultrasound studies performed there are no confirmed deleterious medical side

Fig. 13.2  Microbubble contrast-enhanced image of the right kidney. Split screen view: the image to the reader’s right is a conventional greyscale image; the image to the left is a contrast-specific image showing (normal) intense enhancement of the kidney (arrowheads) and overlying liver in the nephrogram phase. The small area of unenhanced tissue (arrow) is a renal cyst.

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effects of this technique but it is good practice to minimize exposure by adjusting the power output to the lowest possible levels to obtain a diagnostic quality image, and minimizing scan time. This is of particular importance in obstetric imaging when scanning the fetus. Energy is transmitted to the patient during ultrasound examinations and this may cause mechanical or thermal effects. The main mechanical concern is cavitation (the collapse of gas-filled bubbles liberating energy) with a risk of local tissue damage. An estimate of the probability of cavitation occurring is displayed as the mechanical index (MI) on the ultrasound machine display. As gas bubbles are not normally found in the body, cavitation is generally only of concern when ultrasound contrast microbubble agents are used at high acoustic power settings and at gas/tissue interfaces. The probability of tissue heating occurring is displayed as the thermal index (TI). For a more comprehensive review of clinically based ultrasound principles and physics see Correas et al. (2001).

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Fig. 13.3  A normal kidney showing echogenic renal sinus fat centrally, hypoechoic medullary pyramids, and intermediate echogenicity renal cortex peripherally of similar echogenicity to the adjacent liver. There is a small column of Bertin (normal variant) seen in the upper third of the kidney (arrow).

Renal ultrasound Introduction Ultrasound is the primary imaging modality in most kidney disorders. It is the first-choice investigation of renal functional impairment, in particular to exclude obstruction.

Technique and normal appearances To achieve accurate diagnosis requires attention to good ultrasound technique. The kidneys are orientated such that their upper poles are situated posteromedially to the lower poles. This should be borne in mind when orientating the transducer to obtain a true long-axis image of the kidneys. Close anatomical relation to the bowel means the lower poles are frequently obscured by bowel gas, and the upper poles may be obscured by ribs. Scanning usually begins with the patient in the supine position using a 3–5-MHz curvilinear transducer. Views of the right kidney can be facilitated by using the liver as an acoustic window. Scanning intercostally but angling down towards the feet may allow visualization of the lower poles when they would otherwise be obscured by bowel gas and compression can be used to displace gas. Rolling the patient into the lateral decubitus position and deep inspiration may also be of assistance. Fasting is unnecessary preparation for renal ultrasound. It should be remembered that although the kidneys, adrenals, and perinephric fat are enclosed in Gerota’s fascia, the two layers may not be fused inferiorly and medially to the kidneys. This can allow considerable mobility of the kidneys inferiorly and across the midline when rolling some patients onto their side. The renal cortex, medulla, and sinus can be differentiated on ultrasound. The central renal sinus fat is usually echogenic and the medullary pyramids are echo poor. The cortex covers the bases of the pyramids, extends down between them in the columns of Bertin, and is of intermediate echogenicity (Fig. 13.3). The relative reflectivity of the cortex is often compared to the adjacent liver or spleen, with the normal renal cortex being less echogenic (darker) than the normal liver and spleen. Much of ultrasound assessment of the kidney is subjective, but a commonly used quantitative measurement is renal length. Normal adult renal length is 10–12 cm, typically slightly larger on the left side and in male patients. Renal length correlates with the patient’s

height and a decrease in size is seen in old age (Emamian et al., 1993). Other measurements such as renal parenchymal thickness can be performed (where a value of > 10  mm is normal) (Dubbins, 2006). Direct visualization of renal arteries and veins can be achieved at the renal hila or by scanning the aorta and retroperitoneum from a variety of positions. Assessment of the spectral Doppler wave form in the main renal artery is often possible; a peak systolic flow velocity of > 180 cm/s is considered by many to be an indicator of haemodynamically significant renal artery stenosis (Strandness, 1994), while other authors have found the ratio of renal artery to aortic peak velocity more reliable (Li et  al., 2006). Ultrasound assessment of native renal artery stenosis is however frequently technically difficult or impossible and computed tomography (CT) or magnetic resonance imaging (MRI) angiography are more reliable techniques (Halpern et al., 1998). Renal perfusion can usually be readily demonstrated by colour or power Doppler so that even vessels as small as the interlobular arteries can be resolved in slim patients and in transplant kidneys.

Renal parenchymal disease Ultrasound is usually the first imaging investigation requested in patients presenting with acute or chronic kidney injury. The main role of ultrasound in this setting is to exclude renal obstruction, measure renal size, and evaluate parenchymal thickness and echogenicity. Ultrasound is insensitive in differentiating between the different causes of renal parenchymal disease and predicting prognosis. Sonographic changes are inconsistent and non-specific, whatever the underlying pathology. In patients with glomerular disease, ultrasound is often normal in the early stages, but sonographic abnormalities are often seen in patients with tubular-interstitial nephropathies (Quaia and Bertolotto, 2002) (Fig. 13.4). In acute kidney injury, renal size is occasionally increased due to inflammation and oedema but eventually the kidneys will reduce in size and renal parenchymal thickness will be lost. Focal cortical scarring may indicate reflux nephropathy or segmental infarction

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Fig. 13.4  This patient presented with acute kidney injury requiring dialysis. Ultrasound demonstrated increased cortical echogenicity (relative to the liver parenchyma) and loss of corticomedullary differentiation. Renal biopsy was performed showing acute interstitial nephritis.

and a small kidney with a smooth outline suggest the possibility of ischaemic renal disease. There is often increased echogenicity of the renal cortex. Initially the medullary pyramids may be unaffected and become more conspicuous, but the pyramids are also frequently involved leading to loss of corticomedullary differentiation. The ‘end-stage kidney’ appears as a small, shrunken, and echogenic organ that may even be difficult to identify within adjacent perinephric fat. Spectral Doppler ultrasound is unhelpful in determining the aetiology of parenchymal renal disease as all processes tend to show an increase in intrarenal vascular resistance (RI) (Mostbeck et al., 1991). Patients with end-stage renal failure, usually on long-term dialysis, may develop acquired cystic kidney disease (ACKD) (Levine, 1996). Ultrasound will demonstrate small kidneys containing multiple cysts. ACKD is associated with an increased risk of renal cancer with 2–7% of patients ultimately developing a renal carcinoma (Choyke, 2000).

occasionally also seen on ultrasound as an enlarged, elongated unilateral kidney with two distinct renal sinus complexes, often with a different orientation (Goodman et al., 1986). Unilateral renal agenesis has an approximate incidence of 1 in 1000, and is usually associated with compensatory hypertrophy of the contralateral kidney. Duplex kidneys are one of the more common congenital abnormalities found in 2.7–4.2% of patients on urographic series (Mascatello et al., 1997). Although ultrasound cannot usually make a definitive diagnosis, the condition can be inferred when the kidney is large in comparison with the contralateral side with a band of renal cortex dividing the sinus in the interpolar region (Fig. 13.5). The upper moiety may be hydronephrotic due to obstruction by a ureterocele (Nussbaum et al., 1986), which should be sought on bladder scanning. The lower moiety may have a dilated collecting system or cortical scarring secondary to reflux. Hypertrophied columns of Bertin represent a protrusion of normal parenchymal tissue into the renal sinus and are a regular finding on ultrasound, seen in almost half of kidneys in one study (Lafortune et al., 1986). The column is isoechoic with adjacent renal cortex and there should be no contour abnormality of the overlying renal tissue. Splenic (dromedary) humps are a focal bulge of normal renal parenchyma arising from the interpolar region of the kidney, usually on the left side. Many patients also show regular indentations of the renal contour representing persistent fetal lobulation. These conditions need to be recognized as a normal variant rather than a renal mass or evidence of scarring (Bhatt et al., 2007). Ultrasound contrast agents can be very helpful in confirming that pseudomasses are normal renal tissue (Mazziotti et al., 2010).

Ureteric obstruction and renal stone disease (See Chapter 200.) Ultrasound is requested to exclude obstruction, either as a cause of renal functional impairment or in patients with flank pain. Ultrasound is very accurate in detecting hydronephrosis (Ellenbogen et al., 1978), but it should be remembered that ultrasound demonstrates anatomical rather than functional changes (Webb, 2000). Pelvicalyceal dilatation may occur in conditions other than obstruction, such as vesicoureteric reflux, and there may be no dilatation for several hours following the onset of ureteric

Congenital abnormalities and pseudomasses The numerous congenital abnormalities and pseudomasses of the kidneys can be demonstrated with ultrasound. Renal ectopia is most commonly due to incomplete ascent. Pelvic kidneys are found in between 1 in 2200 and 1 in 3000 patients; are more common on the left side and are prone to ureteropelvic junction obstruction and formation of calculi (Cinman et al., 2007). The pelvic kidney can usually be identified by ultrasound but will be malrotated and often hydronephrotic and may be misinterpreted as a pathological mass by the unwary ultrasonographer who fails to note the absence of a kidney in the ipsilateral renal fossa. The most common fusion abnormality is the horseshoe kidney, which occurs in 1 in 400 individuals. The isthmus of tissue joining the lower poles is not always visible on ultrasound as it may be very thin or obscured by bowel gas, and ultrasound diagnosis may be difficult (Strauss et  al., 2000). This condition should be suspected if the kidneys have an abnormal orientation with medial displacement and poor visualization of the lower poles; anterior position of the renal pelvis and a curved renal shape. Crossed fused renal ectopia is

Fig. 13.5  A duplex kidney—it is large and has renal cortex extending into the renal sinus.

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Fig. 13.6  (A) Hydronephrosis—the anechoic collecting system is dilated. (B) The cause of the hydronephrosis is identified; a calculus at the vesicoureteric junction (arrowed). Note the dilated ureter proximal to this.

obstruction (Laing et al., 1985). Hydronephrosis is recognized when the echogenic renal sinus is replaced by communicating fluid-filled anechoic spaces, representing the dilated pelvicalyceal system (Fig. 13.6). It is important to demonstrate this communication as parapelvic cysts can be mistaken for dilated calyces (Fig. 13.7). There is also a delay in the restoration of normal appearances after relief of an obstruction, often for several days and a degree of pelvicalyceal dilatation may persist indefinitely, particularly after relief of longstanding or severe obstruction. A number of grading systems have been proposed for the severity of hydronephrosis but simply stating mild, moderate, or severe is adequate and avoids confusion. The accuracy of ultrasound in diagnosing renal obstruction can be improved by the use of Doppler techniques. Elevated intrarenal vascular resistance in an obstructed kidney can be demonstrated by spectral Doppler examination (Platt et al., 1989, 1993). Colour Doppler ultrasound can also be used to demonstrate absence or abnormality of jets of urine entering the bladder in patients with ureteric obstruction (Burge et al., 1991). These techniques can be of particular value in the evaluation of pregnant patients in whom differentiation between physiological

(A)

hydronephrosis and hydronephrosis due to stone obstruction can be particularly difficult and where the use of imaging investigation using ionizing radiation (CT of the kidneys, ureters, and bladder (KUB) and intravenous urography (IVU)) are undesirable (Hertzberg et al., 1993; Haddad et al., 1995). MRI can be very helpful in difficult cases and is safe after the first trimester (Spencer et al., 2004). Renal stones appear as bright echogenic foci. Within the kidney it can be difficult to distinguish a small stone from the echogenic renal sinus. Sensitivity is improved by using a higher-frequency transducer and ensuring the focus is located at the level of the suspected calculus. Ultrasound has a reported sensitivity of 96% and specificity of 89% for detection of stones in the pelvicalyceal system (with tomography as the gold standard) (Middleton et al., 1988), but the figure is as low as 37% for ureteric calculi (Aslaksen et al., 1990), largely due to overlying bowel gas. In female patients, transvaginal ultrasound can be used to demonstrate distal ureteric stones (Laing et al., 1994). CT has now become the investigation of choice for renal stone disease and should be the first-line investigation for most patients with acute onset loin pain performing

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Fig. 13.7  (A) Apparent hydronephrosis—multiple anechoic structures in the renal pelvis easily mistaken for a dilated collecting system on ultrasound, but subsequently shown to represent parapelvic cysts. (B) On CT urogram, renally excreted contrast is seen in the true collecting system, compressed by the adjacent cysts.

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much better than ultrasound (and IVU) in this situation (Yilmaz et al., 1998).

Solid and cystic renal masses Ultrasound is excellent at differentiating solid from cystic lesions of the kidney. Simple cysts are very common and increase in incidence with age (Ravine et al., 1993). Sonographically, cysts are anechoic, normally spherical, and have posterior acoustic enhancement—the area behind the cyst appears brighter than at the equivalent depth elsewhere on the image. They may be parapelvic, intraparenchymal, or exophytic, in which case observing movement with the kidney on respiration is helpful in making the diagnosis. Benign cysts must be distinguished from cystic malignancy. Septations, loculations, or solid components increase the likelihood of malignancy. The Bosniak classification categorizes cystic lesions by complexity based on their appearances at CT and is used to guide management (Bosniak, 2012). Central to this classification is the degree of enhancement following contrast medium administration. This is easily measured on CT, but only possible on ultrasound using microbubble contrast media (Tamai et  al., 1995). Caution should therefore be applied when extrapolating the Bosniak system to ultrasound practice and most complex cystic renal masses will require further evaluation with CT. A number of other non-malignant renal lesions may appear as complex cystic masses on ultrasound including multilocular cystic nephroma, hydatid disease, abscess, haematoma, and xanthogranulomatous pyelonephritis. The hereditary renal cystic conditions are often encountered during abdominal ultrasound examination. The most common is autosomal dominant polycystic kidney disease (ADPKD) which results in enlarged kidneys with multiple cysts of varying size, randomly distributed throughout the kidney. Ultrasound is sometimes requested to screen a first-degree relative of a patient known to have ADPKD. Diagnostic criteria exist relating the number of cysts to the patient’s age (Ravine et al., 1994; Pei et al., 2009). Screening ultrasound may give a false negative result if performed below 30 years of age (Nicolau et al., 1999). (See Chapters 306–309.) The ultrasound appearances of renal cell carcinoma are variable. Large tumours tend to be heterogenous but predominantly

(A)

hypoechoic. Smaller tumours may be hyperechoic and impossible to distinguish from angiomyolipomas (Vallancien et al., 1990). The presence of vascular invasion into the renal vein and inferior vena cava is readily assessed on ultrasound, although CT is required for complete staging (Fig. 13.8). Other solid renal masses include angiomyolipomas, which are usually brightly echogenic on ultrasound due to the presence of fat. Oncocytomas are usually impossible to distinguish from renal cell carcinoma on ultrasound, although a central scar is said to be characteristic and colour Doppler ultrasonography may show central radiating vessels (Fan et al., 2008). Upper tract transitional cell carcinomas are hard to see on ultrasound; the sensitivity of CT urography or IVU is much greater (O’Connor et al., 2010). Lymphomas have variable appearances which include diffuse infiltration and focal masses and they are typically hypoechoic.

Infection (See Chapters 175–179.) Ultrasound imaging is often requested in cases of suspected pyelonephritis. The findings may include reduced parenchymal echogenicity and increase in renal size. Perfusion defects may be detected using colour or power Doppler; however, the scan is frequently normal. In focal pyelonephritis (lobar nephronia) part of the kidney may be enlarged or of altered echogenicity, either increased, mixed, or decreased (Farmer et al., 2002). The main role of ultrasound is to exclude pyonephrosis or abscess formation. Emphysematous pyelonephritis has a characteristic appearance on ultrasound. The renal parenchyma contains gas which produces bright echoes with heterogeneous (‘dirty’) posterior acoustic shadowing. When the perinephric soft tissues are involved the entire kidney may be difficult to identify. Chronic pyelonephritis secondary to vesicoureteric reflux may cause renal scarring, seen on ultrasound as focal cortical thinning overlying a dilated or clubbed calyx. Dimercaptosuccinic acid (DMSA) scintigraphy is used more frequently in this setting due to its greater sensitivity (Roebuck et al., 1999). In xanthogranulomatous pyelonephritis chronic low grade infection is usually secondary to a central obstructing stone. There are no pathognomonic ultrasound features except that stones may be identified, the affected kidney is

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Fig. 13.8  (A) A large renal cell carcinoma at the upper pole of the right kidney (arrows). (B) Views of the inferior vena cava with Doppler show extension of the tumour into the cava as a filling defect.

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frequently enlarged and contains cystic spaces and fluid collections in the perinephric soft tissues (Tiu et al., 2001). (See Chapter 355.)

Trauma Improved access to CT scanning with modern multidetector row machines has meant that ultrasound is now rarely the first-line investigation for renal trauma. However, the portability of ultrasound means it is still occasionally useful in the emergency department, and it does have a role in follow-up. The key findings are loss of continuity of the renal cortex, perinephric collections of blood or urine, and disruption of renal perfusion. The use of ultrasound contrast agents may improve accuracy but as they are not excreted in the urine cannot be used to detect collecting system injuries.

Renal transplants (See Chapter 282.) About a half of all patients with end-stage renal disease (ESRD) rely on a functioning renal transplant (462 pmp in the United Kingdom). Ultrasound is vital in detection of the complications of renal transplantation, and is used both for regular surveillance and as the first line imaging investigation for graft dysfunction. Renal transplants are typically placed in either iliac fossa, but the axis of the kidney is variable and the scan plane must be tailored to accommodate this. The superficial position of the transplant usually allows it to be assessed easily and a higher-frequency transducer can often be used. The scan technique involves a thorough greyscale assessment of the transplant and evaluation of the surrounding soft tissues for fluid collections. Global renal perfusion is then assessed with colour Doppler followed by a spectral Doppler assessment of several interlobar arteries. Finally the main vascular pedicle and iliac vessels should be examined. In the early postoperative period, peritransplant collections may represent urinomas, haematomas, lymphocoeles, or abscess formation. Ultrasound-guided needle aspiration may be necessary to differentiate between them. Renal artery thrombosis (global or segmental) is easily recognized by an absence of perfusion of all or part of the transplant. Renal vein thrombosis may be difficult to visualize directly but can be inferred when the normal low resistance intrarenal spectral Doppler pattern is replaced by a high-resistance pattern with reversal of diastolic flow (Fig. 13.1). Arteriovenous fistulas and pseudoaneurysms are usually related to previous biopsies and are readily identified with Doppler techniques. A mild degree of hydronephrosis is a common finding and does not necessarily indicate ureteric obstruction but progressive hydronephrosis in the face of deteriorating graft function may require antegrade pyelography or nephrostomy drainage to exclude obstruction. The common causes of early graft dysfunction are acute tubular necrosis (ATN) and rejection. Despite early optimism (Rifkin et al., 1987), ultrasound has been unsuccessful in accurately differentiating between acute rejection and ATN (Kelcz et al., 1990; Chow et al., 2001). In both conditions the intrarenal vascular resistance increases (Saarinen, 1991)  (usually measured by calculating the RI from a spectral Doppler trace of an interlobar artery; the upper limit of normal is < 0.7). Ultrasound-guided biopsy is usually required. Ultrasound can also identify some of the causes of late transplant failure. Peak systolic flow velocities of > 250 cm/s in the main transplant artery indicate a haemodynamically significant stenosis (Patel et al., 2003). Increasing hydronephrosis suggests a ureteric stricture.

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RI measurements are not useful in the diagnosis of chronic allograft nephropathy (chronic rejection) although CEUS shows some promise in this area (Schwenger and Zeier, 2006).

Bladder ultrasound The bladder is usually evaluated through a suprapubic approach with a standard abdominal curvilinear transducer. To achieve useful information requires the bladder to be well distended. Quantification of bladder volume is easily achieved and is of particular importance in patients with lower urinary tract symptoms. This requires measuring distances in both transverse and the longitudinal planes, and is often performed pre and post micturition. A  commonly used formula for converting these linear measurements to an approximate volume is: V = ( H × W × L ) × 0.625 where V is volume, H is height, W is width, and L is length of the bladder. Most ultrasound systems will automatically perform these calculations but the result tends to underestimate bladder volume slightly; accuracy is, however, acceptable for routine clinical use (Nwosu et al., 1998). Three-dimensional ultrasound may give more accurate measurements (Byun et al., 2003) but is not widely available. The normal bladder wall measures 3–5 mm in thickness which may be useful in predicting the severity of bladder outflow obstruction, but measurements depend on the degree of bladder distension and which part of the bladder wall is measured (Oelke et al., 2007). Many bladder abnormalities can be identified and assessed by ultrasound including diverticula, urachal abnormalities, stones, and blood clots. In patients with cystitis, bladder ultrasound is usually normal (but bladder emptying can be assessed). In chronic cystitis there may be bladder wall thickening. The presence of echogenic foci with acoustic shadowing (representing gas) in the bladder wall should alert the ultrasound practitioner to the possibility of emphysematous cystitis. Gas in the bladder lumen, in the absence of instrumentation, is suggestive of a vesicoenteric fistula. Ureterocoeles can be elegantly demonstrated on bladder ultrasound. They are seen as cystic structures projecting into the bladder lumen from the ureteric orifices which can frequently be seen to fill and empty on real-time scanning (Fig. 13.9A). Detection of an ectopic ureterocoele arising from the upper moiety of a duplex kidney inserting below the bladder neck can be facilitated by scanning through the perineum (Vijayaraghavan, 2002). Transitional cell bladder cancer can often be identified on ultrasound as a soft tissue projection into the bladder lumen. Predictably polypoid tumours are much more reliably identified than sessile ones. Tumours < 5 mm in size and those located on the anterior wall or bladder neck are frequently more difficult to identify (Itzchak et  al., 1981)  and ultrasound cannot replace cystoscopy (Ozden et al., 2007). Ultrasound has a limited ability to differentiate transitional cell carcinoma from other bladder masses including other types of bladder tumour, invasive prostate carcinoma, adherent clots, and endometriosis–cystoscopic inspection and biopsy is always required. The simple manoeuvre of rolling the patient onto their side will differentiate dependent debris and clot from posterior tumour. The presence of flow within a bladder mass on Doppler interrogation usually indicates a bladder tumour (Fig. 13.9B).

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(A)

(B)

Fig. 13.9  (A) A ureterocoele, seen as a cystic structures projecting into the bladder lumen from the ureteric orifice. (B) A polypoid transitional cell carcinoma of the bladder with internal blood flow demonstrated by colour Doppler.

Paediatric renal ultrasound ( See Sections 15 and 16.) Ultrasound is the primary imaging modality in paediatric radiology for most conditions, largely because it does not involve ionizing radiation. However, to obtain good images requires the cooperation of the child and parents, warm coupling gel, a relaxed atmosphere, and patience and skill on the part of the ultrasound practitioner. The use of higher-frequency transducers is often possible and transducers with a smaller ‘footprint’ are often used. Urinary tract abnormalities diagnosed in infancy and childhood are sometimes associated with other extrarenal abnormalities or syndromes, for example the ‘VACTERL’ collection of non-randomly associated birth defects. Neonate kidneys are of greater relative size than adults, and charts are available to relate renal size to age (Rosenbaum et al., 1984). The renal cortex is brighter than in adult kidneys, medullary pyramids are also relatively large, corticomedullary differentiation greater, and there is little renal sinus fat (Hricak et al., 1983). By 1 year of age these morphological differences will have resolved and the kidneys resemble those of adults. Antenatal detection of renal tract abnormalities by scans performed in pregnancy is relatively common. Postnatal imaging for antenatal detected hydronephrosis is best performed at 48–72 hours after birth, as relative dehydration in the early postnatal period may produce a false negative result. The diameter of the renal pelvis is often used as a reproducible measure of the degree of hydronephrosis and can be used to guide further management (Sidhh et al., 2006). In most patients renal pelvic dilatation is due to an unobstructed extrarenal pelvis but postnatal ultrasound is required to identify the subgroup of patients with diseases that may require surgical intervention. These conditions include ureteropelvic junction obstruction (UPJO), congenital megaureter, vesicoureteric reflux, posterior urethral valves, duplex kidney, multicystic dysplastic kidney, and prune belly syndrome. Additional imaging investigations such as the micturating cystourethrogram and nuclear medicine diuretic isotope renogram are sometimes required (Belarmino and Kogan, 2006). Autosomal recessive and autosomal dominant polycystic kidney disease can both present in childhood (See Chapter 313). The recessive form also involves the liver leading to hepatic fibrosis

in some patients. In young children with this condition the renal cysts are often tiny and impossible to resolve with ultrasound, the kidneys appearing enlarged and globally echogenic but with time cyst formation may become apparent (Blickman et  al., 1995). ADPKD typically presents in adult patients, but enlarged kidneys containing multiple cysts may occasionally be seen in older children. Renal cysts may also be seen in patients with tuberous sclerosis but characteristically these patients have multiple renal angiomyolipomas, appearing as echogenic nodules within the renal parenchyma. Multicystic dysplastic kidney is another cause of a unilateral cystic kidney; it is usually detected on prenatal ultrasound and may be associated with contralateral UPJO in some patients. In children with urinary tract infection, an ultrasound study is usually performed to exclude a structural abnormality. In infants this is frequently accompanied by a micturating cystogram to exclude vesicoureteric reflux and a DMSA scan to detect renal scarring. The National Institute of Health and Care Excellence (NICE) have produced comprehensive guidelines on the management of urinary tract infection in childhood (NICE, 2007). Wilms tumour (nephroblastoma) is the commonest malignancy in childhood usually occurring in children aged between 2 and 5 years. The commonest presentation is with an abdominal mass. On ultrasound, Wilms tumour is seen as a solid mass replacing all or part of the kidney, often with hypoechoic areas of necrosis (De Campo, 1986)  (Fig. 13.10). Five to 10% are bilateral and there are a number of associated urogenital abnormalities (Breslow et al., 1993). Staging requires CT or MRI. Wilms tumour cannot be reliably differentiated from the more benign entity mesoblastic nephroma on ultrasound. Bladder abnormalities such as neurogenic bladder, stones, cystitis, urachal abnormalities. or rhabdomyosarcoma may be seen on ultrasound. Scanning with a full bladder is usually required. Because of its larger relative size, the normal neonatal adrenal is usually seen on routine ultrasound, unlike adults. There is progressive cortical involution in the first few weeks of life and tables are available for normal adrenal size in the newborn (Scott et al., 1990). Adrenal masses such as neuroblastoma and adrenal haemorrhage may be identified on ultrasound and Doppler may help to differentiate between these two conditions (Deeg et al., 1998).

chapter 13 

Fig. 13.10  Wilms tumour—a large heterogeneous mass arising from and replacing the kidney in a 2-year-old child. With thanks to Dr J. Foster, Derriford Hospital, Plymouth, UK.

ultrasound

insertion of a needle or biopsy trochar under direct visualization. Experience is required to keep the needle in the ultrasound sector and identify the position of the needle tip. Needle holders are available to assist this—they attach to the transducer and ensure the path of the needle remains within the field covered by the beam. The depth of the needle track can be adjusted. Bowel is less easily visualized (and so avoided) on ultrasound than on CT. Many recommend biopsying transversely across the lower pole of the kidneys as this avoids hitting the large vessels at the renal hilum, and yields more nephrons for the histopathologist to assess. Ultrasound is now routinely used in the placement of central venous lines/dialysis catheters, as it allows accurate infiltration of local anaesthetic and confident cannulation of vessels. A chest radiograph will still be required, however, to ensure that the tip is in the desired location—usually at the level of the lower superior vena cava, above the right atrium. Ultrasound has a further role in aiding the safe placement of peritoneal dialysis catheters.

Ultrasound for procedures

Assessment of dialysis fistulae

(See Chapters 12 and 18.) In addition to its extensive diagnostic role, ultrasound is ideally suited to guide interventions (Frede et al., 2001). It allows the

(See Chapter 256.) Ultrasound has a major role in pre-assessment, monitoring, and assessing poorly functioning fistulae.

(A)

(B)

(C)

Fig. 13.11  (A) Doppler ultrasound shows an increased peak systolic flow rate at a stenosis in the draining vein of an arteriovenous dialysis fistula, confirmed by subsequent angiography (B), and treated by angioplasty (C).

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A dialysis arteriovenous fistula (AVF) is a subcutaneous anastomosis between artery and vein, usually in the forearm but occasionally in the leg. A graft is usually made of polytetrafluoroethylene (PTFE) and is a synthetic communication formed in a similar position. AVFs generally have better long-term patency, fewer complications, less morbidity, and improved flow in comparison to grafts (Hirth et al., 1996; Rosas et al., 2003). There is some evidence that the systematic use of preoperative ultrasound to assess the arterial and venous anatomy of a proposed site of fistula formation increases the success rate (Brimble et al., 2002; Ferring et al., 2010). This is often referred to as ‘vein mapping’ and is used to confirm adequate vessel size and patency, absence of significant arterial disease, and exclude vascular anomalies. There is also evidence that prospective monitoring of fistulae and grafts for haemodynamically significant stenoses reduces morbidity, thrombosis, and intervention, improving long-term patency (Smits et al., 2001). Other authorities reserve assessment for those with clinical problems (Teodorescu et al., 2012). Monitoring can take several different forms, including clinical examination, flow measurement during dialysis using dilutional techniques, or Doppler ultrasound to assess the volume of flow. The flow rate in a healthy fistula or graft is up to 2000 mL/min, and there is a risk of thrombosis if the flow rate drops below a critical level. Further investigation and, where necessary, intervention is usually indicated in a graft when the flow volume is below 600 mL/min or drops more than 20–25% on serial ultrasound measurement and in a fistula when flow volume is below 300 mL/min. The ultrasound examination should also assess the entire vascular access circuit to identify abnormalities such as stenoses, thrombosis, intimal flaps and aneurysm/pseudoaneurysm formation in the supplying arteries and draining veins. The accuracy of Doppler ultrasound to assess overall flow rate through a fistula or graft as a screening test for dysfunction is debated. Some studies suggest comparable accuracy to fistulography, but an experienced operator is required and there is considerable interobserver variability (Akoh and Hakim, 1999; Schwarz et al., 2003). It is, perhaps, more universally accepted in assessing a fistula in which dysfunction is already suspected (Teodorescu et al., 2012). In these cases stenosis is visualized as a local increase in flow velocity. A peak systolic velocity of > 400 cm/s or greater than threefold focal increase in peak systolic velocity are parameters used to indicate a significant stenosis (Older, 1998). Ultrasound can often directly visualize thrombosis within the fistula or supplying vessels and this can be confirmed by incomplete or absent luminal filling on colour Doppler examination. The length of thrombus can also be measured and is useful in planning intervention. Formal angiography remains the gold standard for assessment of fistula or graft dysfunction, and allows intervention (Fig. 13.11). Good fistula function requires good arterial inflow and a low pressure venous outflow. Venous stenosis is less common with AVFs than grafts but can be more severe. It usually occurs as a consequence of neointimal hyperplasia at the venous anastomosis or in the draining vein, but can occur within the lumen of synthetic grafts.

Further reading Allan, P. L., Baxter, G. M., and Weston, M. L. (eds.) (2011). Clinical Ultrasound (3rd ed.). Edinburgh: Churchill Livingstone. Pozniak, M. A. and Allan, P. L. (eds.) (2013). Clinical Doppler Ultrasound (3rd ed.). Edinburgh: Churchill Livingstone, Elsevier.

References Akoh, J. A. and Hakim, N. S. (1999). Preserving function and long-term patency of dialysis access. Ann R Coll Surg Engl, 81, 339–42. Aslaksen, A. and Göthlin, J. H. (1990). Ultrasonic diagnosis of ureteral calculi in patients with acute flank pain. Eur J Radiol, 11, 87–90. Belarmino, J. M. and Kogan, B. A. (2006). Management of neonatal hydronephrosis. Early Hum Dev, 82, 9–14. Bhatt, S., MacLennan, G., and Dogra, V. (2007). Renal pseudotumors. AJR Am J Roentgenol, 188, 1380–7 Blickman, J. G., Bramson, R. T., and Herrin, J. T. (1995). Autosomal recessive polycystic kidney disease: long-term sonographic findings in patients surviving the neonatal period. AJR Am J Roentgenol, 164, 1247–50. Bosniak, M. A. (2012). The Bosniak renal cyst classification: 25 years later. Radiology, 262(3), 781–5. Breslow, N., Olshan, A., Beckwith, J. B., et al. (1993). Epidemiology of Wilms tumor. Med Pediatr Oncol, 21(3), 172–81. Brimble, K. S., Rabbat, C. G., Treleaven, D. J., et al. 2002). Utility of ultrasonographic venous assessment prior to forearm arteriovenous fistula creation. Clin Nephrol, 58, 122–7. Burge, H. J., Middleton, W. D., McClennan, B. L., et al. (1991). Ureteral jets in healthy subjects and in patients with unilateral calculi: comparison with color Doppler US. Radiology, 180, 437–42. Byun, S. -S., Kim, H. H., Lee, E., et al. (2003). Accuracy of bladder volume determinations by ultrasonography: are they accurate over entire bladder volume range? Urology, 62(4), 656–60. Chow, L., Sommer, F. G., Huang, J., et al. (2001). Power Doppler imaging and resistance index measurements in the evaluation of acute renal transplant rejection. J Clin Ultrasound, 29(9), 483–90. Choyke, P. L. (2000). Acquired cystic kidney disease. Eur Radiol, 10(11), 1716–21. Cinman, N. M., Okeke, Z., and Smith, A. D. (2007). Pelvic kidney: associated diseases and treatment. J Endourol, 21(8), 836–41. Correas, J. M., Bridal, L., Lesavre, A., et al. (2001). Ultrasound contrast agents: properties, principles of action, tolerance, and artifacts. Eur Radiol, 11(8), 1316–28. De Campo, J. F. (1986). Ultrasound of Wilms’ tumor. Pediatr Radiol, 16(1), 21–4. Deeg, K. H., Bettendorf, U., and Hofmann, V. (1998). Differential diagnosis of neonatal adrenal haemorrhage and congenital neuroblastoma by colour coded Doppler sonography and power Doppler sonography. Eur J Pediatr, 157(4), 294–7. Dubbins, P. (2006). The kidney. In D.L. I. Cochlin, P.A. Dubbins, B. B. Goldberg, et al. (eds.) Urogenital Ultrasound: A Teaching Atlas (2nd ed.), pp. 1–104. Oxford: Taylor and Francis. Ellenbogen, P. H., Scheible, F. W., Talner, L. B., et al. (1978). Sensitivity of grey scale ultrasound in detecting urinary tract obstruction. AJR Am J Roentgenol, 130, 731–3. Emamian, S. A., Nielsen, M. B., Pedersen, J. F., et al. (1993). Kidney dimensions at sonography: correlation with age, sex, and habitus in 665 adult volunteers AJR Am J Roentgenol, 160(1), 83–6. Fan, L., Lianfang, D., Jinfang, X., et al. (2008). Diagnostic efficacy of contrast-enhanced ultrasonography in solid renal parenchymal lesions with maximum diameters of 5 cm. J Ultrasound Med, 27(6), 875–85. Farmer, K. D., Gellett, L. R., and Dubbins, P. A. (2002). The sonographic appearances of acute focal pyelonephritis 8 years experience. Clin Radiol, 57(6), 483–7. Ferring, M., Claridge, M., Smith, S. A., et al. (2010). Routine preoperative vascular ultrasound improves patency and use of arteriovenous fistulas for hemodialysis: a randomized trial. Clin J Am Soc Nephrol, 5(12), 2236–44. Frede, T., Hatzinger, M., and Rassweiler, J. (2001). Ultrasound in endourology. J Endourol, 15(1), 3–16. Goodman, J. D., Norton, K. I., Carr, L., et al. (1986). Crossed fused renal ectopia: sonographic diagnosis. Urol Radiol, 8(1), 13–16.

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Haddad, M. C., Abomelha, M. S., and Riley, P. J. (1995). Diagnosis of acute ureteral calculus obstruction in pregnant women using colour and pulsed Doppler sonography. Clin Radiol, 50, 864–6. Halpern, E. J., Rutter, C. M., Gardiner, G. A., Jr, et al. (1998). Comparison of Doppler US and CT angiography for evaluation of renal artery stenosis. Acad Radiol, 5(8), 524–32. Hertzberg, B. S., Carroll, B. A., Bowie, J. D., et al. (1993). Doppler US assessment of maternal kidneys: analysis of intrarenal resistivity indexes in normal pregnancy and physiologic pelvicaliectasis. Radiology, 186, 689–92. Hirth, R. A., Turenne, M. N., Woods, J. D., et al. (1996). Predictors of type of vascular access in hemodialysis patients. JAMA, 276, 1303–8. Hricak, H., Slovis, T. L., Callen, C. W., et al. (1983). Neonatal kidneys: sonographic anatomic correlation. Radiology, 147, 699–702. Itzchak, Y., Singer, D., and Fischelovitch, Y. (1981). Ultrasonographic assessment of bladder tumors. 1. Tumor detection. J Urol, 126(1), 31–3. Kelcz, F., Pozniak, M. A., Pirsch, J. D., et al. (1990). Pyramidal appearance and resistive index: insensitive and nonspecific sonographic indicators of renal transplant rejection. AJR Am J Roentgenol, 155, 531–5. Lafortune, M., Constantin, A., Breton, G., et al. (1986). Sonography of the hypertrophied column of Bertin. AJR Am J Roentgenol, 146(1), 53–6. Laing, F. C., Benson, C. B., DiSalvo, D. N., et al. (1994). Distal ureteric calculi: detection with vaginal US. Radiology, 192, 545–8. Laing, F. C., Jeffrey, R. B., Jr., and Wing, V.W. (1985). Ultrasound versus excretory urography in evaluating acute flank pain. Radiology, 154, 613–16. Levine, E. (1996). Acquired cystic kidney disease. Radiol Clin North Am, 34, 947–64. Li, J.-C., Wang, L., Jiang, Y.-X., et al. (2006). Evaluation of renal artery stenosis with velocity parameters of Doppler sonography. J Ultrasound Med, 25, 735–42. Mascatello, V. J., Smith, E. H., Carrera, G. F., et al. (1997). Ultrasonic evaluation of the obstructed duplex kidney. AJR Am J Roentgenol, 129, 113–20. Mazziotti, S., Zimbaro, F., Pandolfo, A., et al. (2010). Usefulness of contrast-enhanced ultrasound in the diagnosis of renal pseudotumors. Abdominal Imaging, 35(2), 241–5. McArthur, C. and Baxter, G. M. (2012). Current and potential renal applications of contrast-enhanced ultrasound. Clin Radiol, 67(9), 909–22. Middleton, W. D., Dodds, W. J., Lawson, T. L., et al. (1988). Renal calculi: sensitivity for detection with US. Radiology, 167, 239–44. Mostbeck, G. H., Kain, R., Mallek, R., et al. (1991). Duplex Doppler sonography in renal parenchymal disease. Histopathologic correlation. J Ultrasound Med, 10(4), 189–94. National Institute for Health and Clinical Excellence (2007). Guidelines on Urinary Tract Infection: Diagnosis and Long Term Management of Urinary Tract Infection in Children. Nicolau, C., Torra, R., Badenas, C., et al. (1999). Autosomal dominant polycystic kidney disease types 1 and 2; assessment of US sensitivity for diagnosis. Radiology, 213, 273–6. Nussbaum, A. R., Dorst, J. P., Jeffs, R. D., et al. (1986). Ectopic ureter and ureterocele: their varied sonographic manifestations. Radiology, 159, 227–35. Nwosu, C. R., Khan, K. S., Chien, P. F. W., et al. (1998). Is real-time ultrasonic bladder volume estimation reliable and valid? A systematic overview. Scand J Urol Nephrol, 32(5), 325–30. O’Connor, O. J., Fitzgerald, E., and Maher, M. M. (2010). Imaging of hematuria. AJR Am J Roentgenol, 195(4), 263–7. Oelke, M., Höfner, K., Jonas, U., et al. (2007). Diagnostic accuracy of noninvasive tests to evaluate bladder outlet obstruction in men: detrusor wall thickness, uroflowmetry, postvoid residual urine, and prostate volume, Eur Urol, 52(3), 827–34. Older, R. A. (1998). Haemodialysis access stenosis: early detection with color Doppler US. Radiology, 207, 161–4. Ozden, E., Turgut, A. T., Turkolmez, K., et al. (2007). Effect of bladder cancer location on detection rates by ultrasonography and computed tomography. Urology, 69(5), 889–92.

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Patel, U., Khaw, K. K., and Hughes, N. C. (2003). Doppler ultrasound for detection of renal transplant artery stenosis-threshold peak systolic velocity needs to be higher in a low-risk or surveillance population. Clin Radiol, 58(10), 772–7. Pei, Y., Obaji, J., Dupuis, A., et al. (2009). Unified criteria for ultrasonographic diagnosis of ADPKD. J Am Soc Nephrol, 20(1), 205–12. Piscaglia, F., Nolsøe, C., Dietrich, C. F., et al. (2012). The EFSUMB guidelines and recommendations on the clinical practice of contrast enhanced ultrasound (CEUS), Update 2011 on non-hepatic applications. Eur J Ultrasound, 33, 33–59. Platt, J. F., Rubin, J. M., and Ellis, J. H. (1989). Distinction between obstructive and non-obstructive pyelocaliectasis with duplex Doppler sonography. AJR Am J Roentgenol, 153, 997–1000. Platt, J. F., Rubin, J. M., and Ellis, J. H. (1993). Acute renal obstruction: evaluation with intrarenal doppler and conventional US. Radiology, 186, 685–8. Quaia, E. and Bertolotto, M. (2002). Renal parenchymal diseases: is characterisation feasible with ultrasound? Eur Radiol, 12(8), 2006–20. Ravine, D., Gibson, R. N., Donlan, J., et al. (1993). An ultrasound renal cyst prevalence survey: specificity data for inherited renal cystic diseases. Am J Kidney Dis, 22(6), 803–7. Ravine, D., Sheffield, L. J., Danks, D. M., et al. 91994). Evaluation of ultrasonograhic diagnostic criteria for autosomal dominant polycystic kidney disease 1. Lancet, 343, 824–7. Rifkin, M. D., Needleman, L., Pasto, M. E., et al. (1987). Evaluation of renal transplant rejection by duplex Doppler examination: value of the resistive index. AJR Am J Roentgenol, 148(4), 759–62. Roebuck, D. J., Howard, R. G., and Metreweli, C. (1999). How sensitive is ultrasound in the detection of renal scars? Br J Radiol, 72(856), 345–8. Rosas, S. E., Joffe, M., Burns, J. .E, et al. (2003). Determinants of successful synthetic hemodialysis vascular access graft placement. J Vasc Surg, 37, 1036–42. Rosenbaum, D. M., Korngold, E., and Teele, R. L. (1984). Sonographic assessment of renal length in normal children. Am J Roentgenol, 142, 467–70. Saarinen, O. (1991). Diagnostic value of resistive index of renal transplants in the early postoperative period. Acta Radiol, 32(2), 166–9. Schwarz, C., Mitterbauer, C., Boczula, M., et al. (2003). Flow monitoring: performance characteristics of ultrasound dilution versus color Doppler ultrasound compared with fistulography. Am J Kidney Dis, 42, 539–45. Schwenger, V. and Zeier, M. (2006). Contrast-enhanced sonographu as early diagnostic tool of chronic allograft nephropathy. Nephrol Dial Transplant, 21, 2694–6. Scott, E. M., Thomas, A., McGarrigle, H. H., et al. (1990). Serial adrenal ultrasonography in normal neonates. J Ultrasound Med, 9(5), 279–83. Sidhh, G., Beyene, J., and Rosenblum, N. D. (2006). Outcome of isolated antenatal hydronephrosis: a systematic review and meta-analysis. Pediatr Nephrol, 21(2), 218–24. Smits, J. H., van der Linden, J., Hagen, E. C., et al. (2001). Graft surveillance: venous pressure, access flow, or the combination? Kidney Int, 59, 1551–8. Spencer, J. A., Chahal, R., Kelly, A., et al. (2004). Evaluation of painful hydronephrosis in pregnancy: magnetic resonance urographic patterns in physiological dilatation versus calculous obstruction. J Urol, 171, 256–60. Strandness, D. E. Jr. (1994). Duplex imaging for the detection of renal artery stenosis. Am J Kidney Dis, 24(4), 674–8. Strauss, S., Dushnitsky, T., Peer, A., et al. (2000). Sonographic features of horseshoe kidney: review of 34 patients. J Ultrasound Med, 19, 27–31. Tamai, H., Takiguchi, Y., Oka, M., et al. (2005). Contrast enhanced ultrasonography in the diagnosis of solid renal tumors. J Ultrasound Med, 24(12), 1635–40.

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Teodorescu, V., Gustavson, S., and Schanzer, H. (2012). Duplex ultrasound evaluation of hemodialysis access: a detailed protocol. Int J Nephrol, 2012, Article 508956. Tiu, C. -M., Chou, Y. -H., Chiou, H. -J., et al. (2001). Sonographic features of xanthogranulomatous pyelonephritis. J Clin Ultrasound, 29(5), 279–85. Vallancien, G., Torres, L. O., and Gurfinkel, E. (1990). Incidental detection of renal tumours by abdominal ultrasonography. Eur Urol, 18(2), 94–6.

Vijayaraghavan, S. B. (2002). Perineal sonography in diagnosis of an ectopic ureteric opening into the urethra. J Ultrasound Med, 21(9), 1041–6. Webb, J. A. (2000). Ultrasonography and Doppler studies in the diagnosis of renal obstruction. BJU Int, 86(1), 25–32. Yilmaz, S., Sindel, T., Arslan, G., et al. (1998). Renal colic: comparison of spiral CT, US and IVU in the detection of ureteric calculi. Eur Radiol, 8, 212–17.

CHAPTER 14

Computed tomography Eugene Teoh and Michael J. Weston Introduction Computed tomography (CT) is the most widely used cross-sectional imaging modality. Introduced in the 1970s, CT served as a means of generating images in the transverse plane (axial images) of the internal organs of the patient. It has since evolved to be used to obtain volumes of image data of the relevant body section, which may be manipulated with image processing software to view the internal organs on sub-millimetre sections in multiple planes, and many other applications including surface rendering to allow three-dimensional views. The rudimentary construct of the CT scanner involves an X-ray tube and a row of detectors built into a rotatable gantry. The patient lies on a couch that can move longitudinally through the gantry and images are acquired from relevant sections of the patient’s body when they are in position within the gantry (see Fig. 14.1). The gantry rotates around the patient and X-rays are exposed to acquire data from different angles, which are used to construct the image using image reconstruction algorithms. In modern-day helical CT scanners, this process is rapid, with the gantry in constant rotation and constant exposure and image acquisition being performed as the body part of interest moves through the gantry. The speed of acquisition is increased by the use of multiple rows of detectors, allowing the whole body to be imaged in a matter of seconds. The constructed CT image is laid out as a matrix of pixels, each designated its own CT number or Hounsfield unit (HU). The CT number corresponds to the average density of tissues (and thus level of grey) within the pixel, with a positive correlation between density and CT number. For example, air has a CT number of −1000, water of zero, and bone 500–1500. Since the advent of multidetector CT (MDCT), its use in the evaluation of the urinary tract has expanded widely and it has now replaced intravenous urography (IVU) and the plain radiograph. A variety of CT techniques exist for evaluation of the urinary tract and the choice of technique depends largely on the indication, taking into account other patient factors (e.g. age and contraindications to iodinated contrast).

Contrast agents and practical techniques Contrast agents Iodine is the principal component of contrast medium used in CT, as it provides the radiographic density. Contrast agents may be classified according to their osmolarity (high or low) and composition

(ionic or non-ionic). Both characteristics contribute to the agent’s osmolality (and hence the propensity to cause fluid shifts), the highest being high-osmolar, ionic agents and lowest being low-osmolar, non-ionic agents. In modern-day CT, low-osmolar, non-ionic contrast agents are used, because they are five to ten times safer than their ionic counterparts (The Royal College of Radiologists, 2010). Prior to the examination, checks for contraindications to contrast administration and risk factors for contrast-related adverse effects (drug reactions, nephrotoxicity, and lactic acidosis) must be undertaken (Table 14.1). Essential points to cover are a history of previous contrast reaction, asthma, renal function, diabetes mellitus, allergies, and metformin therapy (The Royal College of Radiologists, 2010). The renal function of the patient must be checked prior to the examination. This should be in the form of the estimated glomerular filtration rate (eGFR), taken within a 3-month period prior to the examination in clinically stable patients, or a 7-day period in patients who have an acute illness or have a history of renal disease (The Royal College of Radiologists, 2010). Special precautions should be applied in patients with renal impairment. This is set arbitrarily as an eGFR of < 60 mL/min/1.73m2 although thresholds and practice will vary depending on local protocols (The Royal College of Radiologists, 2010). The Royal College of Radiologists has published guidance on recommended action for each risk factor (see ‘Safety issues and contraindications’). As with the administration of any drug, patient consent must be obtained before administration. (The Royal College of Radiologists, 2010)

Unenhanced CT KUB CT scan of the kidneys, ureters, and bladder (CT KUB) is the most accurate and now the primary investigation of suspected ureteric colic (The Royal College of Radiologists, 2012). Accuracy rates for this technique are high with reported sensitivities of 96–100% and specificities of 95.5–100% (Tamm et al., 2003). The patient is normally scanned in the prone position with images obtained from the top of the kidneys to the bladder base. The use of oral or intravenous contrast medium is not required. A full urinary bladder is the only prerequisite as this allows more sensitive detection of bladder calculi. Low-dose techniques are now routine, having been shown to maintain high rates of accuracy in detecting stones despite reduced quality of the images of soft tissue structures. With MDCT, the images can be acquired in a single breath-hold but reconstruction of thin sections is essential to avoid missing small calculi, particularly those of low density (Saw et al., 2000).

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X-ray tube

Detector array

Fig. 14.1  Basic schematic of CT scanning.

Renal CT Renal CT is the first-line investigation for the characterization of a renal mass (The Royal College of Radiologists, 2012), previously detected by a different imaging investigation, for example, ultrasound or CT KUB. The examination requires the administration of intravenous contrast medium and routinely involves two phases: an unenhanced phase and a nephrographic phase. The unenhanced phase is acquired before contrast administration. It provides baseline characterization of the density of the lesion to establish enhancement patterns (if any) and detection of high-density

components which may be otherwise obscured with the administration of contrast medium. The nephrographic phase is acquired 100 seconds after administration of intravenous contrast. It produces homogeneous enhancement of the renal parenchyma, allowing for reliable detection of renal masses (Israel and Bosniak, 2005).

Renal transplant CT Although ultrasound examination is the first-line imaging test in patients with a renal transplant, CT is often the next investigation, This is most commonly needed in the postoperative setting (Fig. 14.2).

Table 14.1  Recommended action in patients with increased risk of adverse effects Risk factor

Action

Previous reaction

◆

Establish nature of reaction and causative agent Reassess the need for contrast, risk vs benefit of its use, and consider the use of other investigations ◆ If absolutely necessary, use a different low osmolality agent ◆

Asthma

◆

If symptomatic/not well controlled, defer examination if not urgent, subject to improved management of asthma ◆ If well controlled, reassess the need for contrast and consider the use of alternative investigations

Multiple allergies/ single severe allergy

◆

Renal disease/ diabetes mellitusa

◆

Metformin

◆

◆

If deemed necessary: ◆ Proceed with a low osmolality agent ◆ Maintain close medical supervision for 30 minutes with IV access maintained ◆ Ensure all other safety measures are in place

Establish the nature and sensitivity of allergy Reassess the need for contrast, risk vs benefit of its use, and consider the use of alternative investigations.

Considering the severity of renal impairment, reassess the need for contrast, risk vs benefit of its use, and potential use of other investigations ◆ Use the smallest possible dose of low osmolality agent ◆ Ensure the patient is optimally hydrated before and after contrast injection ◆

If renal function is normal, no further action is required If renal function is abnormal, cessation of metformin for 48 hours is recommended in consultation with the referring doctor

a Coexistent diabetes mellitus in patients with renal impairment carries significant risk.

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computed tomography

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Fig. 14.2  Renal transplant CT. (A) Coronal image of a transplanted kidney in the left iliac fossa acquired in the arterial phase, producing intense cortical enhancement compared to the medulla. The venous phase on the other hand produces a more homogeneous pattern of parenchymal enhancement. (B) The arterial anatomy is clearly delineated in the arterial phase. (C) Reconstructed oblique sagittal image demonstrating the entire length of the transplant artery (and its point of anastomosis onto the external iliac artery).

In most situations, the renal transplant CT involves three phases—an unenhanced phase, followed by an arterial and then a venous phase post contrast administration (Sebastia et al., 2001). In addition to demonstrating the presence and anatomical relationships of common postoperative complications such as collections, each phase has a specific role in the assessment of the transplant patient. The unenhanced phase is helpful in showing areas of haemorrhage or calcification, which are of high density on CT and might otherwise be masked after contrast medium administration. The arterial and venous phases are used to delineate the vascular anatomy and in particular to assess patency. The pattern of graft enhancement on these phases can also demonstrate areas of infarction.

CT urography CT urography (CTU) is the best test for detecting renal calculi, renal masses, and upper urinary tract urothelial cell carcinoma

(A)

(UUT-UCC) (The Royal College of Radiologists, 2012)  and is therefore the ‘one-stop’ imaging investigation of haematuria in the appropriate patient group (discussed in detail in ‘Indications’). The technique involves imaging the urinary tract in three phases:  unenhanced, nephrographic, and excretory, as for the sequence of radiographs performed in IVU (see Fig. 14.3). The unenhanced phase is performed as a CT KUB. The nephrographic phase acquires images of the kidneys (as previously described) after administration of intravenous contrast, allowing detection of renal masses. The excretory phase of the study assesses the collecting systems, ureters and, to a lesser extent, the urinary bladder when optimal luminal opacification of these structures has taken place following contrast excretion by the kidneys. It is best acquired 10–12 minutes after intravenous contrast administration.

(B)

Fig. 14.3  The three phases of CTU. (A) Unenhanced. (B) Nephrographic. (C) Excretory.

(C)

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Renal colic CT KUB is the most accurate, and therefore the primary investigation for suspected ureteric colic. In addition to the high accuracy rates for detection of calculi, it has superseded IVU by eliminating the need for intravenous contrast medium and by being quick to perform. The presence of obstructing calculi and the level of obstruction can be clearly defined. The stone size can be accurately measured, information which influences patient management. Furthermore, extra-genitourinary causes of the patient’s symptoms may be seen as unenhanced CT, for example, appendicitis and diverticulitis (Tamm et al., 2003). Most calculi appear as multifaceted, high-density foci, and the entirety of the kidneys, both ureters, and the bladder should be scrutinized for these. In the absence of calculi, ancillary signs of urolithiasis should be sought, such as hydronephrosis and/or hydroureter caused by distal obstruction, and perinephric and/or periureteric fat stranding indicating inflammation (see Fig. 14.4). These features may indicate recent passage of a stone or the presence of radiolucent calculi (particularly when there is a clear transition point of ureteric obstruction) (Smith et al., 1996).

Complications following renal transplantation Fig. 14.4  Reformatted image of the right kidney in a patient with significant stone disease previously treated with lithotripsy. There is hydronephrosis and hydroureter secondary to the formation of ‘steinstrasse’ (street of stones) within the distal ureter (arrowheads). There are also residual renal calculi predominantly within the lower pole calyx (arrow).

(See Chapter 280.) CT has a role in the evaluation of both immediate and delayed complications of renal transplantation. In the immediate postoperative period, abnormal Doppler waveforms on a transplant ultrasound may indicate the need for further assessment with CT. Arterial and venous thromboses, arterial stenosis, and infarcted grafts can be demonstrated on the triple-phase transplant CT (see Fig. 14.5). The

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Fig. 14.5  Renal transplant complications. (A) Stenosis of the transplant artery (arrow) demonstrated on CT in a postoperative patient with low urine output and abnormal findings on Doppler examination. (B) Diffuse infarction of the lower pole of a transplant kidney characterized by decreased density of the renal parenchyma (arrowheads), compared to the perfused upper pole. (C) Thrombus was also demonstrated as a low-density filling defect (bold arrowheads) within the transplant vein of the same patient, with further areas of infarction (arrows). The distal aspect of the vein remains patent (arrowheads). (D) In a clinical caveat similar to that described in (A), the finding of a kink in the main transplant artery (arrow) explains poor graft perfusion seen on Doppler examination.

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cause and anatomical relationships of other structural abnormalities seen on ultrasound, such as transplant hydronephrosis and collections, can also be characterized in more detail with CT. During long-term follow-up, complications from immunosuppression include opportunistic infection, occult sources of sepsis, and post-transplant lymphoproliferative disorder (PTLD). The use of CT is commonplace in the detection of such complications and in the latter, for staging and follow-up.

Haematuria (See Chapter 46.) The major anxiety in the patient presenting with haematuria is that it is a sign of urological malignancy. The choice of imaging investigation is dictated by the pre-test probability, influenced by two main factors: age and the nature of haematuria. CTU is indicated in older patients presenting with frank haematuria, where the possibility of malignancy is high. There is no clear and agreed age cut-off although evidence from large series support the age of 40 years as a suitable threshold for this indication (Khadra et al., 2000; Edwards et al., 2006). Urolithiasis is the most common benign cause of haematuria in younger patients presenting with frank haematuria, CT KUB is definitely indicated to evaluate for stone disease. The choice of imaging investigation is less clear-cut in subjects with microscopic haematuria. Most cases of upper tract cancers demonstrate persistent haematuria and in the older patient with microscopic haematuria, interval testing to establish persistence should inform the decision on the use of CTU (Edwards et al., 2006). In younger patients with microscopic haematuria, many factors should be considered before requesting CT. Persistence of haematuria, and the presence of pain justify performing a CT. Because of the higher prevalence of renal parenchymal disease in this younger group, suggestive features on clinical and urine dipstick examination, and abnormal renal function should lead first to nephrological assessment rather than CT. A CT KUB is justifiable when there is a corroborating history of renal colic or if ultrasound examination does not show signs of parenchymal disease.

Characterization of renal masses Renal CT is the first-line investigation for the characterization of a renal mass (The Royal College of Radiologists, 2012), usually first detected by a different imaging investigation. Based primarily on its enhancement characteristics and morphological features, renal CT should inform the clinician as to whether the mass is one that needs surgical removal or if it has characteristics justifying follow-up observation (Israel and Bosniak, 2005). When evaluating a lesion, it is also helpful to decide whether it is solid or cystic. Renal cell cancers (RCCs) are the commonest renal parenchymal tumours and are mostly solid (Fig. 14.6A) although 2–5% are predominantly cystic. RCCs vary widely in size, uniformity, and extent on CT, which explains why it cannot be definitively diagnosed using this investigation. It can also not be distinguished from an oncocytoma on CT. Other solid renal masses include angiomyolipomata, lymphomas, and metastases. Of these, an angiomyolipoma can be definitively diagnosed by the detection of fat within the mass (Fig. 14.6B) (fat has a CT number of –60 to –150 HU). Lymphomata and metastases typically feature as multiple bilateral solid masses.

computed tomography

When evaluating a cystic renal mass, the Bosniak renal cyst classification is applied (Israel and Bosniak, 2005). It allows the lesion to be categorized into one of five groups based on morphology and enhancement patterns. These suggest the probable nature of the lesion (Table 14.2). Each category has accompanying clinical management recommendations which should, however, not be adhered to strictly. They should instead serve as a guide to management of individual cases, taking into account factors such as history, co-morbidity, and patient choice, particularly in patients with category III lesions (Fig. 14.7) (Israel and Bosniak, 2005).

Characterization of adrenal masses Adrenal masses are a relatively common incidental finding on CT studies, with adenomas being the commonest and featuring in up to 1.5% of examinations (Dèahnert, 2011). In general, an adrenal mass can be diagnosed as an adenoma on unenhanced CT if its average density is ≤ 10 HU, owing to abundance of lipid content. If the mass is > 10 HU, it should be regarded as indeterminate and

(A)

(B)

Fig. 14.6  (A) An example of an enhancing predominantly solid renal mass (arrows), subsequently diagnosed to be RCC. (B) A fat-containing mass with an average density of −69 HU (arrows), in keeping with an angiomyolipoma.

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(B)

(C)

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Fig. 14.7  (A) A Bosniak category I simple cyst within the upper pole of the right kidney (arrow). (B) A Bosniak category II cyst with a hairline thin calcified septa (arrow). (C) A Bosniak category III cyst within the lower pole of the right kidney with an enhancing thickened and nodular inferior wall (arrow). (D) A Bosniak category IV cystic mass with an enhancing soft tissue component independent of its enhancing thickened wall (arrow).

can be further characterized by assessing its enhancement washout (Korobkin, 2000). Enhancement washout refers to the drop in density between a contrast-enhanced phase and a delayed phase (normally acquired 15 minutes after contrast medium administration). An enhancement washout of > 50% and a delayed phase density < 35 HU would classify the mass as a lipid-poor adenoma. Masses with enhancement washouts < 50% and delayed phase densities > 35 HU remain indeterminate and require further investigation. Depending on various factors such as pre-existing malignancy and size, these take the form of biopsy, surgery, follow-up studies, or radionuclide imaging. Importantly, no specific CT features can differentiate between a functioning and non-functioning adenoma (Korobkin, 2000).

Detection and characterization of urothelial lesions CTU has superseded IVU for imaging the upper urinary tract because of its increased sensitivity and specificity for urothelial abnormality (Anderson et al., 2007). As a cross-sectional technique, it depicts extraluminal pathology and where relevant, pathology within the urachus. UUT-UCC can be seen as enhancing soft tissue density masses and tend to cause filling defects within the opacified urinary tract (Fig. 14.8A, B). Some UUT-UCCs can manifest as circumferential wall thickening which may not even distort the lumen (Anderson et al., 2007).

Bladder tumours can manifest as filling defects within a contrast-opacified bladder (Fig. 14.8C), focal masses, or asymmetric wall thickening, whereas uniform wall thickening tends to represent benign disease (Cohan et al., 2009). It is important to note that in the specific context of suspected bladder cancer, CTU is only an adjunct to cystoscopy (Blick et al., 2012).

Detection and characterization of urinary tract injury The excretory phase of CTU (± unenhanced phase depending on prior imaging) is a non-invasive means of evaluating selected cases with this indication, particularly in cases of suspected ureteric injury where a conventional cystogram might not yield the required information. An example is illustrated in Fig. 14.9. Where there is suspicion of urinary tract injury in a patient who has undergone a trauma series CT, this can be performed with relative ease if the suspicion is raised early as the patient can be scanned after the appropriate excretory phase delay without having to receive a further dose of intravenous contrast (stressing the importance of the secondary survey in trauma). With CT, a better spatial representation of the pattern and extent of injury can be depicted compared to cystography.

Cancer staging As is the case for PTLD and urological cancer, CT is widely used in cancer imaging to establish the presence and extent of distant disease. Staging CT of the chest, abdomen, and pelvis is performed routinely in patients with RCC, 28% of whom have disseminated

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computed tomography

(C)

Fig. 14.8  (A) Axial image demonstrating multiple filling defects within the calyceal system (arrows), subsequently diagnosed to be transitional cell carcinoma. (B) Coronal image on the same patient, demonstrating further disease in the proximal ureter (arrow). (C) Multiple filling defects within the bladder (arrowheads) confirmed to be multifocal bladder cancer.

disease at presentation. The lungs, lymph nodes, liver, and bone are common sites of involvement (Dèahnert, 2011). In bladder cancer, magnetic resonance imaging is the most accurate imaging modality for local staging. Staging CT is indicated for cases of confirmed muscle-invasive bladder cancer, including CTU for complete examination of the upper urinary tracts (Stenzl et al., 2009).

The role of CT in other nephrological disease Beyond the spectrum of transplantation, malignant, and surgical pathology, CT has a more limited role in the evaluation of ‘medical’ renal disease. It may demonstrate features of some disease entities such as nephrocalcinosis in renal tubular acidosis, cortical and papillary necrosis, and AIDS-related nephropathy, amongst others.

(A)

(B)

Safety issues and contraindications (Table 14.1) Contrast medium administration (See Chapter 11.) Several practical measures must be put in place before the administration of contrast (The Royal College of Radiologists, 2010). An appropriately trained doctor should be immediately available to manage severe reactions. Within the confines of the radiology department, an appropriately trained individual should be immediately available to identify and manage severe contrast reactions. Resuscitation facilities should be readily available. A patient should not be left unsupervised within 5 minutes of contrast administration, and should remain on the premises for 15 minutes after (30 minutes in patients with an increased risk of contrast reaction) (The Royal College of Radiologists, 2010). Low-osmolar, iodinated contrast agents are generally associated with a low rate of adverse effects (0.153% in large series data) (Hunt et  al., 2009), the majority of which are mild and can be treated within the radiology department. Severe reactions (anaphylactic spectrum) are rare, constituting 3.3% of all adverse effects (Hunt et  al., 2009). Nonetheless, where identified (see ‘Techniques’), appropriate precautions need to be undertaken to reduce the risk of the adverse effects from contrast administration. These are summarized in Table 14.2. The responsibility for deciding on contrast medium use eventually lies with the supervising radiologist (The Royal College of Radiologists, 2010).

Radiation protection

Fig. 14.9  An example of CTU delineating ureteric injury in a patient presenting with vaginal discharge of urine 3 weeks post pelvic surgery. (A) Contrast opacification of a blind-ending left distal ureter (arrowheads) with a fine track of contrast leading into the vaginal cavity (arrow) which is opacified. (B) A separate communication between the vagina and rectum (arrow) is also demonstrated.

(See Chapter 10.) The main disadvantage of CT is the ionizing radiation dose to the patient. The total effective dose of an optimized CTU protocol (low-dose unenhanced scan, followed by a combined nephrographic and excretory phase) acquired by a modern 64-slice CT scanner is 20.1 mSv (Vrtiska et  al., 2009). This is equivalent to just over 9 years of background radiation in the United Kingdom or an added risk of fatal cancer of 1 in 1000. Hence, despite its superior accuracy, CT examinations of the urinary tract should not be undertaken lightly and as with all diagnostic ionizing radiation examinations, should be justified by the clinical

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Table 14.2  The Bosniak renal cyst classification system Category

Criteria

Management

I (benign)

Non-enhancing, low attenuation (< 20 HU) simple cyst with a hairline-thin wall. No septations, solid components or calcification

No further intervention required

II (benign)

Cystic lesion with one No further of more of the following intervention required characteristics: a few hairline-thin septations which may perceptibly (but not measurably) enhance; thin calcification or short segment of slightly thick calcification along its wall or septa; small (< 3 cm), non-enhancing, high attenuation cyst

IIF (follow-up)

Cystic lesion with one or more of the following characteristics: multiple hairline-thin/minimally thickened septations which may perceptibly (but not measurably) enhance; thick or nodular calcification along its wall or septa; large (> 3 cm), non-enhancing large (> 3 cm), non-enhancing high attenuation cyst

Imaging follow-up to establish stability

III (indeterminate)

Cystic mass with enhancing thickened irregular or smooth walls or septa

Surgery, biopsy

IV (malignant)

Category III features and distinct enhancing soft-tissue components independent of the wall or septa

Surgery

Source data from Israel and Bosniak (2005).

benefit balanced against the radiation risk (The Royal College of Radiologists, 2012). As previously discussed, appropriate selection of patients with a high pre-test probability of disease is key to achieving this balance. This is particularly true in the context of haematuria. This requires clear referral guidelines to be established and adhered to by radiologists and the referrer to have a sound understanding of indications for each examination. Communication between both

parties should always be maintained, to decide on the best imaging pathway for complex cases.

References Anderson, E. M., Murphy, R., Rennie, A. T., et al. (2007). Multidetector computed tomography urography (MDCTU) for diagnosing urothelial malignancy. Clin Radiol, 62(4), 324–32. Blick, C. G., Nazir, S. A., Mallett, S., et al. (2012). Evaluation of diagnostic strategies for bladder cancer using computed tomography (CT) urography, flexible cystoscopy and voided urine cytology: results for 778 patients from a hospital haematuria clinic. BJU Int, 110(1), 84–94. Cohan, R. H., Caoili, E. M., Cowan, N. C., et al. (2009). MDCT urography: exploring a new paradigm for imaging of bladder cancer. AJR Am J Roentgenol, 192(6), 1501–8. Dèahnert, W. (2011). Radiology Review Manual (7th ed.). Philadelphia, PA: Lippincott Williams & Wilkins. Edwards, T. J., Dickinson, A. J., Natale, S., et al. (2006). A prospective analysis of the diagnostic yield resulting from the attendance of 4020 patients at a protocol-driven haematuria clinic. BJU Int, 97(2), 301–5. Hunt, C. H., Hartman, R. P. and Hesley, G. K. (2009). Frequency and severity of adverse effects of iodinated and gadolinium contrast materials: retrospective review of 456,930 doses. AJR Am J Roentgenol, 193(4), 1124–7. Israel, G. M. and Bosniak, M. A. (2005). How I do it: evaluating renal masses. Radiology, 236(2), 441–50. Khadra, M. H., Pickard, R. S., Charlton, M., et al. (2000). A prospective analysis of 1,930 patients with hematuria to evaluate current diagnostic practice. J Urol, 163(2), 524–7. Korobkin, M. (2000). CT characterization of adrenal masses: the time has come. Radiology, 217(3), 629–32. Saw, K. C., McAteer, J. A., Monga, A. G., et al. (2000). Helical CT of urinary calculi: effect of stone composition, stone size, and scan collimation. AJR Am J Roentgenol, 175(2), 329–32. Sebastia, C., Quiroga, S., Boye, R., et al. (2001). Helical CT in renal transplantation: normal findings and early and late complications. Radiographics, 21(5), 1103–17. Smith, R. C., Verga, M., Dalrymple, N., et al. (1996). Acute ureteral obstruction: value of secondary signs of helical unenhanced CT. AJR Am J Roentgenol, 167(5), 1109–13. Stenzl, A., Cowan, N. C., De Santis, M., et al. (2009). The updated EAU guidelines on muscle-invasive and metastatic bladder cancer. Eur Urol, 55(4), 815–25. Tamm, E. P., Silverman, P. M., and Shuman, W. P. (2003). Evaluation of the patient with flank pain and possible ureteral calculus. Radiology, 228(2), 319–29. The Royal College of Radiologists (2010). Standards for Intravascular Contrast Agent Administration to Adult Patients (2nd ed.). London: The Royal College of Radiologists. The Royal College of Radiologists (2012). iRefer: Making the Best Use of Clinical Radiology. London: The Royal College of Radiologists. Vrtiska, T. J., Hartman, R. P., Kofler, J. M., et al. (2009). Spatial resolution and radiation dose of a 64-MDCT scanner compared with published CT urography protocols. AJR Am J Roentgenol, 192(4), 941–8.

CHAPTER 15

Magnetic resonance imaging Kazuhiro Kitajima, Akira Kawashima, and James F. Glockner Introduction Since its introduction in the mid 1980s, magnetic resonance imaging (MRI) has become a powerful imaging tool for the evaluation of patients with renal disease. It is a useful alternative to computed tomography (CT) in patients in whom the use of iodinated contrast media is contraindicated or in patients at risk for radiation exposure. MRI has its particular advantages over other imaging modalities. Many individual properties of the magnetic resonance (MR) signal, such as proton density, relaxation rates (T1 and T2), flow, chemical shift, diffusion and perfusion, contribute to image contrast. The individual weighting of these contributions can be altered by changing the pulse sequence. It demonstrates excellent contrast detail in normal tissues (e.g. kidneys, bladder, and prostate) and pathological lesions are frequently very conspicuous against the background of normal tissue. A variety of MR acquisitions can be performed quickly, within a comfortable breath-hold, and these reduce or eliminate motion artefact as well as acquiring dynamic images following contrast administration. It is an excellent tool for imaging arteries (MR angiography (MRA)) and veins (MR venography (MRV)). A  variety of techniques are available, some of which do not require gadolinium contrast agents. Specific information which can be obtained with MRI include blood flow in arteries or veins, the presence of fat or lipid within a lesion, and apparent diffusion coefficients of normal and pathological tissue.

Principles MRI depends on the magnetic properties of nuclei, predominantly hydrogen, in the form of water molecules. Water protons behave like very small spinning magnets, and in the presence of a large magnetic field will align along the direction of the field, thereby minimizing their energy state and also generating a small net magnetization within the patient. When radiofrequency (RF) pulses of the proper frequency are applied, the water protons absorb energy and alter the orientation of their magnetic vectors with respect to the magnetic field. When the RF pulse is turned off, the protons release energy and relax back to their ground state. The energy released is also in the form of RF waves, which are detected by the antennas or coils within the MRI system and converted into an image. This is akin to spectroscopy. The spatial information required for generation of an image is provided by imposing very

small gradients on the large static magnetic field, thereby changing the resonant frequency of water protons according to their spatial location. There are few but important contraindications to MRI. Patients with pacemakers, ferromagnetic intracranial aneurysm clips, cochlear implants, and metallic ocular foreign bodies cannot be imaged safely. Open or large-bore magnets, sedation, or general anaesthesia may be necessary for examination of paediatric patients or those with claustrophobia. Large patients may be excluded because of limitations in the diameter of the gantry opening or the weight limits of the examining table. MRI can be considered during pregnancy only if the benefit of the procedure outweighs the benefits of alternative non-ionizing diagnostic imaging studies.

Pulse sequences Superior soft tissue contrast resolution is one of the key advantages of MRI over CT. Unlike CT, MRI contrast arises from a complex relationship of many different factors including proton density, magnetic relaxation (T1 and T2 relaxation time constants), magnetic susceptibility, and flow. It is possible to ‘weight’ the relative contribution of these factors and thereby create images with different tissue contrast (i.e. T1 weighted, T2 weighted, proton density weighted, etc.). A pulse sequence is a computer program specifying the details of image acquisition by the MRI system, and by changing various parameters within the sequence it is possible to generate images whose contrast is weighted to different properties of tissue. Two relaxation times, for example, describe the return to the equilibrium state following the initial RF pulse: the T1 relaxation time, or spin lattice relaxation time, is a measure of a proton’s ability to exchange energy with its surrounding chemical matrix, and determines how quickly a tissue can become magnetized. The T2 relaxation time, termed the spin-spin or transverse relaxation time conveys how quickly a given tissue loses its magnetization. By altering two parameters within the pulse sequence, TR (repetition time) and TE (echo time), it is possible to change the image contrast to reflect predominant T1, T2, or proton density weighting.

T1-weighted imaging An image in which the contrast is primarily determined by differences of T1 relaxation values within the tissues is produced by choosing a relatively short TR of 250–600 msec and a relatively short TE of 10–20 msec for a spin echo or fast spin echo pulse sequence. In a T1-weighted image, tissue with short T1 relaxation times appears

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bright. T1-weighted images are a good way to assess morphology and anatomy of organs because of relatively stronger signal intensity (Fig. 15.1A). Breath-hold T1-weighted images can be obtained with three- (3D) or two-dimensional (2D) T1-weighted, spoiled gradient echo pulse sequences, which can improve image quality by reducing motion.

times (e.g. kidney) appears brighter (Fig.  15.1B). Pathological conditions such as tumours, inflammation, or oedema will have long relaxation times and will appear bright on T2-weighted images. Static fluid such as urine appears markedly bright on T2-weighted images due to its very long T2 relaxation times (Table 15.1).

T2-weighted imaging

Gadolinium-based magnetic resonance contrast agents and their side effects

A long TR (2000–4000 msec), long TE (60–120 msec) spin echo or fast spin echo pulse sequence produces a T2-weighted image. In a T2-weighted image, tissue with long T2 relaxation

Intravascular MR contrast agents are used to provide additional image contrast during MRI (Fig.  15.1C–E). Gadolinium is a

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(D)

(E)

Fig. 15.1  Normal kidneys. (A) Axial T1-weighted spoiled gradient echo (SPGR) image without fat suppression obtained at the level of the mid kidneys reveals kidneys of intermediate signal intensity are surrounded by retroperitoneal fat, which appears bright. The renal sinus appears bright because of fat. A = abdominal aorta; C = inferior vena cava; D = duodenum; PH = pancreatic head. (B) The kidneys appear hyperintense on T2-weighted fast spin echo image with fat suppression. The retroperitoneal fat signal intensities are suppressed. Urine in the renal collecting system (arrows = intrarenal collecting systems; P= renal pelves) appears markedly bright similar to cerebrospinal fluid and fluid in the bowel loops. A = abdominal aorta; C = inferior vena cava; D = duodenum; PH = pancreatic head. (C) Breath-hold dynamic enhanced T1-weighted SPGR image with fat suppression 40 seconds after starting IV administration of gadolinium contrast demonstrates increased enhancement of the cortices (straight arrow) with the corticomedullary interface representing the cortical nephrographic phase. a = proximal main renal arteries; v = proximal left main renal vein; A = abdominal aorta; C = inferior vena cava. (D) Dynamic-enhanced T1-weighted SPGR image with fat suppression 70 seconds after gadolinium injection demonstrates homogeneous renal enhancement representing the nephrographic phase. a = proximal main renal arteries; v = proximal left main renal vein; A = abdominal aorta; C = inferior vena cava. (E) Delayed-enhanced T1-weighted image with fat suppression demonstrates excreted contrast material in the renal collecting systems bilaterally, representing the pyelographic phase. P= renal pelves.

chapter 15 

magnetic resonance imaging

Table 15.1  Relative T1 and T2 values for tissues and body fluid in the genitourinary system T1-weighted image T2\T1 T2-weighted image

Absent (markedly low SI)

Long T1 (low SI)

Intermediate T1 (intermediate SI)

Short T1 (high SI)

Long T2 (markedly high SI)

Urine, simple cysts, oedema, inflammation, neoplasm

Proteinaceous fluid (complicated cysts, abscess)

Subacute haemorrhage (extracellular methaemoglobin)

Long T2 (high SI)

High free water tissue (kidneys, testes, peripheral gland of prostate, seminal vesicle, penis)

Subacute haemorrhage (extracellular methaemoglobin)

Intermediate T2

High bound water tissue (adrenal, central gland of prostate, muscle)

Fat (adipose tissue, fatty bone marrow)

Short T2 (low SI) Absent (markedly low SI)

Gadolinium contrast agent

Air, gas, calculi, compact bone, haemosiderin, ion deposition, highly concentrated gadolinium contrast

SI = signal intensity.

lanthanide metal with a k-shell configuration of unpaired electrons, which results in its paramagnetic character. Gadolinium in the free state is extremely toxic; however, when bound to larger stable molecules it is safe for human use. Gadolinium chelates, following intravenous injection, are predominantly eliminated by the kidneys via glomerular filtration. Gadolinium shortens T1-relaxation times of adjacent tissue. Therefore, T1-weighted pulse sequences are used after intravenous (IV) administration of gadolinium-based MRI contrast agents (GBCAs) (0.1–0.2 mmol/kg). Dynamic enhanced imaging, using breath-hold, 3D or 2D T1-weighted, spoiled gradient echo pulse sequences, permits evaluation of the enhancement properties of parenchymal tissue or masses and can also be used to assess renal arteries and veins (Fig. 15.2).

(A)

Adverse reactions Adverse reactions are encountered with a much lower frequency than is observed after administration of iodinated contrast media. The frequency of all adverse events after IV injection of GBCAs ranges from 0.07% to 2.4% (Cohan et al., 2010). The vast majority of these reactions are mild, including coldness at the injection site, nausea with or without vomiting, headache, warmth or pain at the injection site, paraesthesia, dizziness, and itching. Although most adverse events resulting from administration of iodinated contrast media occur within 30 minutes of administration, reactions to GBCA injection can develop after more than 1 hour after GBCA. Reactions resembling an ‘allergic’ response are very unusual and vary in frequency from 0.004% to 0.7%. Rash, hives, and urticaria are the most

(B)

Fig. 15.2  Renal vein thrombus in a 49-year-old man. Enhanced T1-weighted SPGR images with fat suppression in axial (A) and coronal (B) planes reveal filling defect in the right renal vein. Benign simple cyst is present in the right kidney (A). Specimen obtained from percutaneous needle biopsy of the kidney demonstrated membranous glomerulonephritis.

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frequent of this group, and very rarely there may be bronchospasm. Severe, life-threatening anaphylactoid or non-allergic anaphylactic reactions are exceedingly rare (0.001–0.01%). Fatal reactions to GBCAs are very rare. Treatment of moderate to severe adverse reactions to GBCAs is similar to that of moderate or severe reactions to iodinated contrast media. Personnel must be trained and the equipment readily available for management and/or resuscitation of patients receiving intravenous GBCAs for MRI. The frequency of acute adverse reactions to GBCA is about eight times higher in patients with a previous reaction to GBCA. Second reactions to GBCAs can be more severe than the first. Persons with asthma and various other allergies, including to other medications or foods are also at greater risk, with reports of adverse reaction rates as high as 3.7%. Although there is no cross-reactivity, patients who have had previous allergic-like reactions to iodinated contrast media are also in this category. In the absence of any widely accepted policy for dealing with patients with prior contrast reactions (especially to GBCAs) and the need for subsequent exposure to MR agents, it does seem prudent to take precautions in a patient who previously had a reaction to GBCAs. It should be determined if is GBCA necessary, if a different brand could be used, and if 12–24 hours of premedication with corticosteroids and antihistamines should be initiated. This protocol is particularly applicable in patients who had prior moderate to severe reactions.

Nephrogenic systemic fibrosis GBCAs previously were widely believed to be well tolerated and non-nephrotoxic, even in patients with impaired renal function. However, exposure to GBCAs, often used in MRI, in patients with renal failure and those maintained on dialysis has been linked with the development of nephrogenic systemic fibrosis (NSF). NSF is a rare disorder with a scleroderma-like presentation, which may progress to potentially fatal multiorgan-system fibrosing disease (Fig 15.3). NSF was first noted in 1997 and reported to the medical community in 2000 (Cowper et al., 2000). Between 1997 and 2007,

Fig. 15.3  Patterned plaques in nephrogenic systemic fibrosis. Plaques can be seen to be red to purple coloured, thin, and fixed. Reproduced from Girardi et al. J Am Acad Dermatol (2011) 65; 6:1095–1106 with permission.

> 500 cases of NSF in patients with severe renal insufficiency (glomerular filtration rate (GFR) < 30 mL/min/1.73 m2) were reported (Abu-Alfa, 2011), and no documented cases of NSF have occurred in patients with a GFR of > 30 mL/min/1.73 m2 without acute kidney injury. Additional major risk factors include use of high doses or repeat doses of GBCAs a proinflammatory state, acute kidney injury of any severity associated with hepatorenal syndrome, or during the perioperative period after liver transplantation. Because treatment options are currently limited, there has recently been an emphasis on prevention and identification of patients who are at increased risk of developing NSF prior to any GBCA injection. The US Food and Drug Administration (FDA) now recommends that specific types of GBCAs including gadodiamide, gadoversetamide, and gadopentetate dimeglumine should not be used for patients with acute kidney injury or chronic, severe renal insufficiency with a GFR < 30 mL/min/1.73 m2. It is advisable to avoid GBCAs in dialysis-dependent patients unless the possible benefits clearly outweigh the potential risk, and to limit the type and amount in patients with estimated GFR rates < 30 mL/min/1.73m2.

Magnetic resonance angiography MRA is accepted as an accurate and non-invasive technique for evaluation of patients with suspected renal artery stenosis and other renovascular diseases. Contrast-enhanced MRA utilizing a 3D spoiled gradient echo sequence is a time efficient and safe test when compared with conventional catheter-directed arteriography (Fain et al., 2001). Improved hardware and acquisition techniques have led to shorter acquisition times and improved spatial resolution, and state-of-the-art MRA now provides detailed anatomical information of the renal arteries (Fig. 15.4) and has proven highly sensitive and specific for detection of renal artery disease, including

Fig. 15.4  Gadolinium-enhanced three-dimensional MRA reveals normal single main renal arteries bilaterally.

chapter 15 

Fig. 15.5  Bilateral renal artery aneurysms in a patient with Ehlers–Danlos syndrome. Gadolinium-enhanced 3D MRA demonstrates aneurysmal dilation of the bilateral renal arteries at the renal artery bifurcation.

fibromuscular dysplasia, and aneurysms (Figs. 15.5 and 15.6). The visibility of the intrarenal vessels remains limited when compared to conventional angiography. Although false-negative studies are rare on MRA, slight overestimation of the degree of stenosis can

(A)

magnetic resonance imaging

occur and may lead to false-positive diagnoses. The combination of an anatomic MRA acquisition with a velocity-dependent technique (phase-contrast imaging) can help to minimize the tendency to overestimate stenoses (Schoenberg et al., 2002). As with conventional angiography, MRA is an anatomic test, and provides little information regarding the functional significance of a stenosis. It is highly accurate in determining the number of renal arteries, the size of the kidneys, and the presence of any anatomic variants. MRI can be combined with a functional test similar in concept to captopril nuclear renography (Chapter 16). This test, termed MR renography together with MRA may be helpful to determine functional significance in the work-up of renovascular hypertension (Bokacheva et al., 2009). Non-contrast techniques were the first MRA methods developed and include time-of-flight and phase-contrast MRA. These had several limitations, however, including motion artefact, long acquisition times, and insensitivity to in-plane flow, and were largely surpassed by 3D contrast-enhanced methods; however, the advent of NSF has stimulated a renewed interest in non-contrast MR angiography. A recently developed technique employing 3D steady-state free precession (SSFP) with arterial spin labelling has advantages including high vascular signal to noise ratio, fairly rapid acquisition times, and inherent flow compensation (Glockner et  al., 2010; Miyazaki and Akahane, 2012). This has shown sensitivities comparable to contrast-enhanced MRA for detection of renal artery stenosis in limited studies. Non-contrast MRA also tends to overestimate the degree of stenosis by signal loss due to post-stenotic turbulent rapid flow.

Magnetic resonance urography MR urography is an alternative to CT urography (Chapter 14), but is generally not used as the first-line examination for work-up of patients with haematuria. MRI is insensitive to urolithiasis

(B)

Fig. 15.6  Bilateral renal artery stenoses. (A) Gadolinium-enhanced 3D MRA demonstrates a high grade stenosis of the single right main renal artery (short arrow) and focal stenosis of the accessory left renal artery (long arrow). The left main renal artery is negative for stenosis. (B) Digital subtraction catheter-directed aortogram corresponds to the MR angiographic findings of bilateral renal artery stenoses indicated with short and long arrows.

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Fig. 15.7  Coronal view of maximal intensity projection of MR urogram utilizing maximal intensity projection of respiratory gated, heavily T2-weighted fast recovery fast spin echo sequence with use of contrast media. Urine (U) in the right renal collecting system and ureter appears markedly bright similar to fluid filled structures including cerebrospinal fluid (CSF), fluid-filled gastric fundus (S) and bowel loops, and biliary system. Prior left nephroureterectomy and cystectomy with ileal conduit urinary diversion for urothelial carcinoma.

because stones do not have signal characteristics that allow them to be readily detected. Several different approaches are available. One common technique obtains fast heavily T2-weighted hydrographic images using (a) 2D half-Fourier transformation fast spin echo sequences acquired in a single breath-hold and (b) 3D fast recovery fast spin echo sequence with respiratory gating (Fig. 15.7). These static–fluid MR urography techniques provide excellent images of the urinary tract, particularly when dilated and/ or obstructed. Another popular method is gadolinium-enhanced excretory MR urography. This technique is similar in concept to CT urography, and relies on visualizing the excretion of a gadolinium contrast agent in the urinary tract with a 3D, spoiled gradient echo T1-weighted acquisition. Visualization of the collecting system and ureters is further improved with a simultaneous dose of furosemide (5–10 mg) and is particularly useful in assessing non-obstructed urinary tracts (Takahashi et al., 2010). One advantage of MR urography in comparison to CT urography (Fig. 15.8) is that multiple images can be acquired over time until optimal visualization of the urinary tract has been achieved, without any concerns regarding patient radiation.

Functional magnetic resonance imaging Diffusion-weighted imaging (DWI) is a technique employing strong bipolar gradients to create a sensitivity of the MR signal to the thermally induced Brownian (or random walk) motion of water molecules and allowing in vivo measurement of molecular diffusion. The apparent diffusion coefficient (ADC) is a quantitative

Fig. 15.8  Coronal view of maximal intensity projection of MR urogram using breathold, T1-weighted 3D spoiled gradient echo sequence obtained during excretory phase of contrast enhancement. Prior right nephroureterectomy and cystectomy with ileal neobladder urinary diversion for urothelial carcinoma. Neoblad = ileal neobladder.

parameter calculated from DWI which is used as a measure of diffusion. Because the ADC is also dependent on capillary perfusion and water diffusion in the extravascular space, alteration of the ADC provides information regarding microstructural changes. ADC values of the kidneys are higher than ADC values of other abdominal organs, most likely due to high water content and abundant blood supply of the kidneys, with possible contributions from flow in the tubular system. In most studies, the ADC value of the renal cortex is higher than that of the renal medulla, presumably because of the higher perfusion component in the renal cortex. Measurements of renal ADC may be useful for characterization of diffuse disease as well as focal lesions. Restricted diffusion, characterized by high signal intensity on DWI and decreased ADC value, tends to indicate higher cell density (e.g. malignant neoplasm) and increased viscosity (e.g. purulent material in abscess and pyonephrosis). Other functional imaging techniques including diffusion tensor imaging (DTI), blood oxygen level-dependent (BOLD), and arterial spin labelling (ASL) are currently under investigation (Notohamiprodjo et  al., 2010). MR DTI might provide further information about the renal microstructure. It features diffusion measurements along at least six different directions from which the full diffusion tensor and thus the main diffusion direction can be calculated. BOLD MRI is used for non-invasive assessment of the intrarenal oxygen content and uses the paramagnetic effect of deoxyhaemoglobin for indirect depiction of renal oxygenation. ASL techniques can be used to study tissue perfusion without administration of contrast agents. Most ASL techniques, like flow-sensitive alternating inversion-recovery, apply arterial pre-saturation pulses to invert or saturate the magnetization of arterial blood flowing into a recorded section.

chapter 15 

Interpretations and indications Kidney On T1-weighted images, the kidney is outlined by the surrounding retroperitoneal fat (Fig. 15.1A) and the renal cortex is isointense or slightly higher in signal intensity than the medulla, with the degree of corticomedullary differentiation depending on the patient’s age and hydration status. On T2-weighted images, the kidney appears bright (Fig. 15.1B) and the cortex is isointense or slightly hypointense compare to the medulla. The most common uses of MR in the evaluation of the kidneys include MRA of the renal arteries, MRI of indeterminate renal masses shown on other imaging studies (Pedrosa et al., 2008), and staging of known renal malignancies (Reznek, 2004). MR evaluation of indeterminate renal masses should begin with axial T1-weighted images (with and without fat saturation) and axial fast spin echo T2-weighted images. These images allow for proper identification of the mass as well as providing vital tissue characterization with regard to potential fat content or haemorrhagic components. The most vital portion of an MR examination of an indeterminate renal mass is a dynamic gadolinium-enhanced T1-weighted spoiled gradient recalled echo (SPGR) pulse sequence. This is generally performed in an axial or coronal plane in a single breath-hold utilizing a 3D- or 2D-SPGR sequence (Fig. 15.1C–E). The images are acquired before gadolinium administration, at 40 seconds (the cortical nephrographic phase), at 70 seconds (the early homogeneous nephrographic phase), and at 120 seconds (the late homogeneous nephrographic phase) after gadolinium administration. Alternatively, a small test dose of contrast may be administered to determine the timing delay following contrast injection to achieve optimal visualization of the cortical phase. Excreted contrast medium starts to appear in the collecting system (the pyelographic phase) 3 minutes following IV contrast injection. When a renal mass is seen on MRI, the most important characteristic indicating the presence of a malignancy is enhancement of the mass by more than 15% above baseline after IV contrast enhancement (Ho et al., 2002). Subtraction of the pre-contrast from the post-contrast images may be helpful to detect contrast enhancement of a renal mass when the lesion contains haemorrhagic or proteinaceous contents and appears bright on pre-contrast T1-weighted images. MRI is the most accurate method for the detecting renal vein and inferior vena cava (IVC) tumour thrombi in patients with renal cell carcinoma compared to ultrasonography and enhanced CT (Reznek, 2004). MRI is also accurate in determining whether the filling defects in the renal vein and IVC represent bland thrombus or tumour thrombus (Aslam Sohaib et al., 2002). Gadolinium-enhanced MRA and MRV in addition to cross-sectional and 3D display can also provide pre-surgical ‘road mapping’ prior to surgery. This can be especially helpful in nephron sparing surgery. MRA depicts renal arterial and venous lesions with exquisite detail and is also becoming a primary imaging study in patients with suspected renal aneurysms, arteriovenous communications, and renal vein occlusion. These acquisitions can be combined with functional techniques that can be used to make rough assessment of overall and split renal function as well as renal blood flow.

Ureter The intrarenal collecting systems are generally not well visualized on routine imaging unless dilated. The renal pelvis appears dark on T1-weighted images and markedly bright on T2-weighted images,

magnetic resonance imaging

characteristics of a fluid filled structure. The renal pelvis and ureter can be identifiable on T1-weighted images by the surrounding retroperitoneal fat. The normal ureteral wall is not well visualized. MR urography is an alternative to CT urography and excretory urography to assess the upper urinary tract. T2-weighted MR urography utilizing breath-hold heavily T2-weighted images is helpful in depicting a dilated upper urinary tract and in identifying the level of urinary obstruction. Gadolinium-enhanced MR urography is useful to improve detection of upper tract urothelial carcinoma (Takahashi et al., 2010).

Bladder The bladder wall is well demarcated by perivesical fat but is often indistinguishable from the low-intensity urine on T1-weighted images. On T2-weighted images, the low-intensity bladder wall is well outlined by high-intensity urine. With its multiplanar capability and various pulse sequences that allow for fast dynamic enhanced imaging, MRI has the potential to become the imaging modality of choice in staging bladder cancer. With MRI, all of the observations previously possible with CT are not only possible with MRI, but in addition, MRI can identify macroscopic perivesical fat invasion and may be helpful in differentiating between superficial and deep muscle invasion of the bladder wall. MR can detect enlarged abdominal and pelvic lymph nodes. MRI is also useful for the evaluation of patient with stress urinary incontinence. Pelvic floor laxity and abnormalities of the supporting fascia can be demonstrated in incontinent women by obtaining fast sagittal and coronal T2-weighted images at rest and at maximal pelvic floor strain, allowing for the detection and delineation of cystocoeles, rectocoeles, enterocoeles, and uterine prolapse.

Prostate and seminal vesicles The prostate is well outlined by fat and is homogeneous in signal intensity on T1-weighted MR images. The glandular prostate is subdivided into the peripheral gland and the central gland. T2-weighted images provide the best depiction of the prostate zonal anatomy and capsule. On T2-weighted images, the peripheral gland appears as a high-signal-intensity area in the posterior and posterolateral aspect of the gland surrounding the intermediate to low-intensity central gland. In young men, the peripheral gland generally constitutes 70% of the glandular tissues and is very bright on T2-weighted images. In older men, the central gland is composed mainly of the enlarged transitional zone from benign prostatic hyperplasia, and becomes heterogeneous on T2-weighted images. Prostatic cancer typically appears as areas of decreased signal in the peripheral gland on T2-weighted images. Unfortunately, prostatitis, post-biopsy haemorrhage, and atrophy can have a similar appearance. Cancer in the transition zone is shown as a homogeneous low signal mass with indistinct margins on T2-weighted images. Additional techniques including DWI, dynamic contrastenhanced (DCE) imaging, and proton (1H) MR spectroscopy can help further increase the level of confidence in detection and characterization of prostate cancer by providing functional and molecular information (Hoeks et al., 2011). DWI is an emerging clinical tool which does not require contrast administration: recent reports have demonstrated that ADC maps can improve detection and estimation of tumour aggressiveness. DEC imaging is the most common method for direct depiction of tumour vascularity and perfusion characteristics and can improve detection of cancer.

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(A)

(B)

Fig. 15.9  Left suprarenal mass (arrow) appears significantly dark on coronal out-of-phase T1-weighted spoiled gradient echo image (A) compared to that on in-phase image (B), characteristic of benign adenocortical adenoma with abundant intracellular lipids.

1H MR spectroscopy of the prostate is able to depict spectral profiles of cellular metabolites and characterize cancerous lesions by demonstrating reduced citrate and elevated choline peaks. MR spectroscopic imaging combined with MRI can improve tumour localization and volume measurement compared with conventional MRI alone. However, MR spectroscopy needs specialized software and a high level of expertise in scanning and interpretation. The seminal vesicles appear as symmetrically paired ovoid cystic structures. MRI is valuable for the evaluation of patients with ejaculatory dysfunction (e.g. haematospermia, painful ejaculation) and useful in detecting stones, masses, and obstruction of the seminal vesicles and ejaculatory ducts.

Urethra High-resolution MRI with phased-array pelvic and endorectal coils has dramatically enhanced the ability to visualize abnormalities of the female urethra and periurethral tissue. MRI is accurate in diagnosing diverticula of the female urethra. The male urethra is rarely visualized on routine images unless a transurethral Foley catheter is inserted. MRI is useful in staging carcinoma of the male and female urethra.

Adrenal gland (Fig. 15.9) Normal adrenal glands appear as an inverted V or Y configuration. Most normal adrenal glands are well outlined by the retroperitoneal fat on axial T1-weighted images; this may be augmented by thin sectioning or coronal or sagittal views. The gland appears hypointense to the liver and isointense to skeletal muscle. T2-weighted images are unnecessary if the adrenal glands appear normal on high quality T1-weighted images; differentiation of the adrenal cortex from the medulla is generally not possible on routine MR images. Out-of-phase and in-phase chemical shift gradient echo pulse sequences can differentiate benign adrenocortical adenomas from metastases. Benign adrenal adenomas naturally contain a moderate amount of intracellular lipid (lipid rich adenoma), whereas metastases do not. Out-of-phase MR images result in a cancellation of signal in lipid-rich benign adenomas because the signal contributions from intracellular lipid and water oppose each other and cancel the resultant signal within the pixel. The characteristic MR findings of phaeochromocytomas include extremely high signal intensity on T2-weighted images and increased contrast

enhancement. Unfortunately, not all phaeochromocytomas have these imaging characteristics.

References Abu-Alfa, A. K. (2011). Nephrogenic systemic fibrosis and gadolinium-based contrast agents. Adv Chronic Kidney, 18, 188–92. Aslam Sohaib, S. A., Teh, J., Nargund, V. H., et al. (2002). Assessment of tumor invasion of the vena caval wall in renal cell carcinoma cases by magnetic resonance imaging. J Urol, 167, 1271–5. Bokacheva, L., Rusinek, H., Zhang, J. L., et al. (2009). Estimates of glomerular filtration rate from MR renography and tracer kinetic models. J Magn Reson Imaging, 29, 371–82. Cohan, R. H., Jafri, S., Choyke, L. P., et al. (2010). Manual on Contrast Media, Version 7. Reston, VA: American College of Radiology. Cowper, S. E., Robin, H. S., Steinberg, S. M., et al. (2000). Scleromyxoedema-like cutaneous diseases in renal-dialysis patients. Lancet, 356, 1000–1. Fain, S. B., King, B. F., Breen, J. F., et al. (2001). High-spatial-resolution contrast-enhanced MR angiography of the renal arteries: a prospective comparison with digital subtraction angiography. Radiology, 218, 481–90. Glockner, J.F., Takahashi, N., Kawashima, A., et al. (2010). Non-contrast renal artery MRA using an inflow inversion recovery steady state free precession technique (Inhance): comparison with 3D contrast-enhanced MRA. J Magn Reson Imaging, 31, 1411–18. Ho, V. B., Allen, S. F., Hood, M. N., et al. (2002). Renal masses: quantitative assessment of enhancement with dynamic MR imaging. Radiology, 224, 695–700. Hoeks, C. M., Barentsz, J. O., and Hambrock, T. (2011). Prostate cancer: multiparametric MR imaging for detection, localization, and staging. Radiology, 261, 46–66. Miyazaki. M. and Akahane, M. (2012). Non-contrast enhanced MR angiography: established technique. J Magn Reson Imaging, 35, 1–19. Notohamiprodjo, M., Reiser, M. F., and Sourbron, S. P. (2010). Diffusion and perfusion of the kidney. Eur J Radiol, 76, 337–47. Pedrosa, I., Sun, M. R., Spencer, M., et al. (2008). MR imaging of renal masses: correlation with findings at surgery and pathologic analysis. Radiographics, 28, 985–1003. Reznek, R. H. (2004). CT/MRI in staging renal cell carcinoma. Cancer Imaging, 14, 25–32. Schoenberg, S. O., Knopp, M. V., Londy, F., et al. (2002). Morphologic and functional magnetic resonance imaging of renal artery stenosis: a multireader tricenter study. J Am Soc Nephrol, 13, 158–69. Takahashi, N., Glockner, J, F., Kawashima, A., et al. (2010). Gadolinium enhanced magnetic resonance urography for upper urinary tract malignancy. Urology, 183, 1330–6.

CHAPTER 16

Radioisotopes in diagnostic imaging in nephrology Ramya Dhandapani and Sobhan Vinjamuri Introduction to nuclear medicine Nuclear medicine uses functional techniques to assess the different physiological processes of the kidneys. The early understanding of kidney function relied heavily on nuclear medicine imaging. Advances in other modalities such as ultrasound, computed tomography (CT), and magnetic resonance imaging (MRI) are mow preferred for identification of structural abnormalities. There are still some fundamental clinical questions that can be quickly and conclusively answered by isotope imaging techniques such as renal plasma flow, glomerular filtration rate (GFR), renal transit time, and assessment of outflow obstruction. Although the exposure to ionizing radiation should be considered, risk:benefit analysis in individual patients usually favours the clinical value of a test justifying a small additional radiation burden.

Introduction to radioactivity Atoms of all elements are composed of known arrangements of protons, neutrons, and electrons which characterize them as individual nuclides. Nuclides containing the same number of protons, the same atomic number, and the same chemical properties are known as isotopes. The most prevalent isotopes are usually stable and comprise naturally occurring elements. Radioisotopes are unstable members of the group of radioisotopes of an element. The nuclei of radioisotopes undergo rearrangement and change to a stable form, emitting radiation in the process. A radionuclide is a specific radioactive atom, designated by indicating the element and its atomic mass such as iodine-131 or technetium (Tc)-99m. The radiation emitted by a radionuclide enables the detection of extremely small masses, below the limits of chemical detection. It is thus possible to use radionuclides as true tracers for substances without introducing excessive amounts. The radiation emitted by radionuclides is characteristic and unique in terms of the rate of decay, type of radiation, and the energy of that radiation. Radionuclides used in nuclear medicine may emit alpha particles, beta particles, or gamma radiation. Alpha particles are large particles that are emitted from the nucleus with high energy levels. It is a charged particle with a charge of +2 as it has lost two of its electrons and is essentially a helium ion. Beta particles are similar in terms of charge and other physical properties to electrons. While ordinary electrons are found in the electronic shells orbiting the nucleus, beta-minus particles are emitted from the nuclei. They have high kinetic energy. Gamma rays are analogous to energy

packets emitted from the nucleus as part of its emissions to enable the achievement of a more stable physical state. Alpha and beta emitters are commonly used for radionuclide therapy. Gamma rays are more readily transmitted through tissue and therefore the internal administration of gamma emitting radionuclides allows external measurements and imaging of patients using a gamma camera. When the radionuclides are combined with a chemical or a pharmaceutical compound with particular physiological properties, the resultant compound is a radiopharmaceutical. These compounds are subject to strict pharmaceutical controls as is every other medicine suitable for human use. Clinical nuclear medicine usually involves the detection and quantification of ionizing radiation emitted from radioactive substances. A gamma camera produces an image corresponding to the distribution of radioactive substances in the body. Computer display and enhancement of the images with numerical assessment is frequently employed. Tc-99m is the most commonly used radionuclide in nuclear medicine imaging. It has a half-life of 6. 02 hours and this allows patients to attend for outpatient procedures and travel to home after the procedure without any strict radiation precautions. It has a gamma ray energy of 140 keV which is optimally primed for use with the modern gamma camera. The image quality is therefore of a very high quality. It can be readily complexed with a range of compounds to assess different physiological functions in the body and does not have any pharmacological action by itself. It is not toxic and does not elicit an immune response when injected into humans.

Radiation exposure and effective dose Radiation is a property of matter that is all around us and is much more common than people realize. Radiation is analogues to the passing of energy through matter. Most of the radiation does not change the environment it passes through and is called non-ionizing radiation. However, when radiation interacts with the matter as it passes through, then it is called ‘ionizing radiation’. When the energy associated with the radiation is deposited in a particular tissue it is termed as ‘absorbed dose’. This property is useful when a therapeutic benefit is required, however, in most diagnostic settings, this is not helpful and the risk to the individual should be weighed against the benefit of a result that can have a useful impact on patient management.

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Effective dose is an estimate of the total radiation burden to the patient from the exposure to the radioactive compound. This is a useful term that also allows comparison of relative radiation exposure across all modalities involving exposure to different forms of ionizing radiation including X-rays, CT scans, nuclear medicine procedures, and positron emission tomography (PET) scans. Absorbed organ doses in patients and effective doses are not measurable quantities but are based on estimates. Most of what is known about the carcinogenic effects of radiation is derived from studies of atomic bomb survivors. There are additional studies involving medical uses of radiation which have provided some epidemiological data. Radiation exposure is commonly known to be associated with some dose-dependent toxicities such as hair loss and acute radiation sickness. The development of cancer was believed to be a non-dose-dependent association of radiation. However, recent studies have strongly suggested a dose-dependent increase in cancer mortality. Some cancers are caused in children when the exposure occurred during pregnancy (teratogenic), or the risk has passed on to the next generations when the exposure occurred while the patient was not pregnant (genetic).

Principles of imaging the renal tract using radionuclides The three broad functions of the kidney that can be assessed using radionuclides include glomerular filtration, tubular secretion, and cortical function/activity. Agents that are physiologically inert and cleared exclusively by glomerular filtration are used to assess the GFR. These include chromium (Cr)-51 ethylenediaminetetraacetic acid (EDTA) and Tc-99m diethylenetriaminepentaacetic acid (DTPA). Tc-99m MAG3 is a tubular agent that has a higher extraction efficiency from the blood and is easy to prepare. Tc-99m dimercaptosuccinic acid (DMSA) is predominantly a cortical agent that provides an index of cortical function or activity (see Table 16.1).

Routine clinical indications for isotope renography (See Table 16.2) ◆ Assessment of differential kidney function ◆ Assessment of possible pelviureteric obstruction ◆ Assessment of renal damage post pyelonephritis ◆ Renal transplant imaging ◆ Assessment of vesicoureteric reflux ◆ Assessment of renal artery stenosis.

Patient preparation The most important prerequisite for the renogram is an adequate state of hydration of the patient which has to be maintained during the test. It is important to avoid an oliguric state since the result can (because of sluggish urine flow) mimic an obstructive pattern. The bladder is emptied immediately before the examination and the patient is positioned in a sitting or prone position with the gamma camera against their back. It is important to position the patient comfortably as they need to maintain the same posture for 20–40 minutes. After intravenous injection of the appropriate dose of the radiopharmaceutical, images are acquired for 20–40 minutes. The patient is then asked to empty their bladder at the end of the procedure to reduce the radiation dose to the pelvic organs.

Interpretation Studies are analysed by producing clear, summed computer images and defining regions of interest over each kidney and the urinary bladder. Curves are then obtained from the detected count rate against time for each region and the obtained curves are expressed as a percentage of the injected dose. The normal renogram shows three classic phases. Immediately following

Table 16.1  Some common isotopes and their routine clinical indications. Please see text for further details Radioisotope

Common indication(s)

Comment

Cr-51 EDTA

Glomerular filtration

Gold standard for estimation of renal function for research, oncology, and transplant donor evaluation

Tc-99m MAG3

Differential renal function Effective renal plasma flow Captopril renography Transplant renography

Useful to assess function and upper tract obstruction

Tc-99m DMSA

Renal scarring and reflux

Provides information on relative renal function, particularly when kidneys are rotated or at different depths

Tc-99m DTPA

Glomerular filtration

Used in non-obstructed kidneys

F-18 fluoride PET-CT scans

Staging of cancers

Estimate of osteoblastic bone lesions

F-18 FDG PET-CT scans

Screening tool

In systemic illness such as PUO, immunocompromised, vasculitis, metastases, and to assess tumour recurrence post nephrectomy It can also be used to stage post-transplant lymphoproliferative disease

Ga-67 citrate, In-111-labelled leucocytes

Suspected infection

Intense uptake in infection of the kidneys

Table 16.2  Some common clinical questions and suggested procedures. Please see text for further details Clinical question(s)

Procedure to request

Comment

Need accurate estimate of relative renal function

DMSA renal scan

In situations where accuracy is paramount or where renographic estimate is likely to be difficult (e.g. in infants). No information is provided on the status of outflow tract

Suspected upper tract obstruction

Diuresis MAG3 renogram

Estimate of relative function will be routinely provided

Suspected renal scarring

DMSA renal scan

Estimate of relative renal function will be routinely provided

Need accurate estimate of absolute GFR in mL/min

GFR measurement (Cr-51 EDTA)

Could also use Tc-99m DTPA

Need estimate of both relative and absolute GFR

Basic DTPA renogram with GFR

In principle, both can be measured by a single injection of Tc-99m DTPA. However, in children (or adults with compromised renal function) it is preferable to inject Tc-99m MAG3 and Cr-51 EDTA simultaneously. Absolute GFR is measured by blood sampling

Assessment of possible renovascular hypertension

Captopril renogram

Both DTPA and MAG3 can be used

Higher confidence in possibility of acute tubular necrosis

MAG3 renogram

Good perfusion, progressive uptake, and no excretion

Suspected kidney infection

In-111 WBCs or Ga-67 citrate

In-111 white blood cells are not normally excreted by the renal tract. Hence any renal uptake is usually abnormal

Suspected systemic condition such as vasculitis or carcinoma

F-18 FDG PET-CT scanning

FDG PET-CT scans have high sensitivity but lower specificity. Hence further focused testing of abnormal areas is required for confirmation

(A) RENOGRAM

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Left Kidney Rel Uptake 50.9 Right Kidney Rel Uptake 49.1

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Fig. 16.1  (A) Renogram showing normal tracer activity in the bladder. (B) Graph showing normal tracer activity in the bladder.

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Left Kidney Rel Uptake 15.5 Right Kidney Rel Uptake 84.5

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Fig. 16.2  (A) Renogram showing abnormal tracer activity in the bladder. (B) Graph showing abnormal tracer activity in the bladder.

the intravenous injection of the radiopharmaceutical, there is a rapid rise which reflects the vascular supply to the kidney. The second phase is a more gradual slope which corresponds to the renal handling of the tracer by the kidneys and is dependent on various factors such as supply rate, extraction efficiency, intra luminal transit, and excretion. In a normal kidney, the curve reaches a peak at 2–5 minutes and activity starts to leave the renal area which is the beginning of the third phase. At this point the tracer activity starts to appear in the bladder and this is now predominantly the excretory phase. See Fig. 16.1 for normal appearances and Fig. 16.2 for examples of abnormal uptake and excretion.

Limitations of routine isotope renography The value of assessing function and physiology of the renal tract by isotope renography is tried and tested and the benefits usually outweigh the small radiation-associated risks. Because the dynamic phase of renal function needs to be assessed, it is vital that the radioactive tracer or any related medication such as diuretics should not be extravasated into the interstitial tissue at the site of intravenous injection. Insufficient hydration, either self-induced or a result of medication or co-morbidities, can result in delayed uptake and excretion by one or both kidneys and may be misinterpreted as poor function. Significant patient motion during the study may introduce errors in the activity–time curve analysis.

DTPA versus MAG3 renography Tc-99m-labelled DTPA is a physiologically inert compound that is predominantly excreted via the glomeruli (approximately 90% excreted by 4 hours). Up to 10% of the injected activity is bound by plasma proteins and is not available for excretion and therefore the net value may represent a slight underestimation. It does not get secreted or filtered by the tubules and also does not localize to the parenchyma/cortex. Tc-99m labelled MAG3 is a predominantly tubular secretory agent (95% tubular secretion vs 5% glomerular clearance). Although we cannot measure GFR with this agent, another aspect of renal function, that is, effective renal plasma flow can be measured and this represents a surrogate marker of global renal function. Because of the relative ease of preparation of Tc-99m MAG3, better visualization of the renal parenchyma, unpredictable nature of protein binding of Tc-99m DTPA, better image quality, and relative reproducibility of tubular function at poorer renal function, Tc-99m MAG3 is, for most routine indications, the preferred test over Tc-99m DTPA renography.

Choosing MAG3 or DMSA Tc-99m DMSA scanning provides information mainly on cortical or parenchymal activity. This has been widely considered a better test to estimate differential kidney function mainly because it is a parenchymal agent and routine imaging includes anterior as well as posterior. It is particularly useful when both kidneys are located at different depths or rotated with respect to one another. It is also the

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radioisotopes in diagnostic imaging in nephrology

Fig. 16.3  Tc-99m DMSA scanning example.

preferred agent when one kidney is functioning poorly and there is a requirement to assess the differential function to a higher degree of accuracy. Tc-99m DMSA is clearly the agent of choice in suspected pyelonephritis and to assess the extent of cortical damage or scarring. See Fig. 16.3. Tc-99m MAG3 is the preferred agent of choice for any assessment of excretion or possible obstruction. The early parenchymal phase of the Tc-99m MAG3 renogram can be used to assess the parenchymal integrity and the differential function. In routine clinical practice, where there is an expectation that both kidneys are located at the same depth and morphologically similar, Tc-99m MAG3 does provide a good estimate of the renal function. Due to the high radiation burden associated with Tc-99m DMSA renography and the fact that Tc-99m MAG3 provides additional information about renal function, Tc-99m MAG3 can be used to provide an index of differential function in most clinical situations.

Differential renal function Biochemical tests provide a good estimate of overall renal function, but it is useful to know individual kidney function in selected situations such as assessment of potential living kidney donors. Anatomical tests such as an ultrasound and CT rely on size symmetry between the two kidneys and in most cases this should be adequate. However, it is now well accepted that even if the kidneys have the same size, they may contribute differently to the overall renal function. It is important to quantify this asymmetry and isotope tests provide quick, easy and reproducible information.

All three of the commonly used isotopes (Tc-99m DTPA, Tc-99m MAG3, and Tc-99m DMSA) can be used to provide an estimate of the differential function. However, in routine clinical practice, Tc-99m MAG3 is preferred and in patients with known asymmetry of depth or rotated kidneys, Tc-99m DMSA is preferred.

Chromium-51 EDTA estimation GFR is the accepted standard measure of renal function. It is routinely measured using tracers that are cleared exclusively by glomerular filtration, the most common being Cr-51 EDTA and Tc-99m DTPA. GFR is best assessed by Cr-51 EDTA. Clearance values are typically 85% of those using inulin which although gold standard is difficult and time-consuming and is inappropriate for routine clinical use. Since Cr-51 EDTA is expected to be excreted solely by the glomerular filtration route, it has been shown that the rate of disappearance of this compound from the blood is proportional to the renal clearance. Hence there is no need to obtain urinary samples. The commonly used protocols involve at least two timed venous blood samples at 2–3 hours post intravenous injection of the radioactive tracer. The more samples that are obtained, the higher the mathematical accuracy. It is commonly used for monitoring the effect of nephrotoxic drugs, calculating dose in chemotherapy, detection of renal failure with inconclusive serum creatinine values, and in the assessment of potential live donors for transplantation. The values should be interpreted with caution in patients with ascites, oedema, or other expanded body fluid space and in patients receiving intravenous fluids where the renal clearance value may be overestimated. Final values may be corrected for body surface area

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2.1

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Left Kidney Rel Uptake 47.4 Right Kidney Rel Uptake 52.6

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%

U p t a k e

U p t a k e

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Fig. 16.4  (A) Graph showing the captopril phase. (B) Graph showing the captopril phase 2.

rather than for weight, as in the paediatric population, the body surface area corrected GFR increases with age from birth up to 2 years and remains constant into adulthood thereafter.

Captopril renography Captopril renography has been of proven value in the identification of renovascular hypertension in patients with high blood pressure. This test also has a value in the evaluation of asymmetrically sized kidneys and to confirm or exclude functional impairment secondary to renal artery stenosis. The test has two phases: a ‘baseline phase’ to assess the baseline differential function and excretory patterns; a second ‘Captopril phase’ study is done on another day after oral administration of captopril and repeating the renogram. A reduction in renal uptake (unilateral reduction by at least 10%) or prolonged retention of the radiotracer in the renal parenchyma after administration of captopril in comparison to the baseline study is considered a positive indicator of renal artery stenosis causing renovascular hypertension. The captopril phase can also be performed with other angiotensin-converting enzyme inhibitors such as lisinopril or enalapril and the longer-acting medication does not need to be stopped for the ‘captopril phase’ (see Fig. 16.4).

Renal failure Tc-99m MAG3 is the isotope of choice in renal failure as better renal extraction improves visualization of the kidneys. The pattern of the renogram curve can be helpful to identify the cause of acute kidney injury. Prerenal causes will affect the initial perfusion part of the curve. Renal parenchymal diseases affect the tubular function part of the curve and postrenal causes affect the excretory part of the curve. In acute arterial occlusion, there is severely impaired blood flow, nil or poor tracer uptake, with poor or absent excretion. Acute renal vein thrombosis shows an enlarged kidney with poor perfusion and prolonged cortical tracer retention. In causes of prerenal failure such as prolonged hypoperfusion or hypotension, there is normal tracer uptake with delayed excretion and drainage. Parenchymal diseases such as acute glomerulonephritis show symmetrically enlarged kidneys with poor MAG3

clearance. Acute tubular necrosis and urinary tract obstruction show good tracer uptake in the flow phase, as the arterial flow is well established, but no tracer excretion. Correlation with other imaging modalities such as ultrasound/CT may then exclude the presence of a dilated collecting system and aid diagnosis. Renograms in patients with chronic renal failure usually demonstrate small kidneys with little or no tracer uptake, prolonged excretion, and poor drainage.

Renography in transplanted kidneys The main indication for an isotope renogram in post-transplant evaluation is the differentiation of acute tubular necrosis from rejection. Rejected kidneys have poor perfusion as well as poor tubular excretion, resulting in poor visualization of the transplant kidney on the renogram. In acute tubular necrosis, there is normal perfusion/visualization of the transplant kidney but the excretory phase shows no, or minimal, excretion. The urinary bladder is frequently not visualized despite the kidney being visualized. Unfortunately ciclosporin toxicity can also appear similar on a functional aspect and this common differential diagnosis needs exclusion when a transplant renogram is suggestive of acute tubular necrosis. Serial isotope renograms post transplant are helpful in identifying subacute and chronic rejection. See Fig. 16.5.

SPECT-CT and PET-CT scans SPECT (single-photon emission computed tomography) uses radioisotopes that emit gamma rays which are then measured directly using a gamma camera. PET involves injection of a known amount of positron-emitting radiopharmaceuticals and imaging of the whole body or specific organs using a PET scanner. Due to the need for anatomical localization of sites of abnormal tracer activity on PET scan, there was recognition of the need to develop fusion imaging whereby anatomical context can also be provided. Initial efforts were focused on registering PET and CT data obtained separately and using advanced computer software to merge or fuse the two scans. However, this approach was superseded by combining the PET and CT technologies into one

Chapter 16 

(A)

radioisotopes in diagnostic imaging in nephrology (B) 10.5

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Fig. 16.5  (A) Post-transplant renogram showing subacute and chronic rejection. (B) Graph showing post-transplant subacute and chronic rejection.

scanner. The patient therefore undergoes one examination at one sitting and there is minimal patient motion and other technical problems. The CT scan is typically done without intravenous contrast medium enhancement. More recently combined PET-MRI imaging has been introduced with the added advantage of reducing radiation exposure. PET-CT scanners now form the main bulk of functional scanners, with manufacturers slowly phasing out PET only scanners and promoting advances in PET-CT and PET-MRI scanning. One limitation of current PET/CT technology is that data are acquired sequentially rather than simultaneously. This then limits the ability to study functional processes that change between the two scans. Another factor for consideration is the radiation dose to patients. Artefacts caused by patient motion or organ motion between the two sets of images coupled with different breathing protocols for PET and CT can result in problems for data interpretation. Mis-registration between the two sets of images is a recognized disadvantage which is factored in while reporting PET-CT scans. Data acquisition with new hybrid PET-MRI scanners provides the option of real-time simultaneous studies. MRI offers better soft tissue contrast and the challenge of developing viable PET-MRI systems focused on addressing apparent technical incompatibilities such as the fact that PET detectors are based on scintillation crystal blocks which are highly sensitive to even small magnetic fields; and space constraints of fitting in two technologies in one ergonomic system suitable for patient use and convenience. Therefore the

type of sequential imaging performed with PET-CT is not a viable option for PET-MRI and the way forward is to combine both systems into one gantry system.

Renal infection and inflammation Tc-99m DMSA scintigraphy is a highly sensitive and specific tool to detect and confirm renal involvement associated with lower urinary tract infections, especially in the paediatric population. Renal sequelae such as scarring secondary to vesicoureteric reflux can also be assessed in the post -acute or chronic phases. Tc-99m DMSA scintigraphy is recommended for the evaluation of the extent of scarred renal tissue in both children and adults with chronic pyelonephritis. SPECT scanning has been shown to increase the diagnostic confidence for detecting cortical defects associated with pyelonephritis. Renal abscess and focal pyelonephritis may appear as a focal defect with increased uptake in the vascular phase. CT and ultrasound are usually used for correlation. Pyonephrosis on the renal scintigraphy will show no tracer uptake as there is very little viable renal tissue. Gallium (Ga)-67 citrate and indium (In)-111-labelled leucocytes are used in imaging for suspected renal infections. Ga-67 shows non-specific uptake in the kidneys, but has been shown to be particularly useful in acute focal bacterial nephritis which has non-specific ultrasound and CT findings. This commonly occurs in diabetic patients and shows unilateral or multifocal areas of

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Fig. 16.6  Whole-body FDG PET-CT scan.

increased uptake in the kidneys. Unlike Ga-67, In-111-labelled leucocyte scans do not show activity within the urinary tract and are more specific for acute renal infections. These scans can differentiate perinephric abscess from pyelonephritis. Whole-body fluorodeoxyglucose (FDG) PET-CT scans can be used as a screening tool to detect active infection or inflammation involving either the kidney alone or as part of a systemic process in conditions such as vasculitis, glomerulonephritis, polycystic kidney disease, pyrexia of unknown origin (PUO), and in immunocompromised patients. In PUO, PET-CT provides useful diagnostic information in about 40% of patients. Active FDG uptake is seen in the blood vessels affected by vasculitis for which PET has positive predictive value of 93% and negative predictive value of 80%. See Fig. 16.6.

Post-transplant lymphoproliferative disorder FDG PET-CT scan at diagnosis of post-transplant lymphoproliferative disorder will show uptake in all affected lesions and may result in the detection of lesions not seen at conventional CT. This may upstage the disease. Repeating the scan after treatment will also help with prognosis as FDG uptake will have gone from all successfully treated lesions. Persistence of uptake indicates a partial response to treatment. Absence of any uptake in all lesions is a strong predictor of remission. FDG PET-CT has the advantage over conventional contrast medium-enhanced CT in avoiding contrast-related nephrotoxicity in patients with impaired renal function.

Renal tumours Most renal tumours are now detected incidentally when patients undergo cross-sectional imaging for other indications. Nuclear

Fig. 16.7  Example of very high levels of glucose metabolism in FDG PET-CT scan.

medicine and PET tracers are not specific for routine use in the detection and diagnosis of renal cell cancers. FDG hyperactivity is not specific for malignancy. Extensively necrotic primary tumours and lymph nodes show low FDG uptake and may be missed on PET-CT scans. However, when the abnormalities are marked, such as very high levels of glucose metabolism, then there is a higher likelihood of malignancy. See Figs 16.7 and 16.8. FDG PET-CT scans are extremely useful to identify extrarenal sites of renal cancer, tumour recurrence post nephrectomy, and lymph node involvement. Renal metastases are predominantly lytic and may not be detected as hot spots on the Tc-99m methylene diphosphonate isotope bone scan. Scanning with 18-Fluoride is particularly effective in the detection of distant bone metastases thereby influencing therapeutic decisions for staging, assessing recurrence, or new metastasis after initial therapy. See Fig. 16.9.

Renal trauma CT is the gold standard in trauma imaging. Although renal scintigraphy is not routinely performed in the acute setting, it is useful in patients with mild renal injury to monitor and follow-up renal function, presence of urine leak, or acute renovascular compromise. Haematomas are seen as photopenic areas on the renogram. Severe contusions appear as a small functioning kidney while mild contusions may be demonstrated as an enlarged kidney. Persistent focal photopenic defects on follow-up scans are usually areas of infarction. MAG3 renograms have been shown to identify urine leak post renal transplant surgery and post intervention procedures such as percutaneous stenting of renal arteries. However, Tc-99m DTPA

Chapter 16 

radioisotopes in diagnostic imaging in nephrology

DMSA planar versus SPECT Tc-99m DMSA is the isotope of choice to demonstrate the cortical integrity of the kidneys. SPECT is used in correlation with Tc-99m DMSA and other imaging modalities in specific conditions which need to differentiate pathological from non-pathological renal tissue. When Tc-99m DMSA renography shows a cortical abnormality such as a pseudotumour, that is, column of Bertin, dromedary hump, or fetal lobulation, SPECT is useful to demonstrate normal renal tissue uptake at that site, helping to confirm the diagnosis. Photopenic defects on the Tc-99m DMSA may be due to reflux or scarring, cysts, abscess, or neoplasms. These are shown as a true cortical defect on SPECT which then can be evaluated further with other imaging techniques as CT or ultrasound.

Conclusion Nuclear medicine techniques provide a range of diagnostic options in assessing the various physiological and pathological processes of the kidney.

Further reading

Fig. 16.8  Very high levels of glucose metabolism in FDG PET-CT scan in a patient with malignancy.

identifies urine extravasation from the kidneys better than Tc-99m MAG3 which may miss a small subtle leak. There have been case reports in which SPECT scan was useful to identify the exact site of urine leak post renal transplant due to the added advantage of anatomical correlation.

Fig. 16.9  Scanning with fluorine-18 shows bone metastasis.

Achong, D. M. and Tenozrio, L. E. (2003). Abnormal MAG3 renal scintigraphy resulting from dehydration. Clin Nucl Med, 28, 683–4. Aktas, A., Aras, A., Colak, T., et al. (2006). Comparison of Tc-99m DTPA and Tc-99m MAG3 perfusion time-activity curves in patients with renal allograft dysfunction. Transplant Proc, 38, 449–53. Alavi, A., Mavi, A., Basu, S., et al. (2007). Is PET-CT the only option? Eur J Nucl Med Mol Imaging, 34, 819–21. Bar-Shalom, R., Yefremov, N., Guralnik, L., et al. (2006). SPECT/CT using 67Ga and 111In-labelled leukocyte scintigraphy for diagnosis of infection. J Nucl Med, 47, 587–94. Bianchi, E., Pascual, M., Nicod, M., et al. (2008). Clinical usefulness of FDG-PET/CT scan imaging in the management of posttransplant lymphoproliferative disease. Transplantation, 85(5), 707–12. Bleeker-Rovers, C. P., Van Der Meer, J. W., and Oyen, W. J. (2009). Fever of unknown origin. Semin Nucl Med, 39, 81–7. Broekuezen-de Gast, H. S., Tiel-Van Buul, M. M. C., and Van Beek, E. J. R. (2001). Severe hypertension in children with renovascular disease. Clin Nucl Med, 26, 6–609. Caglar, M., Moretti, J. L., Buchet, P., et al. (1998). Enalapril plus frusemide MAG3 scinitgraphy in hypertensive patients with atherosclerosis and moderate renal insufficiency. Nucl Med Commun, 19, 1135–40. Dondi, M., Fanti, S., and De Fabritis, A., et al. (1992). Prognostic value of captopril renal scintigraphy in renovascular hypertension. J Nucl Med, 33, 2040–4. Dondi, M., Monetti, N., Fanti, S., et al. (1991). Use of technetium-99m-MAG3 for renal scintigraphy after angiotensin converting enzyme inhibition. J Nucl Med, 32, 424–8. Esteves, F. P. Taylor, A., Maantunga, A., et al. (2006). 99Tcm MAG3 Renography: normal values for MAG3 clearance and curve parameters, excretory parameters, and residual urine volume. AJR, 187, W610–7. Fanti, S., Dondi, M., Corbelli, C., et al. (1993). Evaluation of hypertensive patients with a solitary kidney using captopril renal scintigraphy with Tc-99m-MAG3. Nucl Med Commun, 14, 969–75. Freeman, L. and Blaufox, M. D. (1999). Renal nuclear medicine including consensus reports. Semin Nucl Med, 29, 146–88. Goldberg, M. A., Mayo-Smith, W. W., Papanicolau, M., et al. (1997). FDG PET characterisation of renal masses: preliminary experience. Clin Radiol, 52, 510–15. Kahn, D., Ben-Haim, S., Bushnell, D. L., et al. (1994). Captopril enhanced Tc-99m MAG3 renal scintigraphy in subjects with suspected renovascular hypertension. Nucl Med Commun, 15, 515–28.

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Prigent, A., Cosgriff, P., Gates, G. F., et al. (1999). Consensus report on quality control of quantitative measurements of renal function obtained from the renogram: International Consensus Committee from the Scientific Committee of Radionuclides in Nephrourology. Semin Nucl Med, 29, 146–59. Rajadhyaksha, C. D., Parker, J. A., Barbaras, L., et al. (2005). Normal and Benign Pathologic Findings in 18-FGD-PET and PET-CT. An Interactive Web Based Image Atlas. Boston, MA: Joint programme in Nuclear Medicine, Harvard Medical School. Ramdave, S., Thomas, G. W., Berlangieri, S. U., et al. (2001). Clinical role of F-18 fluorodeoxy glucose positron emission tomography for the detection and management of renal cell carcinoma. J Urol, 166, 825–30. Rossleigh, M. A. (2001). Renal cortical scintigraphy and diuresis renography in infants and children. J Nucl Med, 42, 91–5. Safaei, A., Figlin, R., Hoh, C. K., et al. (2002). The usefulness of F-18 deoxyglucose whole body positron emission tomography (PET) for re-staging of renal cell cancer. Clin Nephrol, 57, 56–62. Sandler, M. P., Coleman, R. E., Patton, J. A. (2002). Diagnostic Nuclear Medicine (4th ed.). Philadelphia, PA: Lippincott, Williams and Wilkins. Schreij, G., van Es, P. N., van Kroonenburgh, M. J. P. G., et al. (1996). Baseline and postcaptopril renal blood flow measurements suspected of renal artery stenosis. J Nucl Med, 37, 1652–5. Seto, E., Segall, G. M., and Terris, M. K. (2000). Positron emission tomography detection of osseous metastasis of renal cell carcinoma not identified on bone scan. Urology, 55, 286. Sfakianakis, G. N., Al-Skeikh, W., Heal, A., et al. (1982). Comparison of scintigraphy with In-111 leucocytes and Ga-67 in the diagnosis of occult sepsis. J Nucl Med, 23, 618. Shao, Y., Cherry, S. R., Farahani, K., et al. (1997). Simultaneous PET and MR imaging. Phys Med Biol, 42, 1965–70. Wagner, R. H., Karesh, S. M., and Halama, J. R. (eds.) (1999). Questions and Answers in Nuclear Medicine. St Louis, MO: Mosby.

CHAPTER 17

Immunological investigation of the patient with renal disease Jo H. M. Berden and Jack F. M. Wetzels Introduction The kidney is an important site of immune injury mediated by both the innate and adaptive immune systems. Our understanding of the pathogenesis of immunological renal diseases has been improved by study of the time course of changes in kidney morphology in animal models. A good example is the renal injury that occurs after injecting bovine serum albumin (BSA)-anti-BSA complexes into rabbits, a model of serum sickness, and a prototypical example of an immune complex disease. It is now well established that many primary glomerular diseases are immune mediated. Furthermore, the kidney is the target in various infectious diseases, systemic auto-immune disorders and the vasculitides (Couser, 2012; Nasr et al., 2013). Renal injury may be evoked by humoral (antibody) or cellular immune responses. Three mechanisms can be defined. The kidney may be the target of antibodies that are directed against kidney-specific antigens, exciting a type II immunological reaction. Anti-glomerular basement membrane (GBM) disease, caused by antibodies directed against collagen IV, an intrinsic component of the GBM and membranous nephropathy, caused by antibodies directed at podocyte antigens are the best known examples. More often, the kidney is involved in a type III immunological reaction caused by the deposition of immune complexes. These immune complexes may either be formed in the circulation or in situ, by binding of an antibody to an antigen already planted in the glomerular capillary wall. The size, charge, and composition of the immune complexes determine their final localization. The site at which these deposits are formed will determine the pathological changes. Mesangial deposits induce mesangial matrix expansion and haematuria; subendothelial deposits a proliferative glomerulonephritis with capillary loop necrosis and crescent formation; while subepithelial deposits mostly evoke the nephrotic syndrome. The role of cellular immunity in inducing glomerular pathology is less well understood. In glomerular diseases such as minimal change disease, focal segmental glomerulosclerosis, and antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis injury is probably also mediated by cellular responses. Although in vasculitis various auto-antibodies may be present in the circulation there is no evidence of antibody or complement deposition in the kidneys. Laboratory assays which evaluate the immune response, in particular the identification of (auto)-antibodies are valuable tools in establishing a diagnosis and/or monitoring of the activity of the

disease. An alphabetical overview of various immunological markers and associated renal diseases is given in Table 17.1.

Laboratory techniques These assays depend on a number of laboratory techniques. The practising clinician should be aware that test results are dependent on the technique used and be aware of pitfalls.

Electrophoresis Electrophoresis is a simple technique, where a mixture of proteins is applied to an agar gel and separated by applying a charge over the gel. Protein electrophoresis of serum shows a typical pattern with separation of albumin, α1, α2, ß, and γ globulins. More specific assays are based on the principle that antigen and antibody form precipitates. Examples are immunoelectrophoresis or immunofixation, where proteins are separated by charge and then precipitated by antibodies that diffuse into the gel. If specific antibody-antigen reactions occur these are visible as precipitation lines in the gel. In immunonephelometry the antigen and antibody form insoluble complexes in solution that are detected by light scatter. The specificity of these techniques is dependent on the specificity of the antibodies, which can be either monoclonal or polyclonal. When there is antigen excess the assays may provide negative results unless samples are appropriately diluted (Daval et al., 2007).

Indirect immunofluorescence With this technique, antibodies directed at tissue antigens can be visualized. Tissue slides (varying from kidney to specific cell lines) are exposed to various anti-sera. Antibodies directed to antigens that are expressed in the tissue/cells will bind. These antibodies are visualized by adding a fluorescent second antibody directed at human immunoglobulins. The fluorescent pattern and the tissue type provide clues to the nature of the involved antigens, for example, ANCA will cause on granulocytes either cytoplasmic or perinuclear staining and antinuclear antibodies (ANAs) will react with the nuclei of cell lines. Indirect immunofluorescence (IF) on kidney sections together with serological tests are essential tools to diagnose the cause of glomerulonephritis. The indirect IF technique is subject to several pitfalls: the specificity of the antigens is unclear, and most laboratories use only anti-immunoglobulin (Ig)-G antibodies as secondary antibodies, which will not detect IgA or IgM class antibodies.

Table 17.1  Overview of immunological markers in renal diseases Condition ♯

Associated systemic or renal disease

ANA

+

SLE; different auto-immune diseases

ANCA

+

Granulomatosis with polyangiitis; microscopic polyangiitis; renal limited vasculitis; eosinophilic granulomatosis with polyangiitis

Beta-2-glycoprotein 1

+

APS

Cardiolipin

+

APS

Centromere

+

Cutaneous sclerosis

C1q

+

SLE nephritis

DNAse B

+

PSGN

dsDNA

+

SLE

ENA

+

MCTD; Sjögren’s syndrome; scleroderma; SLE

α–fodrin

+

Sjögren’s syndrome

GBM

+

anti-GBM disease; Goodpasture syndrome

Histone

+

drug-induced SLE; SLE

MPO

+

See ANCA

Nucleosome

+

SLE nephritis

Phospholipid

+

See anti-cardiolipin/lupus anticoagulant

PLA2R

+

Membranous nephropathy

Proteinase 3

+

See ANCA

RNP

+

MCTD

SS-A (Ro)

+

SLE; Sjögren’s syndrome

SS-B (La)

+

Sjögren’s syndrome

Streptolysin O (ASO)

+

PSGN

Topo-isomerase I

+

systemic scleroderma

C1q

D

SLE (80%; ♀ > ♂)

CH50

↓/D

See C3/C4

C2

D

SLE (25%, ♀ > ♂), IgA nephropathy, HSP, MPGN type 1

C3

↓/M

SLE (25%, MPGN type 1, DDD, PSGN, aHUS

D

SLE (seldom)

↓

SLE, cryoglobulinaemia

D

SLE (75%)

Factor H

↓/D/M

aHUS, DDD

Factor B

↓/D/M

aHUS, DDD

Factor I

↓/D/M

aHUS, DDD

C3 nephritic factor

+

DDD, partial lipodystrophy

C5-C9

D

SLE (seldom)

+

Cryoglobulinaemia, SLE, Sjögren syndrome, lymphoma, HCV

Immunological marker Screening assays

Antibody specificity

Complement

C4

Miscellaneous Cryoglobulins

(Continued)

chapter 17 

immunological investigation of renal disease

Table 17.1 Continued Condition ♯

Associated systemic or renal disease

↑

IgA nephropathy

D

SLE

Lupus anticoagulant

+

APS, SLE

Paraproteins

+

Amyloidosis, MIDD, cast-nephropathy

vWF protease activity

↓

TTP

Immunological marker IgA

a Meaning of condition: + = present; D = deficiency; M = monitoring; ↓ = decreased; ↑ = increased.

ANA = antinuclear antibodies; ANCA = anti-neutrophil cytoplasmic antibodies; ASO = antistreptolysin O; D = deficiency; DDD = dense deposit disease; ENA = extractable nuclear antigens; GBM = glomerular basement membrane; HCV = hepatitis C virus; HSP = Henoch–Schönlein purpura; HUS = haemolytic uraemic syndrome; M = mutation; MCTD = mixed connective tissue disease; MIDD = monoclonal immunoglobulin deposition disease; MPGN = membranoproliferative glomerulonephritis; MPO = myeloperoxidase; PLA2R = M type phospholipase A2 receptor; PR3 = proteinase 3; PSGN = poststreptococcal glomerulonephritis; TTP = thrombotic thrombocytopenic purpura;

Enzyme-linked immunosorbent assay In the direct enzyme-linked immunosorbent assay (ELISA), antigens are coated on a plate. Antibodies will bind, and can be visualized with a ligand, mostly a secondary antibody labelled with an enzyme. The enzyme converts a colourless substrate into a chromogen that can be measured. In the ELISA, the sensitivity and specificity are determined by the nature and conformational shape of the coated antigens, the sort of ligand, and the epitope specificity of the antibody. The specificity and sensitivity of an ELISA can be increased by an indirect technique, the so-called antigen capture ELISA. In this technique, plates are coated with specific antibodies which bind the antigen under investigation. This technique allows the antigen to remain in its native conformation. Subsequently, antibodies reactive with the captured antigen can then be detected.

Immunoblotting This technique is used to further characterize the antigen specificity of an antibody. The protein mixture is separated depending on size or charge by gel chromatography and the molecules are then transferred to a nitrocellulose membrane. These blots are incubated with serum samples. After binding, antibodies are visualized by enzyme-labelled secondary antibodies. The blotted proteins are identified in a marker lane by labelled antibodies of known specificity, which allows by comparison determination of the antibody specificities in the sample under investigation.

Immunoglobulins Immunoglobulin A (See Chapters 65–69.) Immunoglobulin A  (IgA) comprises 15–20% of all serum immunoglobulins. In serum, most IgA is present in its monomeric form (molecular weight 160 kD). Two classes of IgA are recognized: IgA1 (80%) and IgA2 (20%). Plasma concentrations of IgA may be increased in up to 50% of patients with IgA nephropathy (D’Amico, 1988). Thus, normal values do not exclude the presence of IgA nephropathy. Moreover, elevated levels of IgA can be found in patients with hepatitis, liver cirrhosis, and Sjögren’s disease. A recent study compared serum IgA in Caucasian patients with IgA nephropathy and healthy controls (Moldoveanu et al., 2007). Although mean serum IgA level was increased in the patients, the sensitivity was only 33%. Although specificity was high at 90%,

the comparator group consisted of healthy controls and lacked patients with relevant comorbidity. Recent studies have pointed to the role of aberrantly glycosylated, galactose-deficient IgA1 (GD-IgA1) and the formation of glycan-specific anti-GD-IgA1 antibodies in the pathogenesis of IgA nephropathy. The importance of altered glycosylation is supported by the finding that this altered form of IgA is preferentially present in the mesangial IgA deposits of patients with IgA nephropathy (Allen et  al., 2001). GD-IgA1 is less effectively cleared by the sialoglycoprotein receptor, demonstrates increased binding and activation of mesangial cells, and activates complement through the mannose binding lectin pathway. Moldeveanu and colleagues used a lectin from the garden snail Helix aspersa that specifically binds GD-IgA1 to develop an ELISA assay for the measurement of serum GD-IgA1 (Moldoveanu et al., 2007). They reported increased levels (> 90th percentile of values in 150 healthy controls) of GD-IgA1 in 117 of 153 patients with IgA nephropathy and calculated a sensitivity of 76% and a specificity of 90% for predicting a diagnosis. In this study, serum GD-IgA1 did not correlate with clinical activity. In contrast, Zhao et al. evaluated 275 patients with IgA nephropathy and reported a higher risk of progressive renal failure in patients with higher levels of GD-IgA (hazard ratio 4.76 for the fourth quartile vs first quartile) (Zhao et  al., 2012). In IgA nephropathy the development of disease is dependent on the presence of anti-GD-IgA1 antibodies (mostly of the IgG class). Indeed, in a study of 60 patients with IgA nephropathy, serum levels of IgG anti-GD-IgA1 antibodies were increased and predicted the diagnosis with sensitivity of 88% and specificity of 95% (Suzuki et al., 2009). Moreover, the antibody titre correlated with proteinuria, which strongly supports their pathogenetic role. The association between auto-antibody levels and progression was confirmed recently (Berthoux et al., 2012). IgA deficiency is the most common form of Ig deficiency, occurring in 1 in 800 people. IgA deficiency is associated with an increased incidence of systemic auto-immune diseases including SLE.

Paraproteins Paraproteins are monoclonal immunoglobulins, synthesized and secreted by abnormal clones of B lymphocytes. Paraproteins can consist of the intact immunoglobulin or contain only one part of the immunoglobulin molecule, either the heavy chain (G, A, M, D, or E) or the light chain (κ or λ). The light chains are also known as Bence Jones proteins.

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Paraproteins can be recognized as a single band on serum or urine protein electrophoresis. However, a normal pattern of protein electrophoresis does not exclude the presence of a paraprotein. Protein electrophoresis has a low sensitivity for free light chains (FLC), the amount of paraprotein may be too small and the paraprotein may be masked in the ß-globulin band (Heher et al., 2010). Therefore, if a paraprotein is suspected additional immunofixation electrophoresis of serum and urine is recommended for detecting of a paraprotein and confirming its identity. Various renal diseases can be associated with a monoclonal gammopathy (Heher et  al., 2010). Examples include cast nephropathy, amyloidosis, monoclonal immunoglobulin deposition disease (MIDD), fibrillary and immunotactoid nephropathy, and membranoproliferative glomerulonephritis (MPGN) (see Chapters  60–64). Light microscopic evaluation of the kidney biopsy may already have raised the suspicion of a paraprotein related kidney disease. IF may be pathognomonic by revealing the mere presence of κ or λ light chains. However, IF studies may be misleading. In amyloidosis the paraproteins are modified, and specific epitopes may be lost, resulting in negative light chain staining. Alternatively, the paraprotein may interact with normal, polyclonal IgG (see ‘Cryoglobulins’). In patients with cast nephropathy, free light chains are always found in the urine. In fact, in the majority of patients the light chains are present in relatively large amounts (> 1 g) and account for > 70% of total urinary protein. Of note, urine dipsticks do not detect light chains, therefore in patients in whom myeloma is suspected, the urine must be screened by biochemical techniques. Immunofixation electrophoresis (IFE) lacks 100% accuracy for the diagnosis of paraprotein-related disorders. Using serum and urine IFE the sensitivity for detecting a monoclonal B cell disorder was 92% (Vermeersch et al., 2008b). Sensitivity is even lower in patients with amyloidosis or MIDD. In patients with AL amyloidosis sensitivity of serum IFE is 74–80% (Katzmann et al., 2009; Palladini et  al., 2009)  and of urine IFE 67–83% (Palladini et  al., 2009, Dispenzieri et al., 2009). In patients with MIDD, even when serum and urine immunofixation are combined a paraprotein may be undetected in a significant proportion of patients (4–22%). In a recent study in patients with glomerular patterns of injury (membranous nephropathy or MPGN) and characterized by the presence of monoclonal IgG on IF, a paraprotein was discovered in serum and/or urine by IFE in only 30% of patients (Guiard et al., 2011). The development of a nephelometric assay for FLC’s has allowed the detection and quantitation of small amounts of free κ and λ chains (Katzmann et al., 2002). However, absolute values of FLCs may be misleading. In healthy persons serum κ and λ FLCs increase with age. The κ/λ ratio was independent of age and a diagnostic reference interval of 0.26–1.65 was established (Katzmann et al., 2002). Similarly, absolute levels of FLCs are increased in patients with polyclonal hypergammaglobulinaemia, however the κ/λ ratio is always normal. Use of this assay improved diagnostic accuracy for paraprotein related disorders when compared to IFE:  in 67 patients with multiple myeloma, amyloidosis or MIDD, serum IFE electrophoresis was positive in only 72% of patients, whereas an abnormal κ/λ FLC ratio was found in 96%. Of note, differences were less impressive when urine IFE was used as a comparator, 89% of patients being positive (Katzmann et al., 2002). Similar results were reported by Bochtler et al. (2008), who studied 133 subsequent patients with AL amyloidosis. An abnormal FLC ratio was observed in 116 (87%) of patients and the FLC assay was

significantly better than serum IFE (abnormal in 69%), but not superior to urine IFE (abnormal in 86%) or the combination (abnormal in 92%). Although the FLC ratio may not improve diagnostic accuracy, serum free light measurements may aid in predicting prognosis in AL amyloidosis: median overall survival was 16 months in 644 patients with an abnormal κ/λ ratio, and 64 months in the remaining 86 patients with normal κ/λ ratio (Kumar et al., 2010). Quantitation of FLCs at baseline also improved risk prediction in patients with MGUS, and smouldering multiple myeloma (Rajkumar et al., 2005; Dispenzieri et al., 2009), and changes in FLCs during treatment predicted prognosis in AL amyloidosis (Lachmann et al., 2003). Since free light chains are cleared by glomerular filtration, serum concentrations of free light chains are dependent on glomerular filtration rate (GFR). Indeed, Hutchison et al reported that serum free light chains levels are inversely related to estimated GFR (eGFR) (Hutchison et  al., 2008). In patients with severe chronic kidney disease (CKD) (eGFR < 15 mL/min) serum levels are increased almost tenfold. There are slight differences in sieving characteristics between κ and λ light chains, sieving coefficient being lower for λ chains, that more often exist as dimers in the circulation. As a result, the κ/λ FLC ratio increases with decreasing eGFR. For patients with CKD new reference intervals have been defined: mean ratio 1.12 (95% confidence interval 0.37–3.1).

Cryoglobulins Cryoglobulins are Igs or combinations thereof that precipitate in the cold and re-dissolve upon rewarming to 37°C (Dammacco et  al., 2001). Cryoglobulins are potent activators of the classical complement pathway, which may explain the vasculitis that occurs in medium and small vessels. Renal involvement is seen in up to 25% of patients, and typically manifests as a MPGN (Alpers and Smith, 2008). Cryoglobulins are classified according to their component Igs. Type I cryoglobulins are composed of a single monoclonal Ig, type II cryoglobulins (also called mixed cryoglobulins) comprise a complex of a monoclonal Ig (typically IgM with antiglobulin, i.e. rheumatoid factor activity) and polyclonal IgG, and type III contain polyclonal IgG and/or IgM antiglobulin and polyclonal IgG (see Chapter 151). Renal involvement and the syndrome of ‘essential mixed cryoglobulinaemia’ is seen predominantly in subjects with type II cryoglobulins (Alpers and Smith, 2008). Type II cryoglobulins can be associated with a variety of underlying diseases such as auto-immune diseases, lymphoproliferative disorders, and hepatitis C virus (HCV) infection. In Mediterranean countries, up to 90% of patients with type II cryoglobulinaemia are HCV positive. However, reports differ from countries with a lower prevalence of HCV infections, with a prevalence of HCV ranging from 0% to 22% (Tervaert et al., 2007), (Weiner et al., 1998). In patients with non-HCV cryoglobulinaemia often no underlying cause is found and these 25–40% of patients are referred to as having true essential mixed cryoglobulinaemia. For the detection of cryoglobulins in the serum meticulous precautions at the time of blood collection must be taken, since improper sample handling may prevent detection of cryoglobulins (Allen et al., 2001; Vermeersch et al., 2008a; Motyckova and Murali, 2011). Blood must be drawn into a warm tube, and kept at 37°C during clotting. The serum is then placed in a refrigerator for at least 3 days (type I cryoglobulins usually precipitate within 24 hours, type II and type III cryoglobulins may need up to 1 week).

chapter 17 

The cryoprecipitate can be seen as a white precipitate in the bottom of the tube. After washing to remove passively trapped proteins, the precipitate is dissolved at 37°C and analysed by immunofixation techniques to establish its composition. There is no standard way of reporting and quantitation of cryoglobulins. Cryoglobulins are reported qualitatively (positive or negative) or quantitatively, using cryocrit, total protein content, or the immunoglobulin concentration of the cryoprecipitate. The cryocrit is the percentage of the centrifuged serum that is cryoglobulin. Since there is a lack of correlation between cryoglobulin concentration and symptoms, sensitive methods for the detection of cryoglobulins are advised. Assays now can detect very low concentrations of immunoglobulins in cryoprecipitate of healthy and disease controls. The 97.5th percentile of IgA, IgG, and IgM in diseased controls was proposed as the upper reference value and amounted 2 mg/L, 11 mg/L, and 34 mg/L respectively (Vermeersch et al., 2008a). The serum of patients with cryoglobulinaemia usually contains rheumatoid factor activity, resulting from the presence of an IgM with anti-IgG specificity. Serum complement C4 levels are usually very low, whereas C3 levels may be normal or low. If in a patient with cold insensitivity, purpura, livedo reticularis and/or recurrent skin ulcera no serum cryoglobulins are detected, a diagnosis of cryofibrinogenaemia must be considered (Blain et al., 2000). Cryofibrinogen consists of fibrinogen-fibrin complexes, and precipitates only from plasma (Vermeersch et al., 2008a).

immunological investigation of renal disease

on C3i. Measurements of complement breakdown products such as C3d may be more specific indicators of complement activation. However, local complement activation has a limited effect on systemic complement levels. Conversely, decreased levels of complement do not always reflect immunological injury. Temporary reductions of complement C3 and/or C4 can be found in various

Antibody on microbe

C1r

Classical pathway

MASP

C1s

Lectin pathway

C4 C4a C4b

Alternative pathway

C2 C4b2b P

C3 C2a

H

Microbial cell walls

I

Activation

C3bBb

C3bH B Ba

C3a C5a

D C5

Attaches to C3b receptors on phagocytes

Opsonization

Deactivation

C3b

Complement The complement system is a group of tightly controlled plasma proteins, which after activation evoke potent biological effects. The activation pathways of the complement system, its most important components and regulatory proteins, and the biological effects are shown in Fig. 17.1. However, the crucial regulation of complement activation occurs at the membrane level, either by the constitutive membrane bound membrane co-factor protein (MCP; CD46) or by factor H, which binds to membrane glycoproteins. The complement system can be activated in three different ways, the classical pathway, the alternative pathway, and the MBL (mannose binding lectin) pathway. All three pathways lead to the generation of an enzymatically active C3-convertase, which cleaves the central component C3 into C3a and C3b. C3b converts the C3-convertase to a C5 convertase, which cleaves C5 to C5a and C5b, the initiator of the formation of the C5b–C9 complex, the so-called membrane attack complex. The alternative pathway of complement is constantly active, partly mediated by the activating factor B. To prevent overactivation of the complement system, regulatory (inhibitory) proteins are present such as factors H, I, and MCP. Levels of individual components of the complement system can be measured by immunochemical methods. C3, C4, and factor B are the most frequently measured, and allow differentiation of alternative (low C3, normal C4) and classical (low C3 and low C4) pathway activation. Complement activation is not always reflected by decreased levels of C3 or C4. Sometimes the increased production of complement components, as a result of an acute phase response, counterbalances the increased breakdown. Alternatively, activation of C3 may go unrecognized since antibodies are used that recognize an epitope that is present on intact C3 as well as

MBL

C1q

C5b C9 C9 C9 C9 C9 C9 C9

C6

PMN chemotaxis Mast cell degranulation

C7 C8

Vascular permeability

Membrane attack complex

Phagocytosis

Lysis

Inflammation

Fig. 17.1  Overview of the complement system. C4 is activated either by the classical pathway or by the lectin pathway. The classical pathway is activated by binding of C1q to antigen–antibody complexes. This activates C1r and C1s which cleave C4. The lectin pathway is activated if the mannan binding lectin (MBL) binds to mannose groups on bacteria. This activates the serine proteases MASP 1 and 2 (MBL associated serine protease). MBL and MASP are homologous to C1q, C1r, and C1s. Next, C2 can bind to C4b and then becomes susceptible for cleavage by C1s. After cleavage the classical C3 convertase (C4b2b) is formed, which cleaves C3 into C3b and C3a. The formation of C3b can also result from alternative pathway activation. Spontaneous C3 activation occurs spontaneously at a low level. If this activated C3b binds to factor B, the complex becomes susceptible for cleavage by factor D. This results in the formation of the alternative pathway C3 convertase C3bBb. However, factor H has a higher affinity for C3b than factor B. If factor H binds to C3b the complex is cleaved by factor I resulting in inactivated C3b. Certain microbial surfaces favour the association between C3b and factor B and therefore activate the alternative pathway. Cell surfaces promote the binding of factor H to C3b thereby inhibiting alternative pathway activation. Generation of C3b is a central feature of complement activation, because of the biological functions associated with C3b formation namely opsonization/phagocytosis and pro-inflammatory responses via C3a and C5a. In addition it forms the initial event in the formation of the so-called membrane attack complex C5b–C9 which drills holes in bacteria and cells. Adapted from Playfair JHL, Lydyard PM. Medical Immunology for Students. Churchill and Livingstone 1995; page 8.

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conditions such as atheroembolic disease, sepsis, pancreatitis, severe burns, following the use of contrast agents or inulin, cuprophan haemodialysis, cardiopulmonary bypass, haemolytic crisis in malaria, and porphyria (Hebert et al., 1991). The CH50 activity assay is a functional assay that measures the overall activity of the complement system after activation of the classical pathway. Absent or very low CH50 activity indicates a congenital deficiency of one of the complement components of the classical pathway such as C1q, C2, or C4. We now know that dysregulation of the complement system is often not detected by the standard immunochemical quantitative measurement of the involved proteins. Most abnormalities are caused by genetic mutations that alter the function of the protein, without affecting its production or expression. Function of complement regulatory proteins may also be affected by the presence of specific antibodies, the best example being antibodies against factor H. The complement system is frequently involved in renal diseases. Although for many decades the role of the classical pathway has been emphasized, abnormalities in alternative pathway activation have recently attracted much attention as cause of renal diseases such as the haemolytic uraemic syndrome (HUS) and dense deposit disease (DDD) (Taylor et al., 2010; Pickering and Cook, 2011). In Table 17.2 we provide an overview of complement abnormalities in various renal diseases. Congenital deficiencies of complement components deserve special consideration. There are known associations between complement deficiencies and the development of various renal diseases (Table 17.1). Deficiencies of C1q, C2, and C4 are frequently associated with systemic lupus erythematosus (SLE). C3 nephritic factor is an auto-antibody that is associated with DDD. This auto-antibody prevents the deactivation of C3b by factor H, either by binding to C3b or to factor H. Some patients with DDD and C3 nephritic factor have a typical gaunt appearance with partial lipodystrophy, the absence of fat tissue in the face and upper part of the body. The pathogenesis has been recently clarified: the adipocytes in the upper part of the body produce a protein called adipsin which is identical to factor D. In the presence of C3 nephritic factor complement will be activated and mediate lysis of the adipocytes at these sites.

Antinuclear antibodies ANAs are directed against nuclear constituents, either nucleic acids or nucleoproteins. ANA can be detected in an indirect IF test by applying serum to fixed cell lines. Most commonly used are Hep-2 cells. A positive ANA on indirect IF can be found in up to 10% of aged healthy persons. Various defined specificities of autoantibodies can cause a positive ANA (Table 17.1). The nuclear staining pattern can already provide an indication for the specificity of the ANA (see Fig. 17.2). A homogenous or peripheral staining pattern is mainly caused by antibodies against the nucleosome or its components, double-stranded DNA (dsDNA) or histones. A speckled pattern is caused by antibodies against nucleoproteins like Sm, RNP, SS-A (Ro), or SS-B (La). A centromere or nucleolar staining is suggestive for various scleroderma-associated autoantibodies. The antibody specificities can be more precisely determined by additional testing. Specificity for dsDNA can be determined qualitatively by an indirect IF assay on Crithidia luciliae, a flagellate which contains a kinetoplast with pure dsDNA. For quantitative testing the Farr assay is used, a modified RIA that particularly detects high avidity antibodies. Anti-DNA ELISA systems are available but their specificity is lower because (non-pathogenic) low avidity antibodies are also detected. In recent years it has become apparent that antinucleosome antibodies have a higher sensitivity (especially in lupus nephritis) than anti-dsDNA and an equal specificity (van der Vlag and Berden, 2011). In lupus patients negative for anti-dsDNA 54% were positive for antinucleosome antibodies. In one study it was found that all anti-dsDNA negative patients with lupus nephritis were antinucleosome positive. There are now ELISAs available that test both anti-DNA and antinucleosome antibodies (Biesen et al., 2011). The specificity for other defined nucleoproteins, as suggested by a speckled ANA pattern, can be documented by a number of techniques including an immunodiffusion technique using purified extracted nuclear antigens (ENAs), an immunoblotting technique using either an extract of nuclear and/or cytoplasmic antigens or recombinant autoantigens, or an ELISA using these autoantigens. Well-known specificities include anti-histone, anti-Sm, anti-RNP, anti-SS-A, anti-SS-B, and anti-topo-isomerase I.

Table 17.2  Complement levels in various renal diseases C4

C3

Factor B

PSGN

N(–↓)

↓↓

N?

Only in acute phase

Lupus nephritis

↓↓

↓↓

N

Proportionally decreased

MPGN type I MPGN type II (DDD)

N-↓ N

↓ ↓↓

N ↓

Due to presence C3NeF

GN associated with chronic infections

↓

↓

N

Cryoglobulinaemic GN

↓–↓↓

N or ↓

N

ANCA associated GN

N

N

N

Anti-GBM disease

N

N

N

Haemolytic uraemic syndrome

N

N or ↓

N

↓ = decrease; ↑ = increase; C3NeF = C3 nephritic factor; DDD = dense deposit disease; GN=glomerulonephritis, MPGN = membranoproliferatieve glomerulonephritis; N = normal; PSGN = poststreptococcal glomerulonephritis.

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(A)

(B)

(C)

(D)

immunological investigation of renal disease

Fig. 17.2  The four common patterns of antinuclear staining. Homogenous (A), peripheral (B), speckled (C) and nucleolar (D). Reproduced with permission from Dieppe PA, Bacon PA, Bamji AN, Watt I. Slide Atlas of Rheumatology, vol. 9 SLE. Gower, London, 1984; page 9.6.

Anti-dsDNA (anti-nucleosome) and anti-Sm antibodies are almost pathognomonic for SLE. Both specificities are included in the American College of Rheumatology criteria for SLE. Various other ANAs are also frequently found in patients with SLE and correlate with specific disease associations (see Chapter 163).

cryoglobulinaemia, and MPGN (60–80%). Therefore, their specificity for a particular disease is low. Moreover, anti-C1q antibodies are also present in healthy controls (5%), the prevalence increasing with age (> 70 years, 18% positive).

Anti-C1q antibodies

Antineutrophil cytoplasmic antibodies

The popularity of the use of assays for measurement of circulating immune complexes in the past using C1q as a substrate, revealed the presence of autoantibodies to C1q in many disease conditions (Kallenberg, 2008). These antibodies can be measured by using immobilized C1q either in ELISA or a RIA. To prevent binding of immune complexes or dsDNA to C1q these tests are performed at an increased ionic strength, mostly 0.3 M NaCl. These anti-C1q antibodies are frequently found in patients with proliferative lupus nephritis (30–80%). In many lupus patients anti-C1q antibodies are related to disease activity with higher prevalences and higher titres in active renal disease compared to inactive LN or active non-renal lupus (Kallenberg, 2008). In two studies, anti-C1q levels rose significantly prior to relapses of renal disease (Coremans et al., 1995; Moroni et al., 2001). In the latter study, a sensitivity of 87% and a specificity of 92% was found. However, this was also found in patients with hypocomplementemic urticarial vasculitis (100%) mixed connective tissue disease, polyarteritis nodosa (± 30%),

The discovery of ANCAs has been a major breakthrough when the differential diagnosis includes vasculitis and rapidly progressive glomerulonephritis (RPGN). ANCAs are directed against proteins that are predominantly present in the azurophilic granules of neutrophils. The screening assay for ANCA is indirect IF on ethanol fixed neutrophils. Under these conditions three different patterns can be seen:  cytoplasmic ANCA (c-ANCA), perinuclear ANCA (p-ANCA), or atypical ANCA (x-ANCA) (Fig. 17.3) The difference in staining pattern between c-ANCA and p-ANCA is due to a fixation artefact. Ethanol fixation disrupts the lysosomal membrane and as a result the lysosomal proteins will leak into the cytoplasm. Cationic proteins (targeted by p-ANCA) will be attracted to the negatively charged nuclear membrane giving rise to the perinuclear staining pattern, whereas anionic or neutral proteins will be more evenly distributed in the cytoplasm (c-ANCA). On formalin-fixed neutrophils there are no differences, and only a c-ANCA staining pattern is seen.

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(A)

(B)

(C)

Formalin fixation Strong cationic proteins (e.g. MPO) C-ANCA

Weakly cationic or neutral proteins (e.g. PR3)

Ethanol fixation

P-ANCA

C-ANCA Antibodies to strong cations

Antibodies to neutral proteins or weak cations (e.g. PR3)

Fig. 17.3  Antinuclear cytoplasmic antibodies (ANCA). (A and B) Indirect immunofluorescence techniques using ethanol-fixed neutrophils demonstrate perinuclear (p) (A) and cytoplasmic (c) (B) staining patterns. (C) The use of crosslinking fixatives (e.g. formalin) during the preparation of the neutrophil substrate leads to alterations in granule membranes, allowing positively charged constituents to migrate to the negatively charged nuclear membrane. Because this does not occur with formalin fixation, use of both types of fixation allows the distinction of a true p-ANCA from an antinuclear antibody (ANA). With formalin-fixed neutrophils, antibodies that would cause p-ANCA patterns (on ethanol-fixed cells) will display diffuse granular cytoplasmic staining, whereas nuclear staining would indicate the presence of an ANA. MPO = myeloperoxidase; PR3 = proteinase 3.

P-ANCA must be differentiated from ANAs that also give a perinuclear staining pattern. The effects of fixation can provide a clue. Whereas p-ANCA staining will change to a c-ANCA pattern on formalin fixed neutrophils, the perinuclear staining of ANAs will remain unchanged. Furthermore, ANA staining can be observed when using other cells or tissues, whereas p-ANCA antibodies are neutrophil specific. Most c-ANCA are directed against a 29kD protein called proteinase-3 (PR3). P-ANCA are most frequently directed against myeloperoxidase (MPO), although there are other antigenic targets such as elastase, lactoferrin, lysosyme, cathepsin G, and bactericidal/permeability-increasing protein (BPI). The specificities of x-ANCA are not fully elucidated yet although binding has been observed both to the neutrophil protein tubulin-B5 and the bacterial cell division protein (FtsZ). This cross-reaction is perhaps implicated in the pathogenesis of inflammatory bowel disease (Terjung and Spengler, 2009). This is in line with the observation in a French study in which 71.8% of patients with ulcerative

colitis and 11% of patients with Crohn’s disease were x-ANCA positive, while healthy blood donors showed a prevalence of 0% (Desplat-Jego et al., 2007). The antigenic specificity of ANCA should be tested by the widely available ELISA assays for PR3 and MPO. These commercial assays only detect IgG class antibodies. Although there is concordance between c-ANCA staining and anti-PR3 positivity in ELISA and between p-ANCA staining and anti-MPO positivity, the results are not always congruent and depend on the assay used (Wang et  al., 1997). Of c-ANCA positive sera, 44–84% were positive in a PR3 ELISA, whereas p-ANCA positivity was confirmed by anti-MPO ELISA in 25–75% of cases (Wang et  al., 1997). In the latter study, only 35% of p-ANCA positive sera proved MPO-ANCA positive in capture ELISA, immunoblot and inhibition assays, indicating the frequent existence of other specificities. A  European multicenter analysis revealed that the IF test is more sensitive but the ELISA is more specific (Hagen et al., 1998). However, one should realize that sensitivity and specificity vary between laboratories. Up to 5% of patients with vasculitis are negative in IIF but positive by a MPO or PR3 specific ELISA (Savige et  al., 2000). Granulomatosis with polyangiitis (formerly Wegener’s granulomatosis) is predominantly associated with PR3-ANCA, while microscopic polyangiitis is primarily associated with MPO-ANCA. Of note 20% of patients have the alternative ANCA and 10% are negative. Recently, a new autoantibody specificity was described in ANCA-associated vasculitis. The antibodies were directed against lysosome-associated membrane protein-2 (LAMP-2) (Kain et al., 2008). Intriguingly, LAMP-2 shows partly homology with a bacterial protein FimH present on Gram-negative bacteria. Immunization with FimH in rats led to induction of anti-LAMP-2 and a pauci-immune GN. However, whether anti-LAMP-2 is prevalent in ANCA-positive patients remains to be proven since the results are very conflicting (Kain et al., 2012; Roth et al., 2012). Therefore, at present screening for anti-LAMP-2 is not recommended in patients with suspected ANCA vasculitis.

Anti-GBM antibodies Anti-GBM disease is the prototypic example of antibody mediated renal disease. In anti-GBM disease the glomerulus is severely damaged with a characteristic picture of severe extracapillary proliferation with capillary wall necrosis. Patients present with the clinical picture of a RPGN. In some patients, especially in smokers, the lungs may be involved too. Classical Goodpasture’s syndrome is a pulmonary renal syndrome of RPGN with haemoptysis. A renal biopsy in patients with anti-GBM disease discloses a typical picture on IF, with linear IF of Ig along the glomerular capillary wall. In most cases the causative antibody is of the IgG class, however, patients with IgA and IgM class anti-GBM antibodies have also been described. The antibodies in anti-GBM glomerulonephritis are directed against a conformational epitope on the so called non-collagenous (NC1) domain of the collagen IV α3 chain (Hellmark et al., 1997). This epitope becomes accessible if the dimeric form is converted to the monomeric form. Based on this knowledge, sensitive and specific ELISA assays have been developed using recombinant protein for the quantitative measurement of anti-GBM antibodies. These antibodies can be detected in high titres in almost every patient

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with anti-GBM disease. The sensitivity and specificity of the assay is very high, although low titres can be observed in other unrelated conditions. The positive predictive value (PPV) and negative predictive value (NPV) are therefore very high and are ≥ 95% (Sinico et al., 2006). Of note, the ELISA regularly only detects IgG antibodies, thus the rare patient with IgA or IgM anti-GBM antibodies may remain formally undiagnosed as suffering from anti-GBM disease. In approximately 30% of patients with anti-GBM antibodies, ANCA antibodies are also found (two-thirds with anti-MPO specificity). Although the co-occurrence of ANCA and anti-GBM antibodies is mostly associated with a more favourable course of the disease and in these patients treatment is more efficacious, this is not a uniform finding (Rutgers et al., 2005). The epitope of the anti-GBM antibodies in this subset of patients is not different (Hellmark et  al., 1997). ANCA-activated neutrophils unmasking the cryptic epitope of the NC1-domain is an intriguing, but not proven hypothesis. Recently, it was shown that ANCAs precede the formation of anti-GBM antibodies, lending further support for the pathogenic significance of ANCA in these patients (Olson et al., 2011) Anti-GBM antibodies can also be produced in patients with Alport’s disease after renal transplantation (Brainwood et al., 1998; Kalluri et al., 2000). Alport’s disease is caused by mutations in genes encoding the α3, α4, or α5 chains of collagen IV. These patients do not express these collagen chains in their native forms in their kidneys. After renal transplantation, some patients may develop an aggressive form of anti-GBM disease, causing renal failure. Recent studies have indicated that anti-GBM antibodies can be detected in almost all patients, linear deposits of IgG along the glomerular capillary wall are seen in 20–50% of patients, whereas only a minority develop manifest anti-GBM disease. Several studies have looked for the epitopes to which these antibodies are directed. Although the results are equivocal, many patients have antibodies that are directed against the NC1 domain of collagen α5, especially in patients with X-linked Alport syndrome (Borza, 2007). These antibodies may not be detected by commercial anti-GBM ELISA assays, that use the specific NC1α3 antigen as substrate.

Antipodocytic antibodies It has long been suggested that idiopathic membranous nephropathy (iMN) can be caused by the binding of a circulating antibody to an antigen that is present on podocytes, but the responsible antigen in human membranous nephropathy was elusive. In 2009, the M-type phospholipase A2 receptor (PLA2R) was identified as an important antigenic target (Beck et al., 2009). The authors showed that antibodies against PLA2R, primarily of the IgG4 subclass, were present in approximately 70% of patients with iMN. This study finally proved that iMN is an auto-immune disease. A Western blot technique had been used for the detection of antibodies against PLA2R, but an indirect IF technique is now commercially available. For this assay slides containing biochips of HEK 293 cells transfected with cDNA coding for PLA2R, or non-transfected cells as control are used (Hoxha et al., 2011). A commercial ELISA assay is now available (Dahnrich et al., 2013). Recent studies suggest that antibodies against PLA2R are specific for membranous nephropathy, and particularly the idiopathic form. However, the accuracy of this assay to diagnose iMN and to exclude secondary causes of MN will require prospective studies.

immunological investigation of renal disease

A quantitative assay may be needed for prediction of prognosis and to guide treatment (Hofstra et al., 2012). The role of antibodies against other podocytic antigens such as superoxide dismutase, aldosereductase, and alpha-enolase needs further evaluation.

Antiphospholipid antibodies Antiphospholipid (aPL) antibodies are associated with the antiphospholipid syndrome (APS), which is characterized by (recurrent) arterial and/or venous thromboses and recurrent miscarriages (Levine et  al., 2002). The kidney is often involved, with vascular lesions and variable thrombotic microangiopathy (TMA) (Nochy et al., 1999). APS can be primary (without systemic auto-immunity) or secondary in patients with auto-immune diseases such as SLE (Daugas et al., 2002). The first recognized aPL antibodies were the lupus anticoagulant (LAC) and the anticardiolipin (aCL) antibodies. More recently antibodies to β2-glycoprotein-1 have been identified. The LAC is defined as a serum factor that prolongs phospholipid-dependent coagulation tests in platelet-poor plasma. Examples are the activated partial thromboplastin time (APTT), the kaolin clotting time, or diluted Russell’s viper venom clotting time (DRVVT). Tests are considered positive if the coagulation defect cannot be corrected by addition of normal plasma, but becomes normal after addition of excess anionic phospholipids or freeze-thawed thrombocytes. There are a number of different methods to detect LAC activity (Jacobsen et al., 2001). By only using the standard APTT test, with a cut of value of 67 seconds, 47% of lupus anticoagulant positive samples will be missed (Jacobsen et al., 2001). DRVVT is considered the most sensitive test so the presence of LAC must be ascertained by using at least two independent tests but acknowledging that the accuracy and reproducibility of the test procedures is low. ACL antibodies are directed against cardiolipin, and can be detected in ELISA. Although some aCL antibodies have LAC activity, the results do not always overlap. Thus when searching for aPL antibodies both tests are necessary. Recently, it has been demonstrated that aPL antibodies may be directed at various plasma proteins that have affinity for anionic phospholipids. In fact, aCL antibodies recognize a conformational epitope formed between CL and β2-glycoprotein-1 (β2GP-1). Therefore, β2GPI is an essential co-factor in these assays. Because of this, antibodies to β2-glycoprotein-1 can also be tested directly in ELISA. This is especially necessary in patients in whom there is a high suspicion for APS but negative aCL or LAC tests. For a diagnosis of APS at least one clinical manifestation should have occurred (thrombosis or pregnancy morbidity) together with positive tests 12 weeks apart for one of the aPL assays (aCL, LAC or β2GP-1). The rate of thrombosis increases significantly when more aPL tests are positive: single positivity, 27.6%; double, 38.8; or triple, 66.7% (Lee et al., 2003). Additional specificities include proteins such as prothrombin, protein C, protein S, and annexin V. Of note, these proteins are only recognized in the assays if bound to anionic surfaces. In ELISA antigens must therefore be coated specifically on γ-irradiated polystyrene plates. Although the association between the presence of aPL antibodies and thrombosis is well recognized, the pathogenicity of the antibodies is unclear because aCL antibodies may be detectable in

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2–4% of healthy controls. The incidence is higher (15%) in patients with rheumatoid arthritis or SLE (Merkel et al., 1996). Other specificities are also found with high frequency in patients with SLE. A 21–56% prevalence of antibodies to ß2GP-1, prothrombin, protein C, protein S, and annexin V has been reported in patients with SLE (Nojima et al., 2001). Patients with positive lupus anticoagulant had significantly more frequent antibodies against prothrombin and ß2GP-1, with no difference in the other antibody specificities. The association between LAC, aCL, and aβ2GP-1 antibodies and the APS is well established (Levine et al., 2002). In general, LAC activity is more specific whereas aCL and aβ2GP-1 antibodies show higher sensitivity. However, it was found that the presence of one of the latter antibodies has a poor association with thrombosis (Galli et al., 2003). Understanding of the APS and the role of the aPL antibodies in various clinical manifestations should improve with the development of more specific and better standardized assays. An association was observed between antibodies against ß2GP-1 and prothrombin and arterial thrombosis; between antibodies against protein S and venous thrombosis; and between anti-annexin V and fetal loss (Nojima et al., 2001). These results need confirmation in prospective studies. Patients with an established APS need life-long full oral anticoagulation. For the monitoring of the intensity of oral anticoagulation the international normalized ratio (INR) is used aiming at an INR between 2.0 and 3.0 (Crowther et al., 2003). It should be realized that monitoring of the INR may be difficult in patients with LAC since some available tests may be influenced. In the presence of LAC, values of INR may be falsely elevated, and the level of anticoagulation may be overestimated (Moll and Ortel, 1997).

Antimicrobial responses (See Chapter 164.) The kidney can be damaged in the course of many infectious diseases (see Chapters 77–9). Poststreptococcal glomerulonephritis (PSGN), hepatitis B, and hepatitis C are three important examples.

Antibodies against streptococcal antigens The epidemiology of PSGN has changed in the past three decades (Rodriguez-Iturbe and Musser, 2008). In central Europe, the incidence is low, the elderly being more affected than children. The disease most often occurs in epidemics, and a high incidence is reported in rural communities with low socioeconomic status. PSGN was once regarded the best example of an immune-complex mediated glomerular disease, caused by entrapment in the glomerular capillary wall of circulating immune complexes that consisted of streptococcal antigens and antistreptococcal antibodies. Alternative hypotheses included in situ formation of immune complexes by binding of antibodies against bacterial antigens planted in the GBM, or by binding of antibodies to intrinsic glomerular antigens that mimic bacterial epitopes. The focus has shifted recently to two newly defined, potential nephritogenic antigens, that is, the nephritis-associated plasmin receptor (NAPlr) and the streptococcal pyrogenic exotoxin B (SPEB) (Rodriguez-Iturbe and Musser, 2008). Both NAPlr and SPEB have been detected in glomerular deposits and antibodies against these antigens were present in a high percentage of patients with PSGN, although results differed markedly between populations, suggesting that more than one antigen may be responsible for development of PSGN. The diagnosis of

PSGN requires proof of a prior streptococcal infection. Most studies evaluate the production of antibodies directed against streptococcal antigens. Although antibodies against NAPlr and SPEB may be most relevant, thus far no large study has evaluated their accuracy in diagnosing PSGN. Antibodies against streptolysin O (ASO) are not very accurate, since many patients, in particular those with a streptococcal skin infection, do not produce ASO antibodies. The detection of antibodies directed to other streptococcal antigens, such as deoxyribonuclease B (DNAse B), streptokinase, hyaluronidase, zymogen, GAPDH, and NAD+ glycolase, may be more sensitive and specific for the diagnosis of post-streptococcal infection. In children with acute PSGN the sensitivity of ASO, anti-DNAse and antizymogen titres has been studied (Parra et al., 1998). When compared to controls ROC curves showed superiority of anti-zymogen > anti-DNAse B > ASO. Sensitivity and specificity was 42% and 84% for ASO, 73% and 89% for anti-DNAse B, and 87% and 85% for anti-zymogen. Of note, increased titres of these antibodies were also observed in patients with upper respiratory tract infections or impetigo who did not experience a glomerulonephritis. Thus, the above-mentioned specificity of the assays does not hold if patients with infections, without evidence of renal involvement, are included.

Hepatitis C Hepatitis C is held responsible for various renal diseases such as MPGN type I, membranous nephropathy, and cryoglobulinaemic glomerulonephritis (Lauer and Walker, 2001; Charles and Dustin, 2009). The detection of hepatitis C is based on the detection of antibodies against hepatitis C antigens. An enzyme immunoassay (EIA) is used for screening. This test is very sensitive when used in low-risk population (99%). However, false-positive results are common, thus positive results should always be confirmed by a recombinant immunoblot assay (Chandler, 2000). False-negative results occur particularly in patients with end-stage kidney disease and patients using immunosuppression. Reported sensitivity of an EIA was only 53% and 72% in two studies in dialysis units (Schroter et al., 1997; Kalantar-Zadeh et al., 2005). In patients with high suspicion for hepatitis C, HCV RNA can be detected by rtPCR, a very sensitive method that allows detection of virus as early as 2 weeks after infection. Since viral RNA is unstable, blood samples must be processed within 3 hours. In patients with cryoglobulinaemia, HCV viral RNA may be predominantly contained in the cryoprecipitate. Special care must be taken to draw the blood at 37oC and that the rtPCR is performed on the cryoprecipitate (Charles and Dustin, 2009).

Hepatitis B Various serological markers can be used to evaluate a hepatitis B infection, for example, HBsAg, HBe antigen, and anti-HBs, anti-HBc, and anti-HBe antibodies. The test results provide an indication on the time sequence and the infectivity of the blood (Edey et al., 2010). The presence of HBsAg and HBe suggests infectivity and anti HBs indicates immunity. However, HBsAg and anti-HBs antibody negative patients may be infectious, particularly if organs are used for transplantation. There is a risk of transmission particularly if an organ is transplanted into a non-immune recipient, the risk being higher with liver than with kidney transplantation (Delmonico and Snydman, 1998). It has been suggested that immunosuppressed patients (including patients on haemodialysis) may

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not develop anti-HBs antibodies after exposure to the viral antigen. If suspicion is high, HBV DNA can be detected by sensitive PCR techniques. There is continuous uncertainty about the presence and prevalence of occult HBV infection in HBsAg negative dialysis patients, with estimates ranging from 0% to 26.6% (Fabrizi et al., 2005; Di Stefano et al., 2009).

Immunological studies in patients with specific renal syndromes: guidelines Few studies have specifically assessed the value of testing specific immunological markers in establishing a diagnosis or monitoring disease activity in patients with renal diseases. We will briefly discuss the data that are most relevant for the important clinical renal syndromes and several specific disease entities.

Nephrotic syndrome Two studies have argued against the routine testing of patients with proteinuria or a nephrotic syndrome (Howard et  al., 1990; Chew et  al., 1999). Measurement of ANA, rheumatoid factor, C3, C4, CH50, HbsAg, Venereal Disease Research Laboratory (VDRL) tests, cryoglobulins, and serum and urine electrophoresis did not improve diagnostic accuracy. Therefore, in almost all cases a renal biopsy is needed to establish a diagnosis (Richards et al., 1994). Based on the renal biopsy results (and sometimes on clinical grounds), additional serological tests can be performed that can strengthen the diagnosis and sometimes are helpful during follow-up. If validated, measurement of anti-PLA2R may become the first accurate diagnostic test for a specific cause of the nephrotic syndrome.

Rapidly progressive glomerulonephritis/acute nephritic syndrome (See Chapters 70–4.) In patients with an RPGN glomerular damage occurs often very rapidly and is irreversible if treatment is postponed. Therefore, a rapid diagnosis and early treatment is urgently needed. Serological markers can be very helpful in establishing a diagnosis in these patients. It is important to stress that an examination of the urinary sediment for the presence of dysmorphic erythrocytes and/ or erythrocyte casts remains the most important diagnostic tool to differentiate between RPGN and other causes of acute kidney injury. The presence of high titres of anti-GBM antibodies in patients with a RPGN is virtually pathognomonic for anti-GBM disease. By

Table 17.3  Sensitivity and specificity of ANCA using combined IIF and ELISA in systemic vasculitis Sensitivity

Specificity

Granulomatosis with polyangiitis (formerly Wegener’s disease; N = 97)

73%

98%

Microscopic polyangiitis (N = 44)

67%

98%

Idiopathic RPGN (N = 12)

82%

98%

Adapted from Hagen et al. (1998). Numbers in parentheses represent number of patients in the study cohort. Specificity data are based on 184 disease controls.

immunological investigation of renal disease

using the widely available, specific, and sensitive ELISA, a diagnosis can be made within a day. Although most centres still prefer to perform a renal biopsy, such an approach can be challenged as it may delay institution of treatment with plasma exchange. In patients with anti-GBM antibodies, a search for the presence of ANCA is also warranted. Clinicians should also be aware of the possibility that rare cases of anti-GBM disease are caused by non-IgG antibodies, that will not be detected in the standard assays. Regular assessment of the anti-GBM antibody titres during follow-up may guide treatment. In general, antibodies will disappear within 12–16 weeks after start of treatment, and at that time point immunosuppressive treatment can be rapidly tapered and stopped. Renal transplantation in a patient with anti-GBM disease should be postponed until no anti-GBM antibodies have been detected for at least 3 months. The finding of ANCA has a high predictive value in patients with RPGN. In fact, some authors have argued that in patients with extrarenal symptoms that fit with a multisystemic vasculitis, and who present with acute kidney injury, the detection of ANCA is sufficient for the diagnosis and a renal biopsy may be omitted. Data on the sensitivity and the specificity of ANCA for the diagnosis of systemic or renal limited vasculitis are given in Table 17.3 (Hagen et al., 1998). The diagnostic accuracy is best if ANCA are analysed by both IIF and ELISA techniques. In a meta-analysis, combined testing for c-ANCA/αPR3 and p-ANCA αMPO showed a sensitivity of 85.5% and a specificity of 98.6% for the diagnosis of small vessel vasculitis (Choi et al., 2001). One should realize that the high positive and negative predictive values in this and other studies only hold for patients with a (rapidly) deteriorating renal function, in whom the prevalence of systemic vasculitis is high. Predictability is much lower in patients with moderate renal failure (Falk et al., 2000) (Table 17.4). In such patients a renal biopsy is always needed to establish a diagnosis. Recently, it was demonstrated that during active phases of ANCA-associated vasculitis plasma levels of C3a, C5a, C5b-9, and Bb were elevated while properdin levels were diminished. Activation of the classical pathway was not observed since C4d levels were not different between active and non-active disease. Factor Bb levels were significantly associated with the amount of cellular crescents and clinical disease activity (Gou et al., 2013). These data indicate activation of the alternative pathway (Kallenberg and Heeringa, 2013) It is still unknown whether evaluation of ANCA activity is of value in the follow-up of patients. In 50–60% of patients titres fluctuate parallel to disease activity. In 83% of patients a favourable response to immunosuppressive treatment is accompanied by a negative ANCA test after a median of 12 weeks (Jayne et al., 1995). The predictive value of a rise in ANCA titre (if measured monthly) for a clinical relapse has a sensitivity of 57% and a specificity of 45%. The specificity increases to 72% if transient increases after conversion from cyclophosphamide to azathioprine and rises at the moment of clinical relapse are excluded. These results were confirmed in a recent prospective study. The predictive value for a relapse of a rise in c-ANCA titre in IF was 57% and 71% if assessed in a PR3-ELISA (Boomsma et  al., 2000). However, one should realize that disease quiescence is possible with persistently positive ANCA titres and that relapses can occur while the ANCA test remains negative. Serial measurements of ANCA is therefore of limited use (Tomasson et al., 2012).

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Table 17.4  Estimated positive (PPV) and negative (NPV) predictive values of ANCA for pauci-immune crescentic glomerulonephritis Clinical picture

PPV

NPV

RPGN

98%

80%

Nephritic syndrome and Serum creatinine > 270 μmol/L (A)

92%

80%

Serum creatinine 135–270 μmol/L (B)

77%

98%

Serum creatinine < 135 μmol/L (C)

47%

99%

Adapted from Falk et al. (2000). Data are for adult patients with observed prevalences of pauci-immune crescentic glomerulonephritis 47% for RPGN, 21% for A, 7% for B, and 2% for C.

In patients with a RPGN that are anti-GBM and ANCA negative, additional serological tests may be done based on clinical symptoms or biopsy results (Fig. 17.4). In these patients abnormalities in serum complement levels, the presence of auto-antibodies, or the detection of cryoglobulins or paraproteins are not enough sensitive or specific to establish a diagnosis, however, a positive finding may be helpful in confirming a diagnosis or be used to guide treatment. Abnormalities in complement levels in renal diseases are given in Table 17.2. For most diagnoses there is no evidence that serial measurements of complement can be used to monitor disease activity.

Systemic lupus erythematosus Laboratory tests are pivotal in establishing the diagnosis and may be helpful in monitoring the renal disease activity in patients with SLE. The ANA test is a valuable screening test with high sensitivity but low specificity. If positive, additional tests must be performed to establish antibody specificities. The presence of anti-dsDNA and anti-Sm antibodies establishes a diagnosis of SLE with 85% sensitivity and 93% specificity. Anti-nucleosome antibodies (van der Vlag and Berden, 2011) and antibodies to C1q (Kallenberg, 2008) may be better predictors of renal involvement in SLE; however, prospective data are limited. Complement levels are not helpful in the diagnosis of SLE, but persistently low CH50

values raise the possibility of a deficiency of one of the early complement factors. Antinuclear antibody titres are valuable in monitoring SLE nephritis disease activity during follow-up. If anti-dsDNA is measured monthly by the Farr assay, onset or flares of lupus nephritis will be accompanied by increases of anti-dsDNA titres in > 90% of patients (ter Borg et al., 1990). Measuring of anti-C1q antibodies might be more accurate in predicting renal relapses (Moroni et al., 2001). The likelihood of renal involvement in SLE patients who are negative for anti-C1q or anti-dsDNA antibodies is very low (specificity between 92% and 100%) (Trendelenburg et al., 1999; Moroni et al., 2001). Unfortunately, the anti-C1q assay is not available for routine use. Whether measurement of complement is valuable for monitoring disease activity is still unclear. Monitoring of C4 is not useful, although a low C4 value (< 110 mg/L) at the moment of renal remission, was identified as a risk marker for a flare (Illei et  al., 2002). Measurement of C3 appears more useful, with sensitivities ranging from 20% to 95% and specificities between 74% and 94% (Esdaile et al., 1996). However, in many patients with renal disease activity normal C3 levels are found, whereas decreased C3 levels may be present in patients without renal disease activity. In Table 17.5 the experience in a 6-year prospective study in 228 patients is given (Moroni et al., 2009). It underscores the low PPV of serology for confirming renal lupus flares.

Thrombotic microangiopathy TMA is a descriptive diagnosis and associated with various clinical syndromes (see Chapter 174). Dysregulation of the complement system is an important cause of TMA, a condition also defined as atypical HUS. In patients who present with the triad of Coombs-negative haemolytic anaemia, thrombocytopenia, and renal insufficiency an extensive search for an underlying abnormality is needed. This includes the measurement of the levels of complement proteins, a search for a mutation in the relevant proteins, and the detection of antibodies. Table 17.6 summarizes the recent guidelines (Taylor et al., 2010). Alternative diagnoses must be considered. The presence of aPL antibodies is characteristic for the APS.

Serologic test/IF pattern renal biopsy ANCA+ C3/C4 norm

Anti-MPO+

No extra-renal sympt PI-CGN

ICx markers + C3 low C4 low or norm

Anti-GBM+ C3/C4 norm

Anti-Pr3+

Syst vasc

Asthma eosinophilia

Resp. granulomea

MPA

EGPA

GPA

Anti-GBM

ANA+ anti-dsDNA+ anti-C1q+

[IgA] elevated purpura

Cryoglob + HCV+

Antimicrob resp+

LN

HSP

CGN

PIGN

Fig. 17.4  Diagnostic approach of the patient with a rapidly progressive glomerulonephritis. Anti-GBM = anti-GBM nephritis; CGN = cryoglobulinaemic glomerulonephritis; CSS = Churg–Strauss syndrome; HCV = hepatitis C virus; HSP = Henoch–Schönlein purpura; IC = immune-complex; [IgA] = serum IgA concentration; LN = lupus nephritis; MPA = microscopic polyangiitis; PI-CGN = pauci-immune crescentic glomerulonephritis; PIGN = post-infectious glomerulonephritis; WG = Wegener’s granulomatosis.

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Table 17.5  Positive (PPV) and negative predictive values (NPV) of serological test for lupus nephritis flare Test

Sensitivity

Specificity

PPV

NPV

OR

Anti-C1q

81

71

38

94

11.8 (4.9–18.1)

Anti-dsDNA

70

67

31

91

ND

(reduced) C3

79

51

28

93

2.99 (1.5–5.8)

(reduced) C4

74

64

31

92

3.3 (1.7–6.5)

OR = odds ratio (with 95% confidence interval).

Table 17.6  Investigations in patients with atypical haemolytic uraemic syndrome Exclude other causes Infections: Shiga toxin-producing bacteria; Streptococcus pneumonia;HIV Malignancy SLE, antiphospholipid syndrome Drugs (mitomycin, vincristine, calcineurin inhibitors) Exclude thrombotic thrombocytopenic purpura ADAMTS13 activity < 5% (genetic or immune) Disorders of complement regulation Serum: C3, C4, C3d, factor H, factor I Expression of CD46 on PMBC by FACS analysis Mutation screening: factor H, factor I, factor B, C3, CD46 Auto-antibody screening: factor H, factor B CD46 = membrane cofactor protein; PMBC = peripheral blood mononuclear cells; SLE = systemic lupus erythematosus. Adapted from Taylor et al. (2010).

Thrombotic thrombocytopenic purpura (TTP) is caused by abnormalities in the activity of serum von Willebrand factor (vWF) protease. This protease is a zinc metalloprotease called ADAMTS13, encoded by a gene on chromosome 9q34. This protease rapidly degrades large vWF multimers that are formed in the endothelial cells. A decreased activity of the protease results in the persistence of large vWF multimers in the circulation. These large multimers cause endothelial damage and promote thrombosis. Familial forms of TTP have been associated with a deficiency of this vWF protease. In several families mutations in the ADAMTS13 gene have been found (Levy et  al., 2001). The acquired form of TTP has been associated with a functional loss of vWF protease activity due to circulating antibodies. TTP typically is associated with an ADAMTS13 activity < 5% (Furlan et al., 1998). Tests for these abnormalities are only available in specialist laboratories.

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assessment of the patient with renal disease

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Kain, R., Tadema, H., McKinney, E. F., et al. (2012). High prevalence of autoantibodies to hLAMP-2 in anti-neutrophil cytoplasmic antibody-associated vasculitis. J Am Soc Nephrol, 23, 556–66. Kalantar-Zadeh, K., Miller, L. G., and Daar, E. S. (2005). Diagnostic discordance for hepatitis C virus infection in hemodialysis patients. Am J Kidney Dis, 46, 290–300. Kallenberg, C. G. (2008). Anti-C1q autoantibodies. Autoimmun Rev, 7, 612–15. Kallenberg, C. G. and Heeringa, P. (2013). Complement is crucial in the pathogenesis of ANCA-associated vasculitis. Kidney Int, 83, 16–18. Kalluri, R., Torre, A., Shield, C. F., 3rd, et al. (2000). Identification of alpha3, alpha4, and alpha5 chains of type IV collagen as alloantigens for Alport posttransplant anti-glomerular basement membrane antibodies. Transplantation, 69, 679–83. Katzmann, J. A., Clark, R. J., Abraham, R. S., et al. (2002). Serum reference intervals and diagnostic ranges for free kappa and free lambda immunoglobulin light chains: relative sensitivity for detection of monoclonal light chains. Clin Chem, 48, 1437–44. Katzmann, J. A., Kyle, R. A., Benson, J., et al. (2009). Screening panels for detection of monoclonal gammopathies. Clin Chem, 55, 1517–22. Kumar, S., Dispenzieri, A., Katzmann, J. A., et al. (2010). Serum immunoglobulin free light-chain measurement in primary amyloidosis: prognostic value and correlations with clinical features. Blood, 116, 5126–9. Lachmann, H. J., Gallimore, R., Gillmore, J. D., et al. (2003). Outcome in systemic AL amyloidosis in relation to changes in concentration of circulating free immunoglobulin light chains following chemotherapy. Br J Haematol, 122, 78–84. Lauer, G. M. and Walker, B. D. (2001). Hepatitis C virus infection. N Engl J Med, 345, 41–52. Lee, E. Y., Lee, C. K., Lee, T. H., et al. (2003). Does the anti-beta2-glycoprotein I antibody provide additional information in patients with thrombosis? Thromb Res, 111, 29–32. Levine, J. S., Branch, D. W., and Rauch, J. (2002). The antiphospholipid syndrome. N Engl J Med, 346, 752–63. Levy, G. G., Nichols, W. C., Lian, E. C., et al. (2001). Mutations in a member of the ADAMTS gene family cause thrombotic thrombocytopenic purpura. Nature, 413, 488–94. Merkel, P. A., Chang, Y., Pierangeli, S. S., et al. (1996). The prevalence and clinical associations of anticardiolipin antibodies in a large inception cohort of patients with connective tissue diseases. Am J Med, 101, 576–83. Moldoveanu, Z., Wyatt, R. J., Lee, J. Y., et al. (2007). Patients with IgA nephropathy have increased serum galactose-deficient IgA1 levels. Kidney Int, 71, 1148–54. Moll, S. and Ortel, T. L. (1997). Monitoring warfarin therapy in patients with lupus anticoagulants. Ann Intern Med, 127, 177–85. Moroni, G., Radice, A., Giammarresi, G., et al. (2009). Are laboratory tests useful for monitoring the activity of lupus nephritis? A 6-year prospective study in a cohort of 228 patients with lupus nephritis. Ann Rheum Dis, 68, 234–7. Moroni, G., Trendelenburg, M., Del Papa, N., et al. (2001). Anti-C1q antibodies may help in diagnosing a renal flare in lupus nephritis. Am J Kidney Dis, 37, 490–8. Motyckova, G. and Murali, M. (2011). Laboratory testing for cryoglobulins. Am J Hematol, 86, 500–2. Nasr, S. H., Radhakrishnan, J., and D’agati, V. D. (2013). Bacterial infection-related glomerulonephritis in adults. Kidney Int, 83, 792–803. Nochy, D., Daugas, E., Droz, D., et al. (1999). The intrarenal vascular lesions associated with primary antiphospholipid syndrome. J Am Soc Nephrol, 10, 507–18. Nojima, J., Kuratsune, H., Suehisa, E., et al. (2001). Association between the prevalence of antibodies to beta(2)-glycoprotein I, prothrombin, protein C, protein S, and annexin V in patients with systemic lupus erythematosus and thrombotic and thrombocytopenic complications. Clin Chem, 47, 1008–15.

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Olson, S. W., Arbogast, C. B., Baker, T. P., et al. (2011). Asymptomatic autoantibodies associate with future anti-glomerular basement membrane disease. J Am Soc Nephrol, 22, 1946–52. Palladini, G., Russo, P., Bosoni, T., et al. (2009). Identification of amyloidogenic light chains requires the combination of serum-free light chain assay with immunofixation of serum and urine. Clin Chem, 55, 499–504. Parra, G., Rodriguez-Iturbe, B., Batsford, S., et al. (1998). Antibody to streptococcal zymogen in the serum of patients with acute glomerulonephritis: a multicentric study. Kidney Int, 54, 509–17. Pickering, M. and Cook, H. T. (2011). Complement and glomerular disease: new insights. Curr Opin Nephrol Hypertens, 20, 271–7. Rajkumar, S. V., Kyle, R. A., Therneau, T. M., et al. (2005). Serum free light chain ratio is an independent risk factor for progression in monoclonal gammopathy of undetermined significance. Blood, 106, 812–17. Richards, N. T., Darby, S., Howie, A. J., et al. (1994). Knowledge of renal histology alters patient management in over 40% of cases. Nephrol Dial Transplant, 9, 1255–9. Rodriguez-Iturbe, B. and Musser, J. M. (2008). The current state of poststreptococcal glomerulonephritis. J Am Soc Nephrol, 19, 1855–64. Roth, A. J., Brown, M. C., Smith, R. N., et al. (2012). Anti-LAMP-2 antibodies are not prevalent in patients with antineutrophil cytoplasmic autoantibody glomerulonephritis. J Am Soc Nephrol, 23, 545–55. Rutgers, A., Slot, M., Van Paassen, P., et al. (2005). Coexistence of anti-glomerular basement membrane antibodies and myeloperoxidase-ANCAs in crescentic glomerulonephritis. Am J Kidney Dis, 46, 253–62. Savige, J., Davies, D., Falk, R. J., et al. (2000). Antineutrophil cytoplasmic antibodies and associated diseases: a review of the clinical and laboratory features. Kidney Int, 57, 846–62. Schroter, M., Feucht, H. H., Schafer, P., et al. (1997). High percentage of seronegative HCV infections in hemodialysis patients: the need for PCR. Intervirology, 40, 277–8. Sinico, R. A., Radice, A., Corace, C., et al. (2006). Anti-glomerular basement membrane antibodies in the diagnosis of Goodpasture syndrome: a comparison of different assays. Nephrol Dial Transplant, 21, 397–401. Suzuki, H., Fan, R., Zhang, Z., Brown, R., et al. (2009). Aberrantly glycosylated IgA1 in IgA nephropathy patients is recognized by IgG antibodies with restricted heterogeneity. J Clin Invest, 119, 1668–77.

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Taylor, C. M., Machin, S., Wigmore, S. J., et al. (2010). Clinical practice guidelines for the management of atypical haemolytic uraemic syndrome in the United Kingdom. Br J Haematol, 148, 37–47. Ter Borg, E. J., Horst, G., Hummel, E. J., et al. (1990). Measurement of increases in anti-double-stranded DNA antibody levels as a predictor of disease exacerbation in systemic lupus erythematosus. A long-term, prospective study. Arthritis Rheum, 33, 634–43. Terjung, B. and Spengler, U. (2009). Atypical p-ANCA in PSC and AIH: a hint toward a ‘leaky gut’? Clin Rev Allergy Immunol, 36, 40–51. Tervaert, J. W., Van Paassen, P., and Damoiseaux, J. (2007). Type II cryoglobulinaemia is not associated with hepatitis C infection: the Dutch experience. Ann N Y Acad Sci, 1107, 251–8. Tomasson, G., Grayson, P. C., Mahr, A. D., et al. (2012). Value of ANCA measurements during remission to predict a relapse of ANCA-associated vasculitis--a meta-analysis. Rheumatology (Oxford), 51, 100–9. Trendelenburg, M., Marfurt, J., Gerber, I., et al. (1999). Lack of occurrence of severe lupus nephritis among anti-C1q autoantibody-negative patients. Arthritis Rheum, 42, 187–8. Van Der Vlag, J. and Berden, J. H. (2011). Lupus nephritis: role of antinucleosome autoantibodies. Semin Nephrol, 31, 376–89. Vermeersch, P., Gijbels, K., Knockaert, D., et al. (2008a). Establishment of reference values for immunoglobulins in the cryoprecipitate. Clin Immunol, 129, 360–4. Vermeersch, P., Van Hoovels, L., Delforge, M., et al. (2008b). Diagnostic performance of serum free light chain measurement in patients suspected of a monoclonal B-cell disorder. Br J Haematol, 143, 496–502. Wang, G., Csernok, E., De Groot, K., et al. (1997). Comparison of eight commercial kits for quantitation of antineutrophil cytoplasmic antibodies (ANCA). J Immunol Methods, 208, 203–11. Weiner, S. M., Berg, T., Berthold, H., et al. (1998). A clinical and virological study of hepatitis C virus-related cryoglobulinaemia in Germany. J Hepatol, 29, 375–84. Zhao, N., Hou, P., Lv, J., Moldoveanu, Z., et al. (2012). The level of galactose-deficient IgA1 in the sera of patients with IgA nephropathy is associated with disease progression. Kidney Int, 82(7), 790–6.

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CHAPTER 18

The renal biopsy Ian S. D. Roberts, Philip Mason, and Agnes B. Fogo Introduction The first renal biopsies from living patients were obtained surgically as early as 1896 usually during the course of operations to decapsulate the kidneys in patients with Bright disease, to relieve the ‘intrarenal oedema’, a procedure popular during the first two decades of the twentieth century (Cameron and Hicks, 1997). The first needle renal biopsies were obtained accidentally at attempted liver biopsy in the 1940s. This led Alwall to use the technique to target the kidney directly using the aspiration biopsy needle developed by Iversen in 1944 (but reported in 1952 (Alwall, 1952)). It was the landmark paper by Iversen and Brun in 1951 that popularized the use of the percutaneous renal biopsy (Iversen and Brun, 1951). The technique was refined including positioning the patient prone (originally patients were biopsied sitting) and using a Vim–Silverman (non-aspiration) needle with improved success and tissue adequacy (Muehrcke et  al., 1955). Originally imaging included plain films before and during the procedure, insufflation of perirenal air to aid localization, fluoroscopy, and the use of intravenous urography. Biopsies are now generally performed using real-time ultrasound and a disposable needle with or without a biopsy gun. Initially some pathologists were sceptical that interpretation of such small tissue fragments would be useful. However, after many of the pioneer renal pathologists attended a CIBA foundation meeting in 1955, it became widely accepted. This coincided with the introduction of new techniques for treating specimens for light microscopic, immunohistochemical, and later electron microscopic examination. The percutaneous renal biopsy is now a routine and essential diagnostic tool guiding diagnosis and prognosis.

Indications for renal biopsy Inevitably there is variation between clinicians in the threshold for recommending a renal biopsy. The guiding principle should be whether the biopsy findings are likely to change management or to make a diagnosis that will have some prognostic value. The risk:benefit ratio for each individual patient needs to be taken into account, especially when the risks are higher, for example, in an obese or uncooperative patient or if clotting is impaired. Box 18.1 lists the indications, which are discussed in more detail.

Nephrotic syndrome Most nephrologists agree that biopsy of adults with nephrotic syndrome is essential to establish the diagnosis and plan treatment. It has been argued that adults with nephrotic syndrome should all be treated with a course of corticosteroids (for presumptive

steroid responsive minimal change disease (MCD)) and only those not responding should be biopsied. Two decision analysis papers supporting this policy concluded that routine treatment with high-dose steroids would be associated with fewer complications and lower mortality (Hlatky, 1982; Levey et  al., 1987). However, these were based on old pre-ultrasound complication data and assumed that corticosteroids would be given for MCD, membranous nephropathy, and possibly also membranoproliferative glomerulonephritis. Furthermore, since some adults with MCD may take up to 12 weeks to respond to corticosteroids, the potential risks of high-dose steroids in up to 75% of patients without MCD are themselves significant. Repeated biopsy may be indicated, for example, in patients whose first biopsy suggested MCD but did not respond, or become corticosteroid dependent or resistant to exclude an alternative diagnosis, for example, focal segmental glomerulosclerosis (FSGS) (see Chapter 57). The situation is different in children in whom > 75% (> 90% aged 1–10 years) have steroid sensitive nephrotic syndrome. Most paediatricians recommend biopsy only in those who have not responded to corticosteroids or who have atypical features such as low complement concentrations or microscopic haematuria (although this latter finding is not be particularly discriminative (International Study of Kidney Disease in Children, 1978)).

Acute kidney injury Most acute kidney injury (AKI) is due to prerenal disease, ‘acute tubular injury’ (ATI), or obstruction (see Chapter  220) and the underlying cause is usually obvious and biopsy is unnecessary. Only approximately 4% of AKI is secondary to glomerulonephritis (with or without vasculitis) or acute interstitial nephritis. If the cause is unclear, a biopsy will clarify the diagnosis and inform treatment which might prevent irreversible renal injury. This is particularly important for glomerulonephritis associated with vasculitis, lupus, anti-glomerular basement membrane (GBM) disease and myeloma-associated kidney injury. It is important to remember that patients with these underlying conditions may be systemically ill and therefore also have ATI. Clinical features of systemic diseases, laboratory tests (e.g. antineutrophil cytoplasmic antibodies (ANCAs), antinuclear antibodies (ANAs), complements, anti-GBM antibodies, and myeloma screen) or heavy proteinuria may suggest one of these underlying conditions. A biopsy may also be indicated in patients who fail to recover after an episode of AKI with an identified precipitating cause in order to exclude another diagnosis or to determine the likelihood of recovery.

chapter 18 

Box 18.1  Indications for native renal biopsy ◆ Nephrotic syndrome (except children) ◆ Acute kidney failure without clear cause ◆ Chronic kidney impairment especially with proteinuria and/or haematuria ◆ Non-nephrotic proteinuria ◆ Microscopic haematuria ◆ Systemic diseases with abnormal function and/or heavy proteinuria.

Systemic disease

the renal biopsy

patients are keen to have a diagnosis and are prepared to accept the small risks of the procedure to obtain this.

Microscopic haematuria The arguments for and against biopsy in this setting are similar to those for ‘non-nephrotic’ proteinuria and both may coexist. Isolated haematuria (without collateral evidence of renal disease such as proteinuria), especially in those aged over 40–50  years, needs a urological diagnostic work-up, but renal biopsy is unlikely to change management. Biopsies in this setting most often reveal immunoglobulin (Ig)-A nephropathy IgA or other conditions that are unlikely to progress (Topham et  al., 1994). Individuals with microscopic haematuria being worked up as potential kidney transplant donors should be biopsied since it is important to exclude a glomerulonephritis such as IgA nephropathy.

Patients with known systemic diseases including systemic vasculitis, anti-GBM disease, and systemic lupus erythematosus (SLE) are often biopsied. It has been argued that patients who test positive for ANCA or anti-GBM antibodies do not need a biopsy, since management will not be altered. However, a biopsy may be helpful to assess the acuteness of the illness and the degree of chronic and irreversible damage. Although not very predictive of response to treatment in ANCA-positive disease, and accepting potential sampling bias, the renal biopsy may guide management, for example, in an elderly patient with evidence of long-standing disease (with fibrocellular or fibrous crescents) and much interstitial fibrosis in whom continued immunosuppression may carry significant and unwarranted risks. The decision to biopsy must always be made on an individual basis.

Transplant kidneys

Diabetic patients

Before renal biopsy, the patient should have a renal ultrasound to establish the presence of two normal-sized unobstructed kidneys. Biopsy of a small kidney is associated with higher risks of bleeding and is less likely to provide diagnostic information (small scarred kidneys often show sclerosed glomeruli and interstitial fibrosis with no features of the original disease). The size of the kidney needs to be interpreted in the context of the patient’s size and the cortical thickness should be assessed. Biopsy of a kidney with a cortical thickness of < 1 cm has a higher risk of complications and a lower chance of obtaining diagnostically useful tissue. Anatomic abnormalities, for example, ectopic kidneys, crossed fused ectopia, and horseshoe kidneys, increase the risk of bleeding. Any of these factors need to be taken into account when estimating the balance of risk over benefit for the individual patient. The patient should be tested for any bleeding tendency and have normal clotting tests, a platelet count ≥ 100,000 and not be severely anaemic (haemoglobin < 9 g/dL). The value of the bleeding time test has been controversial. Although there have been reports that it is predictive of complication (Stratta et al., 2007) (although this was only significant for haematoma > 1 cm and not other major or minor bleeding complications), most studies have not found this to be so (Peterson et al., 1998; Manno et al., 2004). Bleeding time in uraemia can be partially reversed with an infusion of DDAVP (0.3 micrograms/kg). The effect is maximal after 30–60 minutes, wearing off by 24 hours. Although not evidence based, many recommend this for patients with poor renal function (e.g. urea > 20 mmol/L or creatinine > 200 µmol/L). Patients with ischaemic heart disease should not be given DDAVP because of the small risk

Diabetic nephropathy commonly occurs in patients with long-standing diabetes. The UKPDS study found that in people with type 2 diabetes the prevalence of microalbuminuria, macroalbuminuria, or an elevated creatinine/renal failure was 25%, 5%, and 0.8% respectively at 10 years following diagnosis (Adler et al., 2003) (see Chapter 149). Patients with diabetic nephropathy usually have evidence of retinopathy and/or neuropathy and when these are present, a biopsy is not usually indicated. In the absence of these complications or with rapid-onset nephrotic syndrome, a short duration of (diagnosed) diabetes and possibly with significant haematuria or systemic disease, a biopsy is indicated to determine if another treatable condition is present. Even when another cause for renal disease is found there is often evidence of diabetic nephropathy as well (Pham et al., 2007).

Proteinuria without the nephrotic syndrome Opinion and threshold for biopsy of patients with ‘non-nephrotic proteinuria’ varies. It is difficult to argue benefit in patients with relatively low-level proteinuria (< 1.5 g/day or PCR of < 150 mg/mmol) and normal renal function. It is true that low-level proteinuria may precede the nephrotic syndrome for most causes of this clinical syndrome (except for MCD). Management of such patients with good blood pressure control and maximizing angiotensin-converting enzyme inhibitor/angiotensin receptor blocker therapy would be the same unless nephrotic syndrome and/or renal impairment were to develop. The threshold for biopsy is lowered as the proteinuria increases but the arguments are essentially the same. Sometimes

Some units routinely take biopsies during implantation either before and/or after reperfusion. This may be helpful in defining the presence or degree of existing disease in the donor kidney, particularly important in older donors. Patients with delayed graft function should be biopsied regularly, even weekly, until function returns to exclude the development of rejection as ATN recovers. This policy may vary depending on the induction therapy (early rejection is unlikely with anti-T-cell-agent induction). Later in the post-transplant course the kidney is biopsied to investigate dysfunction or heavy proteinuria (see Chapters 286 and 289).

The procedure Pre-biopsy evaluation

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Box 18.2  Safety conditions for renal biopsy ◆ Haemoglobin > 9 g/dL ◆ Platelets >100,000 × 109/L ◆ Prothrombin (PT) and activated partial thromboplastin time (APPT) < 1.2 × control ◆ Blood pressure preferably < 160/95 mmHg ◆ Sterile urine.a a

See text.

of coronary artery thrombosis. The use of the thromboelastogram (TEG) has been reported in a single study to be predictive of bleeding following transplant biopsies although the evidence was a 4% drop in haematocrit and only 2% of patients had gross haematuria or a haematoma on ultrasound (Davis and Chandler, 1995). Antiplatelet agents including aspirin, clopidogrel, non-steroidal anti-inflammatory drugs, omega-3 fatty acids, glycoprotein IIb/ IIIa inhibitors, and dipyridamole, all of which increase bleeding time, should be stopped a week before a routine biopsy. Biopsy of patients on aspirin and clopidogrel should be avoided but if absolutely essential then platelet transfusion will reduce the risk of bleeding. Anticoagulants should be reversed or stopped before biopsy and the decision to change to heparin needs to be considered on an individual patient basis. Unfractionated heparin should be stopped at least 6 hours and low-molecular-weight heparin 24 hours before biopsy. Correction of clotting abnormalities with fresh frozen plasma or clotting factors may be necessary for an urgent biopsy. Box 18.2 lists the recommended criteria and Box 18.3 the relative contraindications for renal biopsy. The concern about performing renal biopsy in patients with urinary tract infection is the risk of a post-biopsy perinephric haematoma becoming infected. Anxious patients may need sedation (e.g. with midazolam) to improve the safety of the procedure. A single kidney is often listed as a contraindication but this is relative (all transplant biopsies are of course single functioning kidneys) and for most patients the risk is lower than an open biopsy. Pregnancy is not a contraindication but it is important to be clear that the biopsy will substantially affect management before delivery.

Renal biopsy technique Consent should be taken before the procedure but counselling should be offered as early as possible. An advice leaflet allows the Box 18.3  Relative contraindications for renal biopsy ◆ Single kidney ◆ Small kidneys with thin cortex ◆ Abnormal anatomy including cysts ◆ Obesity ◆ Uncooperative patient ◆ Uncontrolled hypertension ◆ Infection, especially acute pyelonephritis/perinephric abscess.

patient to consider the options at leisure and make a note of questions. The possible complications and their incidence should be provided. These would include the following: ◆ Mild to moderate back/loin pain which usually settles with simple analgesia such as paracetamol ◆ Visible haematuria (up to 5%) which usually clears spontaneously within 24 hours ◆ Local bleeding always occurs but is usually minor and self-limiting; more severe bleeding requiring transfusion (~ 1%) ◆ Bleeding requiring angiographic intervention (≤ 0.5%). See Appendix 18.1 for a standard patient information leaflet which is used in the author’s unit. Biopsies are generally performed under direct ultrasound control using a disposable Tru-Cut® needle or biopsy gun. Biopsy under direct vision has a higher success and lower complication rate than using ultrasound merely for localization. Choice of needle gauge (G) is a matter of personal preference; it is customary to use a 16-G needle for native and 18-G for transplant biopsies. These generally provide adequate sample size with a trend to lower bleeding rates in these single kidneys which may require multiple biopsies. Either kidney may be biopsied, but the left kidney is usually more convenient as it is usually lower so access to the pole is easier. The patient lies prone on one or two firm pillows and the operator first determines the optimum entry site with ultrasound and in which phase of respiration the biopsy will best be performed, aiming for the lower pole through the cortex. The skin is sterilized, sterile gel applied, and the probe covered with a sterile sheath. Local anaesthetic is administered and then, using a spinal (21-G) needle, more local anaesthetic is delivered down to the capsule of the kidney. The biopsy needle is then advanced to the renal capsule under ultrasound visualization, the patient stops breathing as the device is fired and immediately removed, after which the patient can breathe normally. Generally two passes are made to obtain two cores of tissue. Patients unable to lie prone may be biopsied in the sitting position as for a pleural drain. Transplant biopsies are usually more straightforward because the kidney is superficial and does not move with respiration. However, transplant kidneys sometimes lie in unusual orientations and more care needs to be taken to avoid bowel (especially with intraperitoneal kidneys). It is also useful to use colour Doppler to identify large blood vessels, such as the iliac artery and vein, which may lie very close to the lower pole. Patients are kept on bed rest for 4 hours with pulse and blood pressure monitoring and the urine tested for the presence of blood. Renal biopsy is usually performed as a day-case procedure. Patients are discharged home approximately 6 hours after the procedure provided there are no signs of bleeding and the patient will not be alone for the first night. Patients with macroscopic haematuria are kept under observation until this settles (and usually overnight). Very obese patients and those in whom ultrasound localization is difficult may be biopsied using computed tomography (CT) guidance. Other techniques have been described including open surgical and laparoscopic biopsy and, in patients with uncorrectable clotting abnormalities, biopsy is performed via the internal jugular vein but bleeding complications can still occur and the samples are small and often difficult to interpret.

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Complications of renal biopsy Reported complication rates vary widely depending on the era and techniques used. Needle sizes vary from 14 to 18 G and may be automatic or manual with imaging varying from fluoroscopy to ultrasound used for localization or in real time, making comparison unreliable. The best estimates are from a systematic review and meta-analysis, which analysed 34 studies including nearly 9500 adult native kidney biopsies using an automatic needle (14–18 G) under real-time ultrasound (Corapi et al., 2012) and a similar sized review from the Norwegian Kidney Biopsy Register (Tondel et al., 2012) (Table 18.1). As might be expected, apart from needle size, significant haemorrhage is more likely in patients with bleeding abnormalities, poorer renal function (especially acute kidney injury) and higher blood pressures, emphasizing the importance of ensuring that the benefits of biopsy outweigh the risks and that risks are minimized as much as possible. Pain: most patients experience some discomfort in the loin once the local anaesthetic wears off and simple analgesia with paracetamol is usually sufficient. More severe pain might develop with a large haematoma or renal colic or bladder pain might result from significant macroscopic haematuria especially with clots. Haemorrhage: approximately 25% of the cardiac output flows to the kidneys and biopsy inevitably results in some bleeding. Usually the haematoma results in tamponade preventing a large bleed. Bleeding usually settles spontaneously. About 0.5% of patients require a blood transfusion with 16- or 18-G needles (2% with 14-G needles). Macroscopic haematuria with clots should be managed with an irrigation catheter. If the bleeding does not settle, the patient should be referred for angiography with embolization. This rarely fails but surgery and, at the last resort, nephrectomy are very occasionally necessary (Table 18.1). Arteriovenous fistula: arteriovenous fistulae have been reported in up to 18% (in very old literature) but are probably relatively common if looked for in the early post-biopsy period (e.g. with Doppler (Hubsch et al., 1990; Merkus et al., 1993)) but most of these settle

spontaneously within days or weeks. Occasionally a fistula increases in size causing macroscopic haematuria (often with clots) or renal impairment. These require radiological embolization. Other complications: other organs may be inadvertently biopsied, including bowel, liver, spleen, and gall bladder. These are uncommon with the use of ultrasound and an experienced operator. Occasionally a pneumothorax or haemopneumothorax may develop if the pleural cavity breached. Occasionally intracapsular bleeding, which is very painful, has been reported to cause tamponade and persistent renin–angiotensin-driven hypertension (the ‘Page kidney’) (McCune et al., 1991).

Laboratory handling of the renal tissue obtained by biopsy Once the decision to perform a biopsy has been made, it is essential that laboratory and diagnostic procedures are optimized to maximize the information obtained from the biopsy. For the diagnosis of native renal diseases, biopsy and laboratory protocols aim to produce an adequate core of paraffin-embedded renal tissue for light microscopy, snap-frozen renal cortex for immunostaining, and cortex containing at least one glomerulus fixed for electron microscopy (EM) if required. Ideally, the specimen should be examined under a dissecting microscope to identify renal cortex which can be divided while fresh. If this is not possible, for example, when the renal unit and laboratory are in different hospitals, the specimen may be transported in suitable fixatives for light microscopy (LM) and EM, and buffer or a transport medium if frozen tissue for immunostaining is required.

Best practice guidelines in renal pathology Guidelines for the handling and reporting of renal biopsies have been produced in the United Kingdom by the Association of Clinical Pathologists (Furness, 2000)  and The Royal College of Pathologists, and more recently by the Renal Pathology Society

Table 18.1  Incidence of complications of native renal biopsy in adults Complication

Incidence, % (N = 9474) (95% confidence interval) Meta-analysis, Corapi et al. (2012)

Incidence, % (N = 9288) Norwegian Registry (Tondel et al., 2012)

Macroscopic haematuria

3.5 (2.2–5.1)

1.9

Transfusion

0.9 (0.4–1.5)

0.9

Angiographic intervention

0.6 (0.4–0.8)

0.2a

Nephrectomy

0.01

NR

Deathb

0.02

0

Haematomac

85c

NR

Arteriovenous fistulad

NR

a Angiographic intervention and surgery recorded together; nephrectomy, if any, not separately reported. b Two deaths from 8971 procedures reported in a meta-analysis. c Haematoma incidence is unhelpful since small haematomas are almost inevitable and not reported in

meta-analysis or the Norwegian study but 85% in one prospective CT study (Rosenbaum et al., 1978). d Arteriovenous fistula not reported in meta-analysis or the Norwegian study (see text).

NR = not reported.

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(Chang et  al., 2012). The use of histological data in patient management is also included in several of the KDIGO guidelines (Kidney Disease:  Improving Global Outcomes (KDIGO) Glomerulonephritis Work Group, 2012). There is considerable variation between laboratories in methodology used, reflected by an audit of practice in the United Kingdom (Roberts et  al., 2001). Whilst this may in part be a result of differences in expertise and resources, it also reflects a lack of evidence base for many recommendations.

What is an adequate biopsy? An adequate biopsy is one that is sufficient for its purpose, whether that is to make a diagnosis or to provide information to allow disease classification or determination of disease activity, chronicity, and prognosis. How much tissue is required for the accurate diagnosis depends on the disease. For example, a single glomerulus may be sufficient for the diagnosis of glomerular diseases, such as IgA nephropathy or membranous glomerulonephritis, in which the lesions or immune deposits are diffuse, whilst the detection of more focal lesions, in conditions such as primary FSGS, require rather more tissue. Histological classifications of renal diseases frequently include a statement regarding how much tissue is required for application of the classification. This is usually expressed as number of glomeruli, and in many cases was decided arbitrarily. For example, the Banff schema of renal allograft pathology (Racusen et al., 1999; Solez et al., 2008) defines an adequate biopsy as containing at least ten glomeruli and two arterial cross sections. When this was formally tested, it was found that a Banff ‘adequate’ biopsy with two arteries would miss almost a half of vascular rejections, due to sampling artefact (McCarthy and Roberts, 2002). In contrast, the Oxford classification of IgA nephropathy provides an evidence-based recommendation for biopsy adequacy; a minimum of eight glomeruli should be present for glomerular scoring, a value that was determined by analysis of the 265 biopsies on which this classification was based (Roberts et al., 2009).

Stains for light microscopy Commonly used stains for renal biopsies are haematoxylin and eosin (H&E), periodic acid–Schiff (PAS), Jones’ methenamine silver, trichrome (such as Masson’s trichrome (MT), or elastic van Gieson (EVG)), and Congo red. H&E is used for overall evaluation of the biopsy and is particularly valuable in the assessment of cellular infiltrates. PAS detects polysaccharides and glycoproteins. It is used for the demonstration of basement membranes, the brush border of the proximal tubular epithelium, hyaline subendothelial deposits, and assessing mesangial cellularity. Jones’ methenamine silver stain provides detailed visualization of the glomerular basement membrane, revealing abnormalities such as membrane ‘spikes’ in membranous glomerulonephritis. Trichrome stains produce three colours, distinguishing matrix, cytoplasm, and nuclei. They are of value in assessing the amount of fibrosis; mature collagen stains blue with MT and red with EVG. EVG additionally highlights the elastica of arteries, staining it black. This is of value in the assessment of chronic vascular pathology. Congo red is used for the demonstration of amyloid deposits, which it stains pink, producing anomalous colours under polarized light (Howie et al., 2008).

The diagnostic process What is a diagnosis? A potential source of confusion in nephrology is that renal diagnoses are based on different types of information. As a result, multiple terms may be used to describe one condition. Diagnostic labels are based variously on clinical presentation, immunology, histology, and pathogenesis. For example, an ANCA-positive vasculitis (immunological) presents as a rapidly progressive glomerulonephritis (RPGN, clinical) with a pauci-immune necrotizing glomerulonephritis (histological) on renal biopsy. It is important to appreciate the overlap of these different diagnostic labels; in this example, ANCA-positive vasculitis is just one cause of RPGN and a necrotizing glomerulonephritis. The kidney has a limited number of ways to respond to injury and a single morphology may be seen in a number of very different conditions. For example, a nodular glomerulosclerosis may be seen in diabetic nephropathy, light chain deposition disease and in idiopathic nodular sclerosis. In chronic kidney disease, whatever the primary insult, the final common pathway is renal fibrosis with glomerulosclerosis and tubular atrophy. At this stage, histological changes are frequently non-specific and the challenge for the pathologist is to identify clues to the underlying disease process. Equally, one condition can produce diverse morphologies. This is particularly true for lupus nephritis and IgA nephropathy, which can result in almost any pattern of glomerular disease. Many histological ‘diagnoses’ are morphological patterns rather than diagnoses. This is best illustrated by glomerular diseases; morphological labels such as acute proliferative glomerulonephritis (GN), membranoproliferative GN, and FSGS each have a number of potential underlying causes. For glomerulonephritides, there is a link between the target of injury, the morphology, and clinical manifestation. Those conditions that selectively damage the permeability barrier (such as membranous nephropathy and MCD) produce the nephrotic syndrome, with little or no glomerular inflammation or proliferation. Necrotizing glomerular injury with rupture of capillary walls (as in vasculitic glomerulonephritis and anti-GBM disease) results in haematuria and a marked reduction in glomerular filtration rate, producing a RPGN. The exudation of fibrin and cytokines results in reactive extracapillary proliferation, producing a crescentic morphology. Those conditions (such as lupus nephritis and IgA nephropathy) associated with mesangial or subendothelial immune deposits produce glomerular inflammation with the nephritic syndrome and a mesangial or endocapillary proliferative morphology. This latter group show the most variation in morphology and clinical manifestations; when there is associated damage to the permeability barrier there may be heavy proteinuria and the nephrotic syndrome, and if there is necrosis they may present with a RPGN.

Morphological definitions and illustrations The first stage of histological diagnosis is to describe the morphological pattern of disease; to identify the type of lesion and its distribution. To be of clinical value, descriptions must be precise. Vague terminology, such as ‘proliferative glomerulonephritis’ and ‘focal sclerosis’, is to be avoided. Terms such as focal and diffuse, or mild, moderate, and severe, lack precision and reproducibility; lesions should be quantified wherever possible. For example, when a glomerular lesion is described, there should be an indication of the

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The terms focal and diffuse are used to describe the proportion of glomeruli involved, whereas segmental and global refer to the extent of involvement within individual glomeruli. Unfortunately, terminology is not standardized and different definitions are used for focal versus diffuse, and segmental versus global, according to whether sclerosis or proliferation is being described. When describing sclerosis, a focal lesion is one involving some but not all glomeruli and a segmental lesion part of but not all of a tuft. When describing proliferation, a focal lesion is one involving < 50% of glomeruli (rather than < 100%), and a diffuse lesion is one involving most glomeruli, > 50% (rather than all). Similarly, a segmental proliferative lesion is one involving < 50% of a glomerular tuft and a global lesion one involving > 50% of the glomerulus.

hypercellularity:  an increased number of cells within glomerular capillary lumina. These may be endothelial cells or intravascular leucocytes. ◆ Extracapillary proliferation: hypercellularity/proliferation within Bowman’s space, producing more than two cell layers between the capillary tufts and Bowman’s capsule. This appearance is commonly referred to as a cellular crescent which is usually a result of necrosis with exudation of fibrin and cytokines. A collapsing glomerulopathy may be associated with extracapillary proliferation, producing a similar appearance to a cellular crescent. ◆ Necrosis: disruption of the glomerular basement membrane (best appreciated on a silver-stained section) with fibrin exudation and karyorrhexis. The latter may not be evident and the minimum requirement for the definition of a necrotizing lesion is extracapillary fibrin exudation. ◆ Membranoproliferative pattern (also termed mesangiocapillary pattern):  mesangial hypercellularity with thickening of capillary walls. This produces a lobular appearance to the glomerulus. Capillary wall thickening is due to duplication of the glomerular basement membrane, as a reaction to interposed cells and subendothelial or intramembranous deposits.

Proliferation

Sclerosis

percentage of glomeruli in the biopsy showing this lesion. Equally, tubular atrophy should be quantified according to the percentage of cortex involved. In this section we provide a glossary of terms and illustrate common lesions.

Glomerular lesions The normal glomerulus is illustrated in Fig. 18.1.

Distribution of lesions

Proliferation is used to describe an increase in glomerular cells that may result from infiltrating leucocytes or proliferation of endogenous glomerular cells. For this reason, the term hypercellularity is sometime more appropriate than proliferation. Hypercellularity/proliferation is subclassified according to the part of the glomerulus involved. The site of proliferation gives an indication of the underlying cause and is of prognostic and therapeutic importance. The term ‘proliferative glomerulonephritis’ is not recommended. Whilst it is frequently used to describe endocapillary proliferation, it lacks precision and is a potential source of misunderstanding. The various proliferative lesions are illustrated in Fig. 18.2 and defined as follows: ◆ Mesangial hypercellularity: more than three mesangial cells in a peripheral mesangial area in a standard 2–3-micron thick paraffin section. The central stalk of the tuft should not be used for assessing cellularity. Thicker sections give an artefactual impression of hypercellularity.

(A)

◆ Endocapillary

Sclerosis is an increase in extracellular matrix within the glomerulus. In segmental and global glomerulosclerosis, the excess matrix is associated with obliteration of capillary lumina. Globally sclerosed glomeruli that are expanded and solidified by matrix are seen in advanced diabetic glomerulopathy or in a chronic glomerulonephritis. In mesangial sclerosis and nodular glomerulosclerosis, capillaries are patent. An adhesion is continuity with matrix material between glomerular basement membrane and Bowman’s capsule that is separate from an area of sclerosis, that is, capillaries associated with an adhesion are patent. The various types of segmental sclerosing lesions are illustrated in Fig. 18.3. Glomerular obsolescence is collapse of the glomerular tuft with fibrosis in Bowman’s space, and is typical of ischaemic injury. Hyalinosis is the accumulation of non-matrix proteins, that is, insudation of plasma proteins, between the endothelium and the glomerular basement membrane. The hyaline has an amorphous

(B)

Fig. 18.1  The normal glomerulus (A = H&E; B = methenamine silver stain), showing vascular pole and juxtaglomerular apparatus at the bottom. H&E illustrates the normal mesangial cellularity with ≤ 3 mesangial cell nuclei per segment. Silver stain demonstrates the normal glomerular basement membrane that appears smooth and delicate. Note the absence of circular capillary profiles; these are only seen when the capillary wall is abnormally thickened and rigid, as in membranous nephropathy.

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Fig. 18.2  Proliferative glomerular lesions. (a) (PAS) IgA nephropathy showing mesangial hypercellularity in which there are ≥ 4 mesangial cells in a peripheral mesangial area. (b) (H&E) Post-infectious glomerulonephritis showing endocapillary hypercellularity, in which the capillary lumina are filled with infiltrating leucocytes. (c) (H&E silver) Anti-GBM disease showing extracapillary proliferation or cellular crescent, in which there is partial tuft collapse and proliferation of cells within Bowman’s space. (d) (H&E silver) ANCA-associated vasculitis, showing necrosis with capillary wall rupture and fibrin exudation. (e, f) (H&E and silver) Membranoproliferative pattern, showing a lobular appearance of the glomerular tuft with mesangial hypercellularity and thickened capillary walls, with GBM duplication evident on the silver stain.

eosinophilic appearance that is best appreciated in PAS or silver/ H&E-stained sections.

Tubulointerstitial lesions Normal tubules are illustrated in Fig. 18.4. In the normal renal cortex tubules are back to back without significant intervening space. The interstitium comprises peritubular capillaries and a delicate network of myofibroblasts that are not appreciated without immunohistochemistry (IH).

Atrophy and fibrosis Tubular atrophy: atrophic proximal tubules have thick irregular basement membranes and decreased diameter. Atrophic

distal tubules are frequently increased in diameter and contain Tamm–Horsfall protein casts. Sheets of tubules with this appearance are sometimes described as ‘thyroidization’ due to their resemblance to thyroid follicles. Interstitial fibrosis is an increase in extracellular matrix that separates the tubules. There is pronounced proliferation of myofibroblasts in early fibrosis that precedes matrix production. The distribution of interstitial fibrosis and tubular atrophy gives an indication of its aetiology and is illustrated in Fig. 18.4. Sharply delineated segmental atrophy/fibrosis is seen in reflux nephropathy or following healing of cortical infarcts. A multifocal distribution is typically seen in chronic glomerulonephritis or small vessel disease. Diffuse peritubular fibrosis follows inflammatory tubular

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Fig. 18.3  Sclerosing glomerular lesions. (A) (H&E silver) Glomerular tip lesion, showing adhesion and hyalinosis at the tubular pole. This lesion reflects severe proteinuria and may be seen in a number of nephrotic conditions, including primary FSGS, membranous nephropathy and diabetic nephropathy. Patients with tip lesions and otherwise normal glomeruli by light microscopy have a clinical course similar to minimal change disease. (B) (H&E silver) Collapsing lesion, showing tuft collapse with proliferation and swelling of visceral epithelial cells, an indicator of severe podocyte injury. (C) (PAS) Perihilar segmental sclerosis, typical of secondary FSGS, associated with hyperfiltration injury. (D) (H&E) Segmental sclerosis, not otherwise specified, typical of primary nephrotic FSGS. (E) (H&E) A broad-based bland segmental scar, typical of sclerosis following segmental necrosis in vasculitic glomerulonephritis. (F) (H&E) Nodular glomerulosclerosis in diabetic nephropathy, showing patent capillary loops around the mesangial nodules of matrix.

injury, such as a tubulointerstitial nephritis (TIN) in native kidneys or tubulointerstitial rejection in transplants.

Inflammation The significance of interstitial inflammation depends upon the nature of the cells, their distribution and the presence of tubulitis (infiltration of tubules by the inflammatory cells). Examples are illustrated in Fig. 18.5. Tubular atrophy and interstitial fibrosis are usually accompanied by an infiltrate of small lymphocytes, frequently with lymphoid aggregates, whatever the aetiology of the

chronic damage; such infiltrates are not a marker of TIN, but may still be contributing to the fibrotic process. Features of an active TIN are a mixed infiltrate, frequently including plasma cells and eosinophils, around non-atrophic tubules with an associated lymphocytic tubulitis. The type of inflammation may give an indication to the aetiology. Plasma cell-rich infiltrates are typically seen in TIN associated with systemic autoimmune diseases such as Sjögren syndrome and IgG4-associated disease. In the latter, there are numerous IgG4-positive plasma cells and an expansile fibroproliferative interstitial reaction that may result in renal mass lesions on

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Fig. 18.4  Tubular atrophy and interstitial fibrosis. (a, b) Normal renal tubules on H&E/silver and PAS stains. The proximal tubules have abundant eosinophilic cytoplasm. PAS stain demonstrates the brush border on the luminal surface. The tubular basement membranes are closely apposed with very little interstitium evident. (c, d) (H&E and PAS) Focal tubular atrophy, typically seen in chronic glomerular disease. Atrophic proximal tubules are shrunken and have thick basement membranes, highlighted on PAS stain. (e) (H&E silver) Chronic calcineurin inhibitor toxicity showing a striped pattern of atrophy and fibrosis, reflecting the small vessel disease. (f) (H&E) Diffuse peritubular fibrosis in chronic tubulointerstitial nephritis.

imaging. Eosinophils are frequently prominent in drug-associated TIN, and there may also be granulomata. Multiple, often confluent discrete epithelioid granulomata are typical of renal sarcoidosis. Occasional intratubular neutrophils are common in TIN, and do not necessarily indicate the presence of an infective nephritis. Features of acute infective nephritis/pyelonephritis are interstitial oedema with a peritubular infiltrate of neutrophils, a neutrophilic tubulitis and plugs of neutrophils within tubular lumina (see below).

Tubular casts Examples of tubular casts and crystals are illustrated in Fig. 18.6. Red blood cell casts are a marker of glomerular haematuria, unless there has been renal trauma. However, rare such casts may also be

seen in acute TIN. The red cells are frequently dysmorphic and show lysis. Large numbers of red blood cell casts may result in acute kidney injury, even in the absence of severe necrotizing glomerular lesions. Protein casts: the most frequent protein casts are formed largely by Tamm–Horsfall protein (uromodulin), produced in the thick ascending limb of the loop of Henle. These are seen in atrophic distal tubules or in association with filtered proteins in proteinuric states. These casts have a uniform hyaline appearance and, as uromodulin is a glycoprotein, are PAS positive. Other frequently seen casts include (a) light chain casts in myeloma/light chain cast nephropathy, which are PAS negative and have a hard crystalline, fractured appearance with an associated inflammatory reaction;

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Fig. 18.5  Tubulointerstitial inflammation. (a) (H&E) A non-specific infiltrate of small lymphocytes associated with an area of scarring. (b) (H&E) Tubulointerstitial nephritis, showing a mixed interstitial infiltrate of lymphocytes, plasma cells, eosinophils, and neutrophils. (c) (H&E) Renal sarcoidosis, characterized by non-necrotizing epithelioid granulomata. (d) (H&E) A severe neutrophilic tubulitis with neutrophil casts, indicative of ascending bacterial infection. (e, f) IgG4-associated tubulointerstitial nephritis showing a plasma cell-rich interstitial infiltrate. IH (F) demonstrates numerous IgG4-positive plasma cells.

and (b) myoglobin or haemoglobin casts associated with rhabdomyolysis and intravascular haemolysis respectively. These typically have a granular or beaded appearance. Greenish pigmented bile casts with associated ATI may be present in patients with severe liver disease. Crystals: the most frequently seen intratubular crystals are calcium oxalate and calcium phosphate. Calcium oxalate crystals are often sheave-shaped and do not take up standard histochemical stains and are best visualized under polarized light. When extensive, they are indicative of hyperoxalaemia and hyperoxaluria, but are seen in small numbers following ATI. Tubular deposits of calcium phosphate are strongly haematoxyphilic and do not polarize. They are seen in hypercalcaemic states or with large loads of

ingested phosphate, and less frequently following severe ATI, when intraluminal cell debris acts as a nidus for calcification.

Acute tubular injury This may be ischaemic or toxic in aetiology (Fig. 18.7). In ischaemic ATI, there is tubular dilatation and simplification of the epithelium without tubular basement membrane thickening. In toxic injury, there may be cytopathic changes within tubular epithelium, such as cytoplasmic swelling and vacuolation. In severe injury, degenerated epithelial cells are shed into the lumen.

Viral inclusions Certain viral infections, such as polyoma viruses, cytomegalic virus (CMV), and adenovirus, produce characteristic inclusions within

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Fig. 18.6  Tubular casts and crystals. (A) (PAS) PAS-positive Tamm–Horsfall protein (uromodulin) casts in distal tubules. (B) (PAS) Light chain cast nephropathy, characterized by PAS-negative fractured casts with associated inflammation and tubular epithelial injury. (C) (H&E) Granular eosinophilic haemoglobin casts in a patient with AKI associated with intravascular haemolysis. (D) (H&E) Red cell casts, indicative of glomerular haematuria, in a patient with vasculitic glomerulonephritis. (E) (H&E) Calcium phosphate deposits within tubules in a patient with hypercalcaemia and nephrocalcinosis. (F) (H&E polarized light) Calcium oxalate crystals in a patient with AKI associated with enteric hyperoxaluria.

tubular epithelial nuclei. These may be associated with evidence of epithelial injury and an inflammatory reaction (viral nephritis). Special immunostains can identify the specific virus.

Vascular lesions A normal intravenal supply is illustrated in Fig. 18.8. Arcuate arteries at the corticomedullary junction give off radial branches, the interlobular arteries that extend towards the renal capsule. From these branch afferent arterioles, which supply blood to the glomeruli. Arterial and arteriolar lesions may be acute or chronic; the latter may reflect vascular remodelling following earlier acute injury.

Thrombotic microangiopathy describes the morphology of acute microvascular injury, also termed malignant vascular injury (Fig. 18.8). Endothelial activation and injury result in a subendothelial exudate that frequently contains fibrin and red blood cells. There may be luminal thrombosis, particularly of arterioles and glomerular capillaries. The chronic phase is characterized by a proliferative response within the intima (‘onion skin proliferation’) and progressive intimal fibrosis. Arteriosclerosis describes remodelling of the arterial intima, largely comprising elastic fibres, also termed fibroelastosis (Fig. 18.8). This is a ‘normal’ age-related change, accelerated in association with long-standing hypertension.

chapter 18 

(A)

(B)

(C)

(D)

(E)

(F)

the renal biopsy

Fig. 18.7  Acute tubular injury (H&E). (A) Ischaemic acute tubular injury showing widespread tubular dilatation and epithelial simplification. (B) Cytotoxic acute tubular injury, with marked epithelial swelling and degenerative changes, in this case secondary to gentamicin toxicity. (C) Isometric tubular epithelial vacuolation in a renal transplant patient with Tacrolimus toxicity. (D–F) Viral infections in renal transplant patients. Polyoma virus infection (D) with basophilic nuclear inclusions. Adenovirus infection (E) characteristically produces marked tubular necrosis. CMV infection (F) with eosinophilic nuclear inclusions.

Arteritis is characterized by focal necrotizing vascular lesions. A  necrotizing glomerulonephritis, frequently with crescents, is the most frequent lesion in renal involvement by systemic small vessel vasculitis (granulomatosis with polyangiitis, microscopic polyangiitis, eosinophilic granulomatosis with polyangiitis (formerly known as Churg–Strauss vasculitis)), with arterial involvement seen in approximately 20% of cases. Less common is renal involvement in polyarteritis nodosa that involves the larger arcuate and segmental arteries, producing segmental necrosis and aneurysms.

Quantification of histological lesions The renal biopsy provides important information regarding disease activity and chronicity and may therefore be used to guide therapy. Indicators of active disease are necrosis, inflammation, and proliferation, lesions that are potentially responsive to immunosuppressive therapy. Chronic injury is characterized by glomerulosclerosis, interstitial fibrosis, tubular atrophy, and arteriosclerosis, lesions that have been regarded as largely irreversible. It is important for the biopsy report to include measures of the extent of acute and chronic lesions.

153

(A)

(B)

(C)

(D)

(E)

(F)

(G)

(H)

Fig. 18.8  Vascular lesions. (A) (H&E) A normal interlobular artery. (B–D) Arterial TMA. In the acute phase (B), endothelial injury results in marked intimal oedema. This progresses to a healing phase, initially with a proliferative intimal reaction (C) and finally obliterative intimal fibrosis (D, trichrome). (E) (elastic van Gieson) Arteriosclerosis (fibroelastosis), associated with essential hypertension, showing reduplication of the internal elastic lamina. (F) (H&E) A necrotizing arteritis in a patient with ANCA-positive vasculitis. (G) (H&E) Normal afferent and efferent arterioles at the glomerular hilum. (H) (H&E) Marked arteriolar hyalinosis in a patient with diabetic nephropathy.

chapter 18 

Glomerular lesions The total number of glomeruli, together with the number showing each of the glomerular lesions described above, are routinely included in the biopsy report. The percentage of glomeruli showing necrosis, proliferation, crescents, and segmental or global sclerosis has been shown to be of prognostic value in many diseases. For some conditions, this quantitative data forms the basis of a classification, the details of which are disease specific. Widely used schemas include the ISN/RPS classification of lupus nephritis (Weening et  al., 2004)  and the Oxford classification of IgA nephropathy (Cattran et al., 2009).

Tubulointerstitial lesions The extent of tubular atrophy/interstitial fibrosis should be assessed semiquantitatively and is most commonly expressed as a percentage of the cortical area present, to the nearest 10%. Various methods of measuring interstitial fibrosis and tubular atrophy are used (Farris and Colvin, 2012). Tubular atrophy is best visualized on PAS- or silver-stained sections which highlights the thickened basement membranes of atrophic tubules. Trichrome stains demonstrate established interstitial fibrosis but do not reveal the earlier stages characterized by proliferation of interstitial myofibroblasts and production of an immature matrix. Use of quantitative image analysis methods to assess fibrosis may help to improve reproducibility (Farris et al., 2011). The reproducibility of percentage interstitial fibrosis and tubular atrophy depends in part on the pattern of chronic tubulointerstitial damage. For conditions, such as chronic glomerulonephritis, that are associated with a multifocal pattern of atrophy and fibrosis, reproducibility is high, despite observers using different approaches to measurement. In contrast, reproducibility is low for conditions in which there is diffuse cortical involvement such as in chronic damage associated with TIN. Tubular atrophy is a process that develops over time, and the poor reproducibility of the diffuse pattern of chronic tubular injury reflects the difficulty in defining at what point a tubule should be regarded as atrophic. It is also recommended that the extent of interstitial inflammation is assessed. Inflammation is of different significance depending on whether it involves fibrotic or non-fibrotic cortex. In renal transplant biopsies, mononuclear inflammatory cell infiltration in non-fibrotic cortex is a criterion used in the diagnosis of T-cell-mediated rejection, although it has been shown to be poorly reproducible, particularly in the presence of chronic damage (Furness and Taub, 2001).

Arterial lesions It is recommended that lesions of arteries are scored based on the most severe lesions. As there are usually few arteries in a renal biopsy, assessment of arterial lesions is subject to the greatest sampling artefact. Intimal thickening is scored by comparing the thickness of the intima to that of the media in the same segment of vessel, thus allowing for variation in thickness resulting from oblique sectioning of the vessel. The intima is scored variously as normal, and thickened to more or less than the thickness of the media.

Immunohistology Immunohistology is routinely used to demonstrate deposits of immunoglobulins, light chains, and complement. In renal

the renal biopsy

transplantation it is valuable in the diagnosis of viral infections such as polyoma virus and CMV and for detection of complement C4d as a marker of antibody-mediated rejection. Other uses include the typing of leucocyte infiltrates, classification of amyloid deposits, identification of myoglobin casts, and characterization of abnormal matrix proteins. Immunofluorescence microscopy (IF) on frozen sections and IH on paraffin sections may both be used for the demonstration of Ig and complement deposits. IF has the advantage of greater sensitivity and it enables quantitative histology; unlike IH, the level of fluorescence signal correlates closely with the amount of antigen present. Demonstration of linear IgG deposits in anti-GBM disease, and light chain restriction in light chain deposition disease, are frequently only possible using IF on frozen sections. IH has the advantage that it is performed on the same paraffin block as is used for routine light microscopical stains and may be used when there is no frozen core of tissue available for IF. The greater detail of tissue structure in paraffin sections enables more precise localization of deposits using IH, and unlike IF, IH sections are permanent. IH does suffer from greater laboratory artefacts, such as false positivity resulting from plasma proteins in the tissue sections, and false negative stains resulting from masking of epitopes by formalin fixation. Various methods of specimen washing and antigen retrieval are described to minimize these artefacts. Glomerular deposits seen on immunohistology are described by their location (mesangial vs capillary wall; segmental vs global; focal vs diffuse) and appearance (linear, finely granular, coarse granular, pseudolinear). Tubular and vascular deposits are described when present. Clonal cast staining is described. Examples are illustrated in Fig. 18.9.

Electron microscopy EM is valuable, particularly in the diagnosis of glomerular diseases and recurrent disease post transplantation. EM blocks should be prepared for all native renal biopsies. In some centres, EM reports are issued for all cases, whilst in others the pathologist selects which to examine ultrastructurally on the basis of the LM and IF findings. EM is essential for diagnosis in approximately one-quarter of native renal biopsies (Pearson et al., 1994). The ultrastructure of the normal glomerulus is illustrated in Fig. 18.10. EM provides precise localization of glomerular deposits, and reveals internal structure to deposits. It is essential for the diagnosis of fibrillary and immunotactoid glomerulonephritis, conditions that are defined by their ultrastructural appearance. Mesangial, subendothelial, subepithelial and intramembranous deposits are illustrated in Fig. 18.10. In addition to demonstration of deposits, EM is used to quantify the extent of podocyte foot process effacement in proteinuric conditions. Abnormalities of the GBM usually require EM for diagnosis. Whilst localized thinning of the lamina densa may be seen at sites of injury, diffuse thinning is indicative of thin membrane nephropathy or early stage or a carrier state of Alport syndrome. In established renal involvement in Alport syndrome, there is multilayering of the thin lamina densa, producing a basket-weave appearance. Amorphous thickening of basement membranes is a feature of diabetic glomerulopathy. EM plays a central diagnostic role in the diagnosis of certain storage

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(B)

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Fig. 18.9  Immunofluorescence. (a) Linear GBM positivity for IgG in anti-GBM disease. (b) Granular capillary wall positivity for IgG in membranous nephropathy. (c) Mesangial positivity for IgA in IgA nephropathy. (d) Mesangial and capillary wall positivity for C3 in C3 glomerulonephritis. (e) Mesangial and tubular basement membrane positivity for kappa light chains in light chain deposition disease. (f) Positivity for lambda light chains in tubular casts in light chain cast nephropathy.

diseases, including Fabry disease and lecithin cholesterol acyltransferase deficiency.

Diagnostic algorithms In Table 18.2, the characteristic features of some selected glomerular diseases are summarized. In the final section, we present a series of diagnostic algorithms for glomerular diseases (Fig. 18.11), based

on selected appearances at LM. These illustrate how morphology is refined to a diagnosis using IF, EM, and clinical data. The algorithms are not intended to be comprehensive and do not include some of the rarer causes of these patterns of disease. In the first algorithm, in which glomeruli are normal by LM, the range of diagnoses considered is dependent upon the clinical features and urinary findings, emphasizing the importance of clinicopathological correlation in the diagnostic process.

(A)

(B)

(C)

(D)

(E)

(F)

(G)

(H)

Fig. 18.10  Electron microscopy. (A) The normal capillary wall, comprising lumen at the bottom, lined by a fenestrated endothelium. The outer aspect of the basement membrane is covered by podocyte foot processes. The podocyte cell body is at the top. (B) Minimal change disease, showing diffuse effacement of podocyte foot processes. (C) Paramesangial deposits in IgA nephropathy. (D) Subepithelial deposits with membrane spikes in membranous nephropathy. (E) Subendothelial deposits in C3 glomerulonephritis. (F) Intramembranous electron dense deposits in dense deposit disease. (G) Mesangial deposits of IgG with a fibrillary internal structure in fibrillary glomerulonephritis. (H) Lipid inclusions within podocytes and endothelial cells in a patient with Fabry disease.

Table 18.2  Summary of characteristic abnormalities of selected glomerular diseases Disease and typical clinical presentation

LM pattern

IF

EM, other findings

Mesangia

Subepithelial

Subendothelial

+

+ (wire loop)

Mesangial and capillary wall deposits Reticular aggregates in endothelial cells

−

Mesangial ± subendothelial deposits

Haematuria/nephritis Lupus ISN/RPS class IV (diffuse proliferative)

Endocapillary proliferation in > 50% of glomeruli, wire loops

+

IgA nephropathy

Mesangial proliferation

+ IgA dominant

−

Post-infectious GN

Endocapillary proliferation intracapillary neutrophils

−

+ Coarse chunky C3 dominant

Haemolytic-uraemic syndrome

Arteriolar/glomerular thrombosis Endothelial swelling

−

−

−

Endothelial injury Expansion of lamina rara interna

Minimal change disease

Normal

−

−

−

Effacement of podocyte foot processes

Focal segmental glomerulosclerosis

Segmental glomerulosclerosis

± IgM, C3

−

−

Effacement of podocyte foot processes, no deposits

Diabetic nephropathy

Increased mesangial matrix, ± nodules, thick GBM, hyalinized arterioles

−

−

−

Thick GBM without deposits

Membranous GN

Spikes on Jones’ silver stain

−

+ granular IgG, C3

−

Subepithelial deposits

Amyloid

Pink, acellular material in glomeruli, Congo Red +

−

−

−

Fibrils

All Igs, C3, C4

Irregular, hump-like deposits on top of GBM

Nephrotic syndrome

RPGN Anti-GBM disease

Global or segmental necrosis of linear staining of GBM glomeruli, crescents IgG, C3

Granulomatosis with polyangiitis

Focal segmental necrosis of glomeruli, crescents

−

−

−

No deposits

Microscopic polyangiitis Focal segmental necrosis of glomeruli, crescents

−

−

−

No deposits

EM = electron microscopy; IM = immunofluorescence microscopy; LM = light microscopy.

No deposits

(A)

Normal Glomerulus by LM

Isolated Haematuria IgA

Nephrotic proteinuria Negative IMF

IgAN

Thin GBM

Normal GBM

Thin GBM Alport early/carrier

Non-renal haematuria

(B)

Negative IMF

IgG + C3

MCD or unsampled FSGS

MN Stage I

Endocapillary proliferation

IgA

C3 dominant

IgAN

IgG + C3

Full House IgG,A,M,C3,C1q

Non-organised deposits

Fibrillary deposits

Infection/other ICGN

Fibrillary GN

Subepithelial humps, no DD

Subendothelial deposits

Postinfectious GN

C3 GN

Monoclonal Ig

Organised deposits Lupus nephritis

Proliferative GN with monoclonal deposits

Linear IgG

Dense deposit disease

Anti-GBM disease

Glomerular crescents

Negative IMF

Pauci-immune vasculitic GN

Cryoglobulinaemic GN

Intramembranous linear dense deposits

(D)

(C)

IgM + IgG

Double contours of GBM Positive IMF

Granular disease specific IMF

Various immune complex GN

Negative IMF

Subendothelial deposits

Subepithelial/ intramembranous deposits

Widening of LRI, no deposits

Various diseases, see endocapillary proliferation

Chronic MN

Chronic TMA

Fig. 18.11  Algorithms for analysis of renal biopsies. (A.) Normal glomeruli by light microscopy in a patient with renal disease (B.) Glomeruli show endocapillary proliferation (C.) Glomeruli show crescents (See also chapter 70) (D.) Glomerular basement membrane shows double contours (see also chapter 80). Key: Light microscopy (LM) Pink; Immunofluorescence Green; Urinalysis Yellow; Diagnoses shown in Blue. Abbreviations C3: Third component of Complement. DD: Dense deposits. GBM: glomerular basement membrane. ICGN: Glomerulonephritis characterised by immune complexes. GN: Glomerulonephritis. Ig: Immunoglobulin (hence IgA for Immunoglobulin A, IgG etc). IgAN: IgA nephropathy. IMF: immunofluorescence. LM: light microscopy. LRI: Lamina rara interna. MCD: Minimal change disease. MN: Membranous nephropathy (= membranous glomerulonephritis). TMA: Thrombotic microangiopathy.

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References Adler, A. I., Stevens, R. J., Manley, S. E., et al. (2003). Development and progression of nephropathy in type 2 diabetes: the United Kingdom Prospective Diabetes Study (UKPDS 64). Kidney Int, 63(1), 225–32. Alwall, N. (1952). Aspiration biopsy of the kidney, including a report of a case of amyloidosis diagnosed through aspiration biopsy of the kidney in 1944 and investigated at an autopsy in 1950. Acta Med Scand, 143, 430–5. Cameron, J. S. and Hicks, J. (1997). The introduction of renal biopsy into nephrology from 1901 to 1961: a paradigm of the forming of nephrology by technology. Am J Nephrol, 17, 347–58. Cattran, D. C., Coppo, R., Cook, H. T., et al. (2009). The Oxford classification of IgA nephropathy: rationale, clinicopathological correlations, and classification. Kidney Int, 76, 534–45. Chang, A., Gibson, I. W., Cohen, A. H., et al. (2012). A position paper on standardizing the nonneoplastic kidney biopsy report. Clin J Am Soc Nephrol, 7, 1365–8. Corapi, K. M., Chen, J. L., Balk, E. M., et al. (2012). Bleeding complications of native kidney biopsy: a systematic review and meta-analysis. Am J Kid Dis, 60(1), 62–73. Davis, C. L. and Chandler, W. L. (1995). Thromboelastography for the prediction of bleeding after transplant renal biopsy. J Am Soc Nephrol, 6(4), 1250–5. Farris, A. B., Adams, C. D., Brousaides, N., et al. (2011). Morphometric and visual evaluation of fibrosis in renal biopsies. J Am Soc Nephrol, 22, 176–86. Farris, A. B. and Colvin, R. B. (2012). Renal interstitial fibrosis: mechanisms and evaluation. Curr Opin Nephrol Hypertens, 21, 289–300. Furness, P. N., Taub, N., and Convergence of European Renal Transplant Pathology Assessment Procedures (CERTPAP) Project (2001). International variation in the interpretation of renal transplant biopsies: report of the CERTPAP Project. Kidney Int, 60, 1998–2012. Furness, P. N. (2000). ACP. Best practice no 160. Renal biopsy specimens. J Clin Pathol, 53, 433–8. Hlatky, M. A. (1982). Is renal biopsy necessary in adults with nephrotic syndrome. Lancet, 2(8310), 1264–8. Howie, A. J., Brewer, D. B., Howell, D., et al. (2008). Physical basis of colors seen in Congo red-stained amyloid in polarized light. Lab Invest, 88, 232–42. Hubsch, P. J., Mostbeck, G., Barton, P. P., et al. (1990). Evaluation of arteriovenous fistulas and pseudoaneurysms in renal allografts following percutaneous needle biopsy. Color-coded Doppler sonography versus duplex Doppler sonography. J Ultra Med, 9(2), 95–100. International Study of Kidney Disease in Children (1978). Nephrotic syndrome in children: prediction of histopathology from clinical and laboratory characteristics at time of diagnosis. A report of the International Study of Kidney Disease in Children. Kidney Int, 13(2), 159–65. Iversen, P. and Brun, C. (1951). Aspiration biopsy of the kidney. Am J Med, 11, 324–30. Kidney Disease: Improving Global Outcomes (KDIGO) Glomerulonephritis Work Group (2012). KDIGO Clinical Practice Guideline for Glomerulonephritis. Kidney Int Suppl, 2, 1–274.

Levey, A. S., Lau, J., Pauker, S. G., et al. (1987). Idiopathic nephrotic syndrome. Puncturing the biopsy myth. Ann Internal Med, 107(5), 697–713. Manno, C., Strippoli, G. F., Arnesano, L., et al. (2004). Predictors of bleeding complications in percutaneous ultrasound-guided renal biopsy. Kidney Int, 66(4), 1570–7. McCarthy, G. P. and Roberts, I. S. (2002). Diagnosis of acute renal allograft rejection: evaluation of the Banff 97 guidelines for slide preparation. Transplantation, 73, 1518–21. McCune, T. R., Stone, W. J., and Breyer, J. A. (1991). Page kidney: case report and review of the literature. Am J Kidney Dis, 18(5), 593–9. Merkus, J. W., Zeebregts, C. J., Hoitsma, A. J., et al. (1993). High incidence of arteriovenous fistula after biopsy of kidney allografts. Brit J Surg, 80(3), 310–12. Muehrcke, R. C., Kark, R. M., and Pirani, C. L. (1955). Technique of percutaneous renal biopsy in the prone position. J Urol, 74(3), 267–77. Pearson, J. M., McWilliam, L. J., Coyne, J. D., et al. (1994). Value of electron microscopy in diagnosis of renal disease. J Clin Pathol, 47, 126–8. Peterson, P., Hayes, T. E., Arkin, C. F., et al. (1998). The preoperative bleeding time test lacks clinical benefit: College of American Pathologists’ and American Society of Clinical Pathologists’ position article. Arch Surg, 133(2), 134–9. Pham, T. T., Sim, J. J., Kujubu, D. A., et al. (2007). Prevalence of nondiabetic renal disease in diabetic patients. Am J Nephrol, 27(3), 322–8. Racusen, L. C., Solez, K., Colvin, R.B., et al. (1999). The Banff ‘97 working classification of renal allograft pathology. Kidney Int, 55, 713. Roberts, I. S., Cook, H. T., Troyanov, S., et al. (2009). The Oxford classification of IgA nephropathy: pathology definitions, correlations, and reproducibility. Kidney Int, 76, 546–56. Roberts, I. S. D. and Davies, D. R. (2001). Handling of renal biopsies: different approaches reflect a lack of evidence for what constitutes ‘best practice’. J Clin Pathol, 54, 413–14. Rosenbaum, R., Hoffsten, P. E., Stanley, R. J., et al. (1978). Use of computerized tomography to diagnose complications of percutaneous renal biopsy. Kidney Int, 14(1), 87–92. Solez, K., Colvin, R. B., Racusen, L. C., et al. (2008). Banff 07 classification of renal allograft pathology: updates and future directions. Am J Transplant, 8, 753–60. Stratta, P., Canavese, C., Marengo, M., et al. (2007). Risk management of renal biopsy: 1387 cases over 30 years in a single centre. Eur J Clin Invest, 37(12), 954–63. The Royal College of Pathologists (n.d.). Datasets and Tissue Pathways. [Online] www.rcpath.org/publications-media/publications/datasets/ datasets-TP.htm Tondel, C., Vikse, B. E., Bostad, L., et al. (2012). Safety and complications of percutaneous kidney biopsies in 715 children and 8573 adults in Norway 1988-2010. Clin J Am Soc Nephrol, 7(10), 1591–7. Topham, P. S., Harper, S. J., Furness, P. N., et al. (1994). Glomerular disease as a cause of isolated microscopic haematuria. Q J Med, 87(6), 329–35. Weening, J. J., D’Agati, V. D., Schwartz, M. M., et al. (2004). The classification of glomerulonephritis in systemic lupus erythematosus revisited. Kidney Int, 65, 521–30.

CHAPTER 19

Clinical trials: why and how in nephrology Richard Haynes, Martin J. Landray, William G. Herrington, and Colin Baigent Introduction Randomized trials are an indispensable tool for nephrologists seeking to improve outcomes for their patients. Randomized trials are the best method for identifying (and quantifying) the benefits and risks of interventions in clinical practice, and the only method that can (when properly conducted) eliminate bias (Collins and MacMahon, 2001). The incremental improvements in survival seen in cardiology and cancer medicine are due in part to the widespread acceptance and conduct of large randomized trials. Nephrology lags far behind most specialities in medicine in its evidence base (Strippoli et al., 2004) to the detriment of patients under the care of nephrologists and also the larger number of patients with more moderate kidney disease who are cared for by doctors from other specialities. Furthermore, most trials in nephrology have been too small to provide reliable answers and therefore to change clinical practice. Many commonly used treatments currently recommended in nephrology guidelines have never been tested in an adequately sized randomized trial (e.g. phosphate binders, immunosuppressants, and dialysis techniques) and therefore their effects are not known. In some cases, they could be doing substantial harm. This lack of a robust evidence base will become more important in the coming decades as the global population ages and the epidemic of type 2 diabetes mellitus matures, with a consequent rise in the numbers of patients with kidney disease. In order to provide the best possible care for our current and future patients, there is an urgent need to design, fund, and conduct large randomized trials capable of addressing important clinical questions in nephrology.

Why do we need large randomized trials in nephrology? Moderate effects are the best that can be expected Some treatments have large, and hence obvious, effects on survival: for example, it was clear without the need for any randomized trials that prompt treatment of diabetic coma or ventricular fibrillation can save lives. However, in part because of these striking successes but also because of large effects of treatments on intermediate disease markers (such as antihypertensives on blood pressure), the hopes of large treatment effects on major clinical outcomes (such

as end-stage renal disease or mortality) in nephrology, have been unrealistically high. In general, where substantial uncertainty remains about the efficacy of a treatment on major clinical outcomes in nephrology, its effects on such outcomes are probably either negligibly small, or only moderate (i.e. a 15–20% proportional reduction in risk), rather than large (Box 19.1). This may be because a variety of pathophysiological mechanisms are responsible for progression of chronic kidney disease (CKD) or its complications (e.g. cardiovascular disease), only one of which may be appreciably influenced by any one particular therapy. The existence of (at best) moderate effects of most treatments is strongly suggested by meta-analyses of various therapies in nephrology (Jafar et al., 2001; Webster et al., 2004) and by the typically modest strength of epidemiological relationships between known causal risk factors that are modifiable by treatment (e.g. blood pressure) and outcomes that are common in CKD (such as heart failure). In nephrology, failure to acknowledge that treatment effects are likely to be, at best, moderate has led to randomized trials of promising treatments that have almost always been too small: many apparently negative trials may have missed potentially worthwhile benefits (e.g. EVOLVE, which tested cinacalcet in haemodialysis patients with secondary hyperparathyroidism (Chertow et al., 2012)) whilst on the other hand, exaggerated claims of efficacy have proved to be unrealistically large and have not changed clinical practice (e.g. trials of antioxidants in haemodialysis patients (Boaz et al., 2000; Tepel et al., 2003)).

What types of studies are required to assess promising treatments in nephrology? If moderate differences in outcome resulting from promising treatments are to be detected reliably, then any errors in studies of such treatments need to be much smaller than this. This requirement necessitates a study design that both excludes biases and minimizes random error. Randomization eliminates bias by ensuring that each type of patient can be expected (but for the play of chance) to have been allocated in similar proportions to different treatment strategies. This means that only the treatment effect and random differences should affect the final comparisons of outcome. The way to guarantee very small random differences is to design studies that include large numbers of relevant events that are potentially preventable by the treatment under consideration.

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Box 19.1  Reasons why the effects of treatment on clinical outcomes are usually moderate (but still worthwhile) ◆ Clinical outcomes have multiple causes: diseases such as atherosclerosis or progressive kidney disease have multiple causal risk factors. On the other hand, treatments are usually specific to one particular cause, so can only mitigate the risk attributable to that cause, but not the others. Therefore, claims of large treatment effects are usually not plausible. ◆ Large treatment effects on intermediate outcomes do not usually translate into large effects on clinical outcomes:  certain drugs do have large effects on measurements such as blood pressure, proteinuria, or acute rejection rates. Although these effects may be large, for the reasons given above, they are unlikely to translate into large effects on clinically important outcomes such as development of end-stage renal disease, transplant failure, or death. The easy availability of measuring such outcomes and their association with clinical outcomes often leads to the false assertion that treatments which reduce such intermediate outcomes will improve clinical outcomes. ◆ There are also some reasons specific to nephrology that may limit the magnitude of expected treatment effects: for example, the dose of drug used may be limited by either risk of nephrotoxicity or systemic toxicity due to reduced renal clearance, and patients with kidney disease are prescribed many medications which may limit compliance to study treatment. It is often claimed that by collecting enough information about various prognostic features, it is possible to make statistical adjustments to correct for any such differences between the types of patients who, in an observational (i,e. non-randomized study), receive the different treatments that are to be compared. Such methods, which are often carried out on routinely collected healthcare data (such as dialysis registries, for example), aim to achieve comparability between those entering the different treatment groups, but they cannot be guaranteed to do so, and they often fail seriously (Deeks et al., 2003). The difficulty is that some important prognostic factors may be unrecorded, while others may be difficult to assess exactly and hence difficult to adjust for reliably. Although there are examples of non-randomized studies in which the estimated effects of treatment appear quantitatively close to those observed in analogous randomized trials, there are many examples where they are not, being either quantitatively incorrect—so that drugs appear either misleadingly promising or of misleadingly low efficacy—or even qualitatively incorrect, when a harmful drug might appear effective (or vice versa) (Grady et al., 1992; Rossouw et al., 2002). This residual bias means such analyses should not be used to guide treatment decisions, however large the dataset. Random errors are minimized by ensuring that there are sufficient patients recruited to yield the number of ‘endpoints’ that is required for reliable statistical inference. However, it is not sufficiently widely appreciated just how large clinical trials need to be in order to detect moderate differences reliably. This can be illustrated by the recent Study of Heart and Renal Protection (SHARP): the 17% relative reduction in major atherosclerotic events (non-fatal myocardial infarction, coronary death, ischaemic stroke, or revascularization procedure) was statistically significant among around

9000 patients, but if this trial had only been half as large (say, 4500 patients) then it would have had about a one in three chance of missing this clinically worthwhile treatment benefit (at a type 1 error rate of 5%) (Baigent et al., 2011). When trials are not large enough to detect such treatment effects, their results can be very misleading. First, since it is mathematically impossible for small trials to be statistically significant unless they yield large treatment estimates, small trials are useless unless the drug under study is a miracle cure (in which case a randomized trial would not be necessary to identify its effect). In all other cases, such trials will not help to distinguish between effective (hence useful) treatments and those that are useless or even harmful. It is common for positive results from small single-centre trials not to be replicated when larger multicentre trials are completed. In nephrology, for example, two small trials of renal replacement therapy (RRT) for acute kidney injury appeared to show a reduction in mortality in those allocated to more intensive RRT (Ronco et al., 2000; Schiffl et al., 2002), but in subsequent multicentre trials, no such difference was found (Palevsky et al., 2008; Bellomo et al., 2009). Small trials can even be misleading about the direction of treatment effect. For example, a small trial of the inotrope vesnarinone suggested that it halved the risk of death in patients with heart failure (13 vesnarinone versus 33 placebo deaths, P = 0.002) (Feldman et al., 1993). However, when the same regimen was tested among a larger population of similar patients, mortality was in fact increased (292 vesnarinone versus 242 placebo deaths, P = 0.02) (Cohn et al., 1998). Very large trials may also have the benefit of being able to address efficacy and safety in a wide range of different types of patients, in whom treatment effects may differ.

How do we achieve large, successful randomized trials in nephrology? Assuming that a new treatment is moderately effective among patients with CKD, four main requirements need to be satisfied by a randomized trial if it is to be able to establish this fact reliably (Table 19.1): 1. Randomization of a large sample of patients (to produce a large number of endpoints) 2. Maintenance of compliance with the randomized treatment allocation (to maintain study power) 3. Ascertainment of relevant study outcomes 4. Appropriate statistical analysis.

Randomization of a large sample of patients In order to recruit large numbers of patients into trials, it is necessary to streamline trial procedures to ensure wide eligibility criteria and efficient recruitment.

Trial eligibility: using the uncertainty principle Randomization can be offered only if both doctor and patient feel substantially uncertain as to which of the trial options is best. The question then arises:  ‘Which categories of patients about whose treatment there is such uncertainty should be offered randomization?’ The answer is all of them, welcoming the heterogeneity that this will produce. (For example, either the treatment of choice will turn out to be the same for men and women, in which case the

chapter 19 

clinical trials: why and how in nephrology

Table 19.1  Main requirements for successful large clinical trials and methods to achieve them Requirement

Study design

Study conduct

Effective recruitment

Consider a pilot study Use uncertainty principle to determine eligibility Avoid unnecessary exclusion criteria Heterogeneity is a strength in a randomized controlled trial population

Pre-screen Simple consent process Detailed baseline phenotyping (including biological samples)

Good compliance

Aim for as large a difference in relevant and measurable intermediate outcome (such as biomarkers) as possible Use pre-randomization run-in period

Use procedures that maintain investigator and participant compliance Leave patient management as much as possible to managing physician

Reliable ascertainment of outcomes

Make trial long enough to accrue sufficient efficacy and safety information

Use simple case report forms

Employ on-site monitoring only where necessary

Minimize number of study visits and biological samples

Central statistical monitoring

Use routine collected data where possible to supplement study data

Review by unblinded Data Monitoring Committee

Use electronic data capture methods that check and validate data entry and point of entry

Select a clinically important outcome

Analyse by the principle of intention to treat

Ensure outcome is modifiable by study treatment

Consider appropriate methods to analyse subgroups (tests of heterogeneity and trend with allowance for multiplicity of testing)

Relevant outcomes and appropriate analysis

trial might as well include both, or it will be different, in which case it is particularly important to study both.) This approach of randomizing the full range of patients in whom there is substantial uncertainty as to which treatment option is best was used in the ASTRAL trial of renal arterial revascularization for renal artery stenosis (Fig. 19.1) (Wheatley et al., 2009): ◆ If the doctor or the patient were reasonably certain, for any reason, that they did wish to revascularize the affected kidney(s) in that particular patient, the patient was not eligible for entry into ASTRAL. ◆ Conversely, if the doctor or the patient were reasonably certain, for any reason, that they did not wish to revascularize the affected kidney(s) in that particular patient, the patient was likewise not eligible for entry into the trial. ◆ If, but only if, the doctor and patient were substantially uncertain what to recommend, the patient was eligible for randomization between the renal revascularization (by angioplasty with or without stenting) versus standard medical care alone. In ASTRAL, there were substantial differences between individual nephrologists in the types of patients about whom they were uncertain. For example, most (but not all) were convinced that patients with severe bilateral renal artery stenosis should be revascularized, but there were differing views about the precise threshold of stenosis above which a patient should be offered such treatment and whether it should be used among those with stable kidney function. The use of the uncertainty principle enabled the trial to yield at least some direct evidence in a broad range of patients. As a result of the wide and simple entry criteria adopted by ASTRAL, 806 patients were randomized (which is almost four times more than all previous trials of renal revascularization combined), no patient received a treatment which they or their doctor believed to be contraindicated, and the study was able to provide some evidence about the

effects of renal revascularization with which to plan the treatment of future patients. Other trials in nephrology have also taught us the importance of embracing uncertainty. During the course of the placebo-controlled TREAT trial of erythropoietin, the investigators were initially criticized for choosing a placebo comparator, but as other data emerged during the trial they were then criticized for the haemoglobin target selected in the active arm. Eventually, the TREAT trial provided reliable data to show that a policy of increasing haemoglobin from 9 g/dL to about 13 g/dL does not reduce cardiovascular disease in patients with diabetes and CKD and may in fact cause harm (Pfeffer et al., 2009).

Efficient recruitment The chief focus of a large-scale trial designed to detect effects on major clinical outcomes should be to address its main hypothesis. In order to maximize the probability of a successful trial, it may be prudent to limit expenditure on measurements or substudies which are not strictly necessary. Such expenditure takes resources away from the primary purpose of the trial, and may be a poor investment if the trial fails to answer its primary question due to insufficient size. When designing a new study, it is wise to consider every proposed visit, measurement and assay critically. It is often possible to reduce costs without loss of scientific information. In SHARP, for example, it was not considered necessary to collect blood and urine for analysis in the central laboratory at every 6-month study visit, and it was possible to estimate changes in blood lipid profile among a random 10% subsample of patients at 1 year and 4 years after randomization (SHARP Collaborative Group, 2010). Moreover, knowledge of the results of some blood assays whilst the trial is in progress may not be necessary, and can be done more cheaply in batches at a later date in stored frozen samples.

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assessment of the patient with renal disease Renal artery stenosis detected

Should patient be offered immediate revascularization?

Doctor(s) or patient reasonably certain that revascularization is not appropriate: patient is ineligible

Doctor(s) or patient reasonably certain that revascularization is appropriate: patient is ineligible

Doctor(s) and patient substantially uncertain whether to revascularize: uncertainty implies eligibility

Group 1: allocated to medical therapy only

Group 2: allocated to medical therapy plus revascularization

6% get revascularization (median delay 20 months)

83% get revascularization (median delay 1 month)

Fig. 19.1  Demonstration of uncertainty principle in recruitment. The ASTRAL trial investigated renal artery revascularization in renal artery stenosis (Wheatley et al., 2009). Because individual clinicians differed in the types of patients about whom they were uncertain, no category of patient about which there was widespread uncertainty was excluded, so the trial result is widely applicable.

Pilot studies can be helpful in testing and refining processes and determining what data are essential to collect (and conversely what are unnecessary) (Box 19.2). They are also better suited to providing more detailed assessments of the treatments on intermediate variables such as blood pressure or cholesterol. It is feasible to study such measures in detail in a small number of closely followed patients and provide both accurate and reliable results, whereas such close observation is neither feasible nor necessary in a larger trial assessing clinical outcomes. One important aspect of the trial that can be piloted is the identification, invitation and recruitment of trial participants; time spent testing and modifying these in a pilot study can yield substantial rewards in terms of more rapid recruitment into the subsequent larger trial. One method of enhancing recruitment that can be very time- and cost-effective is pre-screening. For many renal trials this can be achieved by identifying potentially eligible patients from renal unit databases while other aspects of the study are still being set-up (e.g. before full ethics or regulatory approvals). Potentially eligible patients are asked if they would consider entering a trial so that there is a pool of patients who can be rapidly invited when the study can formally begin at a site. Pre-screening also acts as an assessment of a site’s performance and of whether eligibility criteria need to be modified.

Box 19.2  Information that can be obtained from pilot studies ◆ Recruitment: ease and rates of recruitment; test methods (e.g. pre-screening). ◆ Trial materials: assess acceptability of patient information leaflets and consent forms. ◆ Tolerability of study treatment:  will inform likely compliance in larger study which will have a major impact on statistical power. ◆ Efficacy on intermediate outcomes:  whilst assessment of effects on an intermediate outcome (e.g. cholesterol) may help in estimating likely effects on clinical outcomes for the main trial, in the absence of a suitable (i.e. causally relevant) intermediate outcomes effect size estimates in pilot studies will generally be of little value (since they are subject to large random errors). ◆ Trial procedures: identify trial procedures that patients or trial clinic staff find hard to adhere to, allowing their modification (or, perhaps even their removal if they are not absolutely necessary).

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Maintenance of compliance A high incidence of participants randomly allocated to active treatment stopping that treatment (‘drop-outs’) or those allocated to placebo starting open-label active treatment (‘drop-ins’) quickly erodes a trial’s ability to test the study hypothesis (that is, in statistical terms, it reduces study power): as much effort should be put into ensuring trial participants remain on their study treatment as goes into recruiting them in the first place. In general, every one treatment drop-out or drop-in post-randomization has the equivalent effect to recruiting two fewer patients in the first place. Such considerations were a key part of the design of SHARP, which included a pre-randomization 6-week placebo run-in period. Such a period allows participants, investigators, and other doctors time to reconsider a particular patient’s participation, enables tests to check eligibility to be conducted, and is a test of compliance so that those participants who were unlikely, or unable, to take study treatment long-term can be excluded prior to randomization (Lang et al., 1991). Similarly, the exclusion of patients with prior myocardial infarction or coronary revascularization was intended to reduce the chances of ‘drop-in’ statin therapy among such patients. Compliance can also be improved by minimizing the impact of the study on the patients’ daily routine and clinical care. Trials should also have procedures to identify participants who discontinue study treatment so any modifiable reasons can be addressed.

Reliable ascertainment of relevant study outcomes The reliable collection of study outcomes is ensured by careful study design. To prevent incomplete outcome ascertainment introducing bias, all participants should also continue to attend study visits even if they have stopped treatment or had a study outcome. Where feasible, a study may be ‘double blind’ so that both investigators and participants do not modify their reporting of events (consciously or subconsciously) by knowledge of treatment allocation. Alternatively, open-label studies can minimize these biases, providing that study endpoints are not subjective and that information is collected and assessed systematically (e.g. using registry data), blinded to the randomized treatment allocation. In order to facilitate complete follow-up, study visits should aim to collect the minimum information required from patients. Collecting vast amounts of data just because it might prove interesting both increases costs and makes patients less likely to attend visits, both of which work against a successful outcome. Using electronic case report forms is also an efficient method to check completeness (and internal validity) of data during the study clinic visit, rather than through time-consuming (and costly) checks of data later. To ensure high-quality follow-up, trial site staff should receive adequate training and support, and key aspects of study performance should be monitored (Landray et al., 2012). Traditionally, ‘on-site monitoring’, where a member of the trial team visits a study site, has been considered to be a standard requirement. However, the detection of missing data (or fraud) is more effectively—and much more efficiently—achieved by review of centrally collected data (Pogue et  al., 2013). In addition, statistical techniques can often be used to identify sites which are ‘outliers’ in measures of performance (e.g. study visit duration and frequency of adverse event reporting) and therefore require further scrutiny (perhaps including a targeted on-site visit) (Valdes-Marquez et al., 2011). It

clinical trials: why and how in nephrology

is important not to waste large amounts of resources trying to identify every item of data. Small amounts of missing or incorrect data are likely to be randomly distributed between arms and therefore, provided the study is large enough, unlikely to reduce the reliability of the results. Ultimately, the most important monitoring during a trial is that performed by an independent data monitoring committee, which regularly reviews unblinded interim analyses of the study’s emerging findings. These analyses are a more informative means of reviewing the safety of the study treatment than individual reports of adverse events.

Appropriate choice of study outcomes and statistical analyses Appropriate choice of study outcomes In any trial, it is important to select a primary outcome that is both clinically important and sensitive to the study treatment. Most ‘surrogate’ outcomes (e.g. albuminuria) are not true surrogates (Prentice, 1989) and trials based on these can be misleading: for example, although dual renin–angiotensin system blockade reduces albuminuria more than monotherapy (Jennings et  al., 2007), no trials have yet identified improvements in clinical outcomes (but larger trials have shown harm) with this strategy (Parving et al., 2012; Fried et  al., 2013). When combining endpoints, outcomes should be of broadly similar clinical significance and likely to be affected in the same direction. For example, trials of antithrombotic therapy need to consider ischaemic and haemorrhagic stroke subtypes separately for net benefit in different types of participant to be accurately estimated, whereas a trial of blood pressure-lowering treatment might reasonably combine them (since the epidemiological associations of blood pressure and each of the stroke subtypes are qualitatively similar). Total mortality is one of the least sensitive endpoints and is rarely an appropriate outcome because an effective treatment is likely to yield a moderate benefit on just a few causes of death, and as a result is likely to have only a small effect on total mortality. Any effect is also very dependent on the patterns of mortality in the population tested and the result may not be generalizable to a different population.

Appropriate statistical analysis Even after proper randomization of a large number of patients, bias can be introduced by inappropriate statistical analysis. For example, excluding patients who do not adhere to their study treatment will introduce bias if the prognosis of those excluded from one treatment group differs from that of those excluded from another (which is highly likely as the side-effects of active treatment are often more pronounced in those that are sicker). The Coronary Drug Project provides an example of this: patients who took at least 80% of their allocated clofibrate had lower mortality after 5 years than those who did not (15.0% versus 24.6% respectively, P = 0.0001). However, the difference was even more striking between patients who did or did not take at least 80% of their allocated placebo (15.1% versus 28.3%, P < 0.00001) (The Coronary Drug Project Research Group, 1980). It is therefore essential that the primary analysis of any trial should compare the outcome among all those originally allocated one treatment (regardless of how long they took the treatment for or even if they ever did) with the outcome among all those allocated to the comparator group (i.e. according to the ‘intention-to-treat’ principle).

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When faced with an overall trial result, doctors frequently want to know for which group of patients is this treatment particularly helpful (or harmful). Looking at the treatment effect in the subgroup of interest in isolation is not appropriate; the correct question is whether the treatment effect observed in that subgroup differs significantly from the overall treatment effect. In other words, the overall treatment effect becomes the new ‘null’ hypothesis and the relevant test is whether the subgroup of interest differs from this (using tests for heterogeneity or trend to test this hypothesis). In the absence of evidence of heterogeneity, the best resolution is to emphasize the overall results of the trial (Collins and MacMahon, 2001). It is often asked whether patients who respond better to a given treatment (e.g. have a larger than average blood pressure reduction with an antihypertensive) gain more benefit in terms of clinical outcomes. Not infrequently, these subgroups are defined after randomization and this can introduce bias. Groups of patients defined by different post-randomization responses are very unlikely to differ only randomly from each other and it is highly possible that some of these other differences might be related to the risk of the clinical outcome of interest. Inferences drawn from such non-randomized comparisons of ‘responders’ versus ‘non-responders’ are biased and could be misleading. For example, the RENAAL study of losartan versus placebo in diabetic nephropathy showed that allocation to losartan reduced the risk of death, dialysis or doubling of creatinine by 16% (P = 0.02) (Brenner et al., 2001). A subsequent analysis of the study suggested that patients who had a larger reduction in albuminuria derived the most renoprotective benefit (Eijkelkamp et al., 2007). However, the groups of patients were defined by post-randomization changes in albuminuria and therefore the analyses are potentially biased. Such analyses can be performed if response to treatment is defined prior to randomization. For example, in the Heart Protection Study, all potential participants entered a run-in phase with during which they received simvastatin 40 mg. Measurement of blood lipids at the beginning and end of run-in allowed for a pre-randomization assessment of low-density lipoprotein-lowering ‘responsiveness’. Therefore, post-randomization outcomes could be compared between active and control therapies within each response group without affecting the balance of randomization (Heart Protection Study Collaborative Group, 2002).

Summary If clinical outcomes for patients with kidney disease are to improve it is essential that nephrologists embrace the concept of large randomized trials, the concept of being uncertain, and join collaborative studies of the key clinical questions. Both randomization and large sample sizes are vital if the trial results are to be a reliable test of the intervention and if the results are to change clinical practice. Large trials require careful design and conduct to ensure that doctors can be confident that the answers they give are relevant to their practice. A doctor treating an individual patient may feel that the large trial results deny the individuality of their patient. However, one of the main reasons trials need to be so large is precisely because patients are so different from one another. Only by randomizing large numbers can the effect of treatment be distinguished from natural variation in clinical course. Trial populations may not be representative of all the patients doctors meet in routine clinical practice, but

an unrepresentative trial population can still provide a generalizable trial result. It is very unlikely that the patients recruited into a large randomized trial are so different from ‘normal’ patients that the proportional effect of treatment would differ in some meaningful way. From such data, the effect of treatment on ‘real-world’ observed absolute risk can be predicted. Large randomized trials therefore remain the only way to reliably answer questions about the efficacy and safety of many treatments in nephrological practice and are key to improving clinical outcomes.

References Baigent, C., Landray, M. J., Reith, C., et al. (2011). The effects of lowering LDL cholesterol with simvastatin plus ezetimibe in patients with chronic kidney disease (Study of Heart and Renal Protection): a randomised placebo-controlled trial. Lancet, 377, 2181–92. Bellomo, R., Cass, A., Cole, L., et al. (2009). Intensity of continuous renal-replacement therapy in critically ill patients. N Engl J Med, 361, 1627–38. Boaz, M., Smetana, S., Weinstein, T., et al. (2000). Secondary prevention with antioxidants of cardiovascular disease in endstage renal disease (SPACE): randomised placebo-controlled trial. Lancet, 356, 1213–8. Brenner, B. M., Cooper, M. E., de Zeeuw, D., et al. (2001). Effects of losartan on renal and cardiovascular outcomes in patients with type 2 diabetes and nephropathy. N Engl J Med, 345, 861–9. Chertow, G. M., Block, G. A., Correa-Rotter, R., et al. (2012). Effect of cinacalcet on cardiovascular disease in patients undergoing dialysis. N Engl J Med, 367, 2482–94. Cohn, J. N., Goldstein, S. O., Greenberg, B. H., et al. (1998). A dose-dependent increase in mortality with vesnarinone among patients with severe heart failure. Vesnarinone Trial Investigators. N Engl J Med, 339, 1810–6. Collins, R. and MacMahon, S. (2001). Reliable assessment of the effects of treatment on mortality and major morbidity, I: clinical trials. Lancet, 357, 373–80. Deeks, J. J., Dinnes, J., D’amico, R., et al. (2003). Evaluating non-randomised intervention studies. Health Technol Assess, 7, iii–x, 1–173. Eijkelkamp, W. B., Zhang, Z., Remuzzi, G., et al. (2007). Albuminuria is a target for renoprotective therapy independent from blood pressure in patients with type 2 diabetic nephropathy: post hoc analysis from the Reduction of Endpoints in NIDDM with the Angiotensin II Antagonist Losartan (RENAAL) trial. J Am Soc Nephrol, 18, 1540–6. Feldman, A. M., Bristow, M. R., Parmley, W. W., et al. (1993). Effects of vesnarinone on morbidity and mortality in patients with heart failure. Vesnarinone Study Group. N Engl J Med, 329, 149–55. Fried, L. F., Emanuele, N., Zhang, J. H., et al. (2013). Combined angiotensin inhibition for the treatment of diabetic nephropathy. N Engl J Med, 369(20), 1892–903. Grady, D., Rubin, S. M., Petitti, D. B., et al. (1992). Hormone therapy to prevent disease and prolong life in postmenopausal women. Ann Intern Med, 117, 1016–37. Heart Protection Study Collaborative Group (2002). MRC/BHF Heart Protection Study of cholesterol lowering with simvastatin in 20,536 high-risk individuals: a randomised placebo-controlled trial. Lancet, 360, 7–22. Jafar, T. H., Schmid, C. H., Landa, M., et al. (2001). Angiotensin-converting enzyme inhibitors and progression of nondiabetic renal disease. A meta-analysis of patient-level data. Ann Intern Med, 135, 73–87. Jennings, D. L., Kalus, J. S., Coleman, C. I., et al. (2007). Combination therapy with an ACE inhibitor and an angiotensin receptor blocker for diabetic nephropathy: a meta-analysis. Diabet Med, 24, 486–93. Landray, M. J., Grandinetti, C., Kramer, J. M., et al. (2012). Clinical trials: rethinking how we ensure quality. Drug Information Journal, 46, 657–60. Lang, J. M., Buring, J. E., Rosner, B., et al. (1991). Estimating the effect of the run-in on the power of the Physicians’ Health Study. Stat Med, 10, 1585–93.

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Palevsky, P. M., Zhang, J. H., O’Connor, T. Z., et al. (2008). Intensity of renal support in critically ill patients with acute kidney injury. N Engl J Med, 359, 7–20. Parving, H. H., Brenner, B. M., McMurray, J. J., et al. (2012). Cardiorenal end points in a trial of aliskiren for type 2 diabetes. N Engl J Med, 367, 2204–13. Pfeffer, M. A., Burdmann, E. A., Chen, C.-Y., et al. (2009). A trial of darbepoetin alfa in type 2 diabetes and chronic kidney disease. N Engl J Med, 361(21), 2019-32. Pogue, J. M., Devereaux, P. J., Thorlund, K., et al. (2013). Central statistical monitoring: detecting fraud in clinical trials. Clin Trials, 10, 225–35. Prentice, R. L. (1989). Surrogate endpoints in clinical trials: definition and operational criteria. Stat Med, 8, 431–40. Ronco, C., Bellomo, R., Homel, P., et al. (2000). Effects of different doses in continuous veno-venous haemofiltration on outcomes of acute renal failure: a prospective randomised trial. Lancet, 356, 26–30. Rossouw, J. E., Anderson, G. L., Prentice, R. L., et al. (2002). Risks and benefits of estrogen plus progestin in healthy postmenopausal women: principal results from the Women’s Health Initiative randomized controlled trial. JAMA, 288, 321–33. Schiffl, H., Lang, S. M., and Fischer, R. (2002). Daily hemodialysis and the outcome of acute renal failure. N Engl J Med, 346, 305–10.

clinical trials: why and how in nephrology

SHARP Collaborative Group (2010). Study of Heart and Renal Protection (SHARP): randomized trial to assess the effects of lowering low-density lipoprotein cholesterol among 9,438 patients with chronic kidney disease. Am Heart J, 160, 785–794 e10. Strippoli, G. F., Craig, J. C., and Schena, F. P. (2004). The number, quality, and coverage of randomized controlled trials in nephrology. J Am Soc Nephrol, 15, 411–9. Tepel, M., Van Der Giet, M., Statz, M., et al. (2003). The antioxidant acetylcysteine reduces cardiovascular events in patients with end-stage renal failure: a randomized, controlled trial. Circulation, 107, 992–5. The Coronary Drug Project Research Group (1980). Influence of adherence to treatment and response of cholesterol on mortality in the coronary drug project. N Engl J Med, 303, 1038–41. Valdes-Marquez, E., Hopewell, J. C., Landray, M., et al. (2011). A key risk indicator approach to central statistical monitoring in multicentre clinical trials: method development in the context of an ongoing large-scale randomized trial. Trials, 12, A135. Webster, A. C., Playford, E. G., Higgins, G., et al. (2004). Interleukin 2 receptor antagonists for renal transplant recipients: a meta-analysis of randomized trials. Transplantation, 77, 166–76. Wheatley, K., Ives, N., Gray, R., et al. (2009). Revascularization versus medical therapy for renal-artery stenosis. N Engl J Med, 361, 1953–62.

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SECTION 2

The Patient with fluid, electrolyte, and renal tubular disorders

20 An overview of tubular function  171 Matthew A. Bailey

21 Sodium transport and balance: a key role for the distal nephron  189 Laurent Schild

22 Water homeostasis  197 David Marples and Søren Nielsen

23 Potassium homeostasis  204 Alain Doucet and Gilles Crambert

24 Renal acid–base homeostasis  218 Carsten A. Wagner and Olivier Devuyst

25 Phosphate homeostasis  225 Heini Murer, Jürg Biber, and Carsten A. Wagner

26 Calcium homeostasis  231 Francesco Trepiccione and Giovambattista Capasso

27 Magnesium homeostasis  243 Pascal Houillier

28 Approach to the patient with hyponatraemia  249 Ewout J. Hoorn and Robert Zietse

29 Approach to the patient with hypernatraemia  261 Robert Zietse

30 Approach to the patient with oedema  272 Jean-Marie Krzesinski and Eric P. Cohen

31 Approach to the patient with salt-wasting tubulopathies  282 Detlef Bockenhauer and Robert Kleta

32 Approach to the patient with polyuria  291 Daniel G. Bichet

33 Clinical use of diuretics  299 David H. Ellison and Arohan Subramanya

34 Approach to the patient with hypo-/hyperkalaemia  323 Charles S. Wingo and I. David Weiner

35 Approach to the patient with metabolic acidosis or alkalosis  339 Mitchell L. Halperin and Kamel S. Kamel

36 Approach to the patient with renal tubular acidosis  363 Stephen B. Walsh

37 Approach to the patient with hypercalcaemia  372 Dennis Joseph and Theresa A. Guise

170

38 Approach to the patient with hypocalcaemia  378 Agnès Linglart and Anne-Sophie Lambert

39 Approach to the patient with hypo-/ hyperphosphataemia  384 Judith Blaine, Hector Giral, Sabina Jelen, and Moshe Levi

40 Approach to the patient with hypomagnesaemia  397 Martin Konrad and Karl P. Schlingmann

41 Approach to the patient with renal Fanconi syndrome, glycosuria, or aminoaciduria  412 Detlef Bockenhauer and Robert Kleta

CHAPTER 20

An overview of tubular function Matthew A. Bailey Introduction

The proximal tubule

Of the approximately 180 L of water that is filtered by the renal glomeruli per day, only 1–2 L is excreted. Similarly, the filtered sodium (Na+) load of approximately 25,000 mmoL/day is mostly reabsorbed with only approximately 150 mmol/day being excreted. Chloride (Cl−), bicarbonate (HCO3−), calcium (Ca2+), and magnesium (Mg+) have a proportionally similar excretory profile and although secretion is important for the homeostasis of some electrolytes (particularly potassium and acid–base balance), the metabolic activity of the kidney is chiefly concerned with reabsorption. Most of the transport processes are coupled to the reabsorption of sodium, either directly through specialized transport proteins, or indirectly due to transepithelial electrochemical gradients. The ‘pump-leak’ model is evident in most nephron segments: the heteromeric Na+/K+-ATPase extrudes three sodium ions across the basolateral membrane in exchange for two potassium ions, generating a low intracellular sodium concentration and a steep electrochemical gradient for passive entry across the apical membrane via a selection of specialized transport proteins (Fig. 20.1). This process accounts for most of the energy used by the kidney, which is derived chiefly through oxidative metabolism. Renal tissue, which accounts for approximately 0.5% of total body mass, consumes approximately 7% of whole-body oxygen. Renal blood flow is about 20% of the cardiac output and the renal cortex is maintained at the high partial pressure of oxygen required for aerobic metabolism. The proximal tubule is the powerhouse for reabsorptive processes. It reclaims all of the filtered glucose, amino acids, and small proteins, 80% of HCO3− and phosphate (PO4), and 70% of Na+, Cl−, potassium (K+), and water. The loop of Henle contributes a further 25% to overall reabsorption of Na+, Cl−, and K+ with significant amounts of Ca2+ and Mg+ also being reabsorbed, particularly in the thick ascending limb of Henle (TALH). The distal tubule and collecting duct can fine-tune overall urinary excretion in accordance with systemic balance requirements: Na+, Cl− Ca2+, and Mg2+ are all reabsorbed here, whereas the net movement of K+ is secretory. Water reabsorption in the distal nephron is variable, being influenced by vasopressin and thus hydration, and to a lesser extent, volume status. In the first part of this chapter, the major transport pathways and regulatory features for each nephron segment is described. The focus here is on the transepithelial flux of Na+,K+, and water. In the second part, other important aspects of renal homeostasis, including urine concentration and acid–base balance, are summarized. Throughout the chapter, the key transport proteins are given and, where different, the name of the encoding gene is also provided.

The main function of the proximal tubule is the bulk reabsorption of the glomerular filtrate. The majority of this occurs in the proximal convoluted tubule. Two structural features of the proximal tubule cells enhance the surface area for transepithelial flux. First, microvilli project into the tubule lumen from the apical membrane (the brush border membrane); and second, the basolateral membrane is extensively invaginated. Transport in the proximal tubule is driven by the Na+/K+-ATPase, expressed in the basolateral membrane. Hydrolysis of ATP by this transporter underpins the high metabolic requirements of the proximal tubule, which has mitochondria densely packed to the basolateral membrane. The proximal tubule lies mainly in the cortex and relies predominantly on aerobic metabolism. This contrasts with those segments in the medulla that are in a relatively hypoxic environment and therefore have capacity for anaerobic metabolism. The proximal tubule is prone to hypoxic insult and mitochondrial dysfunction, which may reflect intrinsic differences in the mitochondrial population, compared to the distal tubule (Hall et al., 2009). In the proximal tubule, the NHE3 isoform of Na+/H+ exchange is the main transporter responsible for Na+ entry into the cells but a whole battery of specialized transporters are also expressed in this membrane, which couple sodium entry to movement of other species (Fig. 20.2A). Despite the prominence of NHE3, mutations in the encoding gene (SLC9A3) have not yet been associated with human renal disease. This reflects the redundancy of renal sodium transport:  NHE3 knockout mice have increased distal sodium reabsorption (Bailey et al., 2004) due to activation of the renin–angiotensin–aldosterone system (RAAS); the mild hypotension and alkalosis reflect an absorptive defect in the colon. Aquaporin-1 (AQP1) water channels are constitutively expressed in both membranes, and contribute to the high water hydraulic permeability of the proximal segment. Deletion of AQP1 in mice diminishes substantially transcellular flux and this channel is required for near isosmolar reabsorption observed in the proximal tubule (Vallon et al., 2000). However, water transport in the proximal tubule as a whole remains robust and the low-resistance paracellular shunt provides a route, perhaps the major route, of transport for water across the proximal tubule. Experiments in rodents suggest that the tubular fluid is relatively hypotonic, the transepithelial osmotic gradient of < 10 mosmol/kg being sufficient to drive large amounts of water reabsorption (Green and Giebisch, 1989). It is possible, however, that reabsorption can occur in the absence of an osmolar gradient due to ‘electro-osmosis’ or current-induced fluid movement (Fischbarg, 2010).

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Lumen

Interstitial fluid

Na+ Na+ Na+

K+ K+

X Na+ Y

Apical membrane

Basolateral membrane

Fig. 20.1  Prototypical renal tubule cell. The Na+/K+-ATPase or ‘sodium pump’ in the basolateral membrane maintains a low intracellular Na+ concentration, generating an electrochemical gradient for passive entry of Na+ across the apical membrane. The transport protein facilitating movement of Na+ across the apical membrane varies along the nephron. Cotransport (symport and antiport) systems use the kinetic energy of Na+ entry to move another species against its electrochemical gradient. A simple system—an ion channel for Na+—is found in the distal nephron.

The apical membrane of the proximal tubule also expresses a range of enzymes that change the composition of luminal fluid. Carbonic anhydrase type IV, for example, is vital to bicarbonate reabsorption (see later) and enzymes such as neutral endopeptidase (Walter et  al., 1997)  and ecto-5′-nucleotidase can influence the tubular concentration of paracrine agents (Shirley et al., 2009)  and thereby influence tubular function in downstream segments.

Transport processes The initial short segment of the proximal tubule (S1 segment) reclaims most of the filtered glucose, amino acids, and phosphate by cotransporters directly coupled to sodium entry into the cell down its electrochemical gradient (Fig. 20.2A). Glucose crosses the apical membrane via the low-affinity, high-capacity, sodium-dependent transporter SGLT2 (SLC5A2), and exits via GLUT2. The reabsorption of glucose (and amino acids) is electrogenic, generating a small, lumen negative, transepithelial potential difference (PD) (approximately −2mV) in the S1 proximal tubule. Although this should favour the paracellular reabsorption of Cl−, the reflection coefficient of the S1 PT for Cl− is 0.9 (a reflection coefficient of 1.0 indicates complete impermeability of the S1 epithelium to the solute in question). Cl− reabsorption therefore lags behind that of Na+ and water, causing the tubular fluid Cl− concentration to rise slightly. Thus, Na+ reabsorption in the S1 PT is preferentially ‘coupled’ to that of HCO3−. This process is indirect: sodium entry into the cell via the NHE3 exchanger causes secretion of a proton into the tubule lumen. An apical H+-ATPase also contributes to proton secretion. In the tubule fluid, protons combine with the filtered HCO3− to produce carbonic acid and ultimately carbon dioxide (CO2) and water. This is discussed in more detail in the later section on HCO3−. The remainder of the proximal convoluted tubule is the S2 segment. In the cortex, the proximal straight tubule also consists of S2 cells, with S3 cells being limited to the medulla. The preferential reabsorption in the S1 segment of NaHCO3 relative to NaCl creates a modest tubular-fluid to plasma concentration gradient for

Cl−. The gap junctions between S2 cells have a high permeability for Cl−, allowing paracellular reabsorption down its concentration gradient. Since electrogenic Na+ reabsorption in the S2 segment is now limited by the low tubular fluid concentration of glucose and amino acids, paracellular Cl− flux causes a reversal of the transepithelial PD (now +2 mV). This lumen positivity favours paracellular reabsorption of cations (Fig. 20.2B) but the majority of sodium transport is thought to be transcellular via apical NHE3. The gradient also favours anion secretion and there is substantial plasma to tubule fluid concentration gradient for HCO3−. However, this gradient is not dissipated due to low paracellular permeability for HCO3− backflux. Micropuncture evidence suggests that the S1 and S2 proximal tubule reabsorbs approximately 70% of the filtered Cl−. Not all of this occurs by paracellular diffusion as some transporters, coupled ultimately to the basolateral Na+/K+-ATPase, help to reabsorb Cl− by the transcellular route (Fig. 20.3). The apical entry path is either secondary or tertiary active transport involving members of Slc26 anion exchanger family (Sindic et al., 2007). These transporters are multifunctional, capable of transporting, in addition to Cl−, anions such as sulphate, iodide, formate, oxalate and HCO3−. CFEX (SLC26A6) has an important role here. Expressed on the apical membrane, CFEX is capable in expression systems of chloride/formate, chloride/oxalate and chloride/bicarbonate exchange (Aronson, 2006). Studies in Slc26a6 null mice indicate that the primary mode in vivo is to mediate oxalate-dependent Cl− reabsorption in the proximal tubule. The chloride exits via a basolateral K+Cl− cotransport mechanism (probably KCC1) or through Cl− channels. K+ reabsorption pathways in the proximal convoluted tubule are not completely resolved. Much of the potassium that enters the cell via the Na+/K+-ATPase is recycled across the basolateral membrane via K+ channels (channels of KCNK and KCNJ gene families have been identified) or K+Cl− cotransport. The isoforms KCC3 (SLC12A6) and KCC4 (SLC12A7) have been identified (Jentsch, 2005). These basolateral exit routes for K+ are crucial for maintaining efficient transepithelial Na+ flux, stabilizing intracellular K+ concentration (and cell volume) in the face of fluctuating rates of transepithelial Na+ transport (Hamilton and Devor, 2012). K+ channels have also been identified in the apical membrane but the electrochemical gradient would favour secretion of K+ into the tubular fluid. A small secretory component has been identified but the physiological purpose of this is unknown. There is some evidence for active transport mechanisms, particularly the non-gastric isoform of H+/K+-ATPase (ATP12A), but the contribution that this pathway makes under normal circumstances is probably minor (Malnic et al., 2013). K+ reabsorption lags behind that of Na+ in the S1 segment, causing the tubular fluid concentration to rise slightly. Combined with the lumen positive PD in the S2 segment, paracellular reabsorption by simple diffusion is sufficient to account for the majority of K+ reabsorption by the proximal tubule. A certain proportion of K+ reabsorption is mediated by paracellular solvent drag. Since the S2 segment has a high K+ permeability (reflection coefficient of < 0.4), significant amounts of this solute can be entrained by vectoral fluid flux. The proximal straight tubule, or pars recta, has similar mechanisms of transport for the major electrolytes as the PCT and reabsorption here is also effectively isosmolar. Notably, the proximal straight tubule expresses in addition to AQP1, AQP7 (Sohara et al.,

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an overview of tubular function

(A)

S1 Interstitial fluid

Lumen Na+ glucose Na+ amino acids Na+ NPT2a phosphate Na+ NHE3 H+ SGLT2

Na+ K+ K+ K+ Cl–

–2mV

H2O AQP1 AQP1 (B) Lumen

S2

SGLT1

Interstitial fluid

Cl–

+

Na+ NHE3

Na+

K+

H+ K+

Na+ K+

K+ Cl–

H2O AQP1 AQP1

Fig. 20.2  The major Na+ transport pathways in the proximal tubule. (A) In the S1 segment, Na+ transport is coupled to that of glucose, amino acids, and bicarbonate, generating a small lumen negative transepithelial potential difference. (B) In the S2 segment, this potential becomes lumen positive..

2009), which contributes both to transepithelial water flux and the generation of a concentrated medulla due to its permeability to glycerol. Studies in isolated tubule segments have identified a lower Na+/K+-ATPase activity per unit length and the capacity for Na+ transport is approximately 50% that of the PCT. The straight segment reclaims (‘mops up’) the remaining 10% of the filtered glucose load with the high-affinity, low-capacity Na+-dependent cotransporter, SGLT1 (SLC5A1), providing entry across the apical membrane; GLUT1 facilitates basolateral exit. A major difference in the proximal straight tubule is that net K+ flux is secretory, which may reflect the diminution (or reversal) of the concentration gradient for paracellular K+ diffusion coupled to transcellular secretion via apical K+ channels. The PST is also an important site for the secretion of organic acids and bases, via multiple members of the SLC22A gene family (Fig. 20.4). For organic acids, the first step of the secretory process is basolateral anion exchange, in which

the exit of dicarboxylate (chiefly α-ketoglutarate) down a concentration gradient is coupled to entry of organic acids (Rizwan and Burckhardt, 2007). The main transporters are OAT1 and OAT3. Apical efflux utilizes a range of transporters, including OAT4 (humans only) and URAT-1, and is not rate limiting. For bases, the overall pathway is similar, with the basolateral entry step being rate limiting and mediated by the selective exchangers, OCT1 and OCT2 (Jonker and Schinkel, 2004). Clinically important organic acids (e.g. non-steroidal anti-inflammatory drugs, diuretics, penicillin) and bases (e.g. amiloride, cimetidine and atropine) are substrates for this secretory system, which is often, therefore, defined from a pharmacological viewpoint. Indeed, the prototypic substrate for organic acid secretion is p-aminohippurate, this being the central tenet for its clearance being used as an index of effective renal plasma flow. Creatinine is secreted via the organic base pathway, which can lead

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S2

Lumen

NHE3 H+

Na+

Na+

H+

K+ K+ Cl–

HCOO–

HCOO– CFEX

Interstitial fluid

K+ Cl–

Cl–

Fig. 20.3  Tertiary active transport facilitating transepithelial Cl− reabsorption.

to an overestimation of creatinine clearance and thus glomerular filtration rate (GFR). The physiological role of this system is less well characterized. URAT1 contributes to the urate homeostasis. Urate is the end product of purine metabolism in humans (rodents express uricase allowing metabolism of urate to allantoin) and hyperuricaemia is a risk factor for chronic kidney and cardiovascular disease (Filiopoulos et al., 2012). URAT1 reabsorbs urate in exchange for lactate or nicotinate and is therefore an attractive therapeutic target in patients with hyperuricaemia/gout. Corticosteroids have affinity for the OAT proteins and dopamine and adrenalin are transported via the organic base route. These systems may therefore contribute to intrarenal recycling of physiological active hormones. Gene deletion studies in mice also indicate that OAT3 may contribute to blood pressure control (Vallon et al., 2008), suggesting a role in Na+ and fluid homeostasis.

Major control mechanisms Autoregulation of blood flow is an intrinsic property of the vasculature that stabilizes renal perfusion in the face of fluctuating

S3 Lumen OAT4

Cl–

URAT1

OA–

Interstitial fluid α-KG2–

OAT1

–

OAT3

OA

Na+ K+ H+ OB+

OCT1 OB+

OCT3

Fig. 20.4  The S3 proximal tubule is a major site for the secretion of organic acids (OA) and bases (OB), which include several classes of clinically important drugs. The physiological role of this system is less well defined.

blood pressure. Autoregulation is so efficient that renal blood flow can be largely independent of blood pressure in the physiological range (Cupples, 2007). Whole-kidney autoregulation is governed through the combined influence of at least two mechanisms: tubuloglomerular feedback (see ‘The macula densa’) and the intrinsic myogenic response of the vascular smooth muscle. These regulatory systems have different, but overlapping, operational frequencies. Of the two major components, only the intrinsic myogenic response is both necessary and sufficient for full whole-kidney autoregulation (Cupples, 2007). The myogenic response operates along the preglomerular vascular tree, responding to increased transmural pressure by channel-mediated calcium influx and reflex vasoconstriction of the vascular smooth muscle. The exact signalling mechanisms are not defined, but local release of ATP is implicated, causing vasoconstriction through activation of P2X1 channels (Shirley et al., 2013). Glomerulotubular balance (GTB) is a mechanism intrinsic to the haemodynamic and structural properties of the proximal tubule. It ensures that fluctuations in GFR, and therefore filtered solute load, are matched by near proportionate changes in proximal tubular reabsorption such that the fractional reabsorption of the proximal tubule is held almost constant. This serves to prevent loss of solute in the event of increased GFR; as GFR drops it maintains delivery of sodium to the distal nephron, permitting efficient regulation of potassium and proton secretion. Mechanistically, the balance of peritubular Starling forces is a major component of GTB. Thus, when filtration rate rises due to an increased filtration fraction, the oncotic pressure in the peritubular capillaries is elevated, stimulating reabsorption. When GFR rises without a change in filtration fraction (influenced by glomerular plasma flow), flow-dependent reabsorption may contribute to GTB. The underlying mechanism of flow-dependence couples mechanical forces exerted on the microvilli to altered intracellular calcium signalling (Weinbaum et  al., 2010)  and modulation of paracrine agents, such as ATP (Shirley et al., 2013), dopamine (Du et al., 2012b) or angiotensin II (Du et al., 2012a). Pressure natriuresis is the process through which increases in arterial blood pressure lead to an increase in renal sodium excretion. This process has infinite feedback gain, that is, it continues to

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function until blood pressure returns to set point. Pressure natriuresis is central to the integrated maintenance of sodium homeostasis and thus the long-term regulation of blood pressure (Wadei and Textor, 2012). When kidney function is normal, sustained elevation of arterial pressure of only a few mmHg can evoke large changes in sodium excretion. An impaired pressure natriuresis response is a hallmark of hypertensive states or conditions of sodium sensitivity of blood pressure (Ivy and Bailey, 2014) (Fig. 20.5). Pressure natriuresis is ascribed to a diminution of sodium reabsorption in the proximal tubule following an acute increase in blood pressure. The main mechanism is an increase in renal interstitial hydrostatic pressure, particularly in the medulla. Decapsulation of the kidney prevents the rise in interstitial pressure and blunts the natriuresis, indicating that the response is largely determined by physicochemical factors. Pressure natriuresis occurs in the absence of large changes in renal blood flow due to autoregulation. This at first seems counterintuitive, since the increase in systemic arterial pressure must be transduced to the kidney by changes in renal perfusion pressure. One possibility is that medullary vasculature autoregulates less well than the cortex and small increases in whole-kidney blood flow exert large changes in the medulla. Nevertheless, factors that directly affect tubular sodium transport, particularly those that determine nitric oxide bioavailability are important (Menzies et al., 2015). Renal sympathetic nerve activity (RNSA) can directly stimulate proximal tubule sodium transport by increasing NHE3 and Na+/ K+-ATPase activity. RSNA is increased following a reduction in circulating volume or a modest (< 5 mmol/L) elevation of plasma sodium, such as can occur following a meal. RNSA thus contributes to an integrated view of blood pressure control (Kopp, 2011). Renal sympathetic overactivity has long been linked to sodium retention in experimental hypertension and recent clinical data indicate that bilateral sympathetic efferent denervation effects sustained reductions in blood pressure in selected hypertensive patients (Esler et al., 2010). Plasma angiotensin II, within the physiological range stimulates sodium transport by activation of NHE3; pharmacological levels

Salt excretion

Sodium intake (× Normal)

8 6

B High salt intake

4 A

2

C

50

100

150

Normal salt intake 200

MABP (mmHg)

Fig. 20.5  Renal function curve showing the relationship between arterial blood pressure (MABP) and renal sodium excretion. (A) The equilibrium pressure that is maintained through adjustment in sodium balance. (B) On sustained increases in salt intake, the function curve shifts to the left to give a higher level of excretion at any given pressure. (C) If natriuretic capacity is impaired, the curve shifts to the right and is flattened, so that a higher equilibrium pressure is required to match sodium output to input.

an overview of tubular function

are inhibitory. These effects are mediated by AT1 receptors on the basolateral membrane. Glucocorticoids, acting via the glucocorticoid receptor, stimulate the reabsorption of sodium bicarbonate due to activation of NHE3 and the Na+HCO3− cotransporter on the basolateral membrane. From an integrated standpoint, however, glucocorticoids promote natriuresis, kaliuresis and phosphaturia. Some of this reflects inhibition of transporters but the majority reflects the haemodynamic effects of increased GFR (Hunter et al., 2014). Intraluminal factors produced by proximal tubule cells will regulate transport by autocrine/paracrine signalling. The concentration of angiotensin II in tubular fluid is much higher than that in plasma due to the intrarenal RAAS (Ramkumar and Kohan, 2013). Angiotensin II can either stimulate (via AT1R) or inhibit (via AT2R) proximal transport by regulating NHE3 activity. Similarly, the proximal tubule has a local dopaminergic system (Zhang et al., 2012); activation of D1 receptor contributes to the natriuretic effect of dopamine. Finally, intraluminal nucleotides inhibit NHE3 activity via P2Y1 receptors (Shirley et al., 2013).

The loop of Henle The loop of Henle comprises the proximal straight tubule (see above), the thin descending limb, the thin ascending limb, and the cortical and medullary segments of the TALH. The thin descending limb has a high expression of AQP1 in both apical and basolateral membranes and has, therefore, a large hydraulic permeability. The permeability to sodium varies among species and transepithelial transport processes are not defined. In contrast to the proximal tubule, the thin limb is simple epithelial of flat cells with short microvilli and few mitochondria. Consistent with this, only a low level of Na+/K+-ATPase activity has been detected and few other transport proteins detected:  there is evidence for Cl− and K+ channels and K+Cl− cotransport and a sodium-borate (or potentially Na+HCO3− transporter), encoded by SLC4A11, has been identified (Groger et al., 2010). UT-A2 (SLC14A2) transporters have been identified in the thin descending limb, giving this segment a high permeability to urea. The transition from thin descending limb to thin ascending limb is evident only in the deep nephron population and occurs just before the bend of the loop. The thin ascending limb is a simple epithelium, having low water permeability due to absence of any aquaporin. Although Na+/K+-ATPase activity is low, the epithelial permeability to sodium, potassium and chloride is high. A  bumetanide-sensitive pathway, presumably a Na+-K+-2Cl− cotransporter has been identified in several species (Nishino et  al., 2007)  and mRNA for NHE1, NHE3, H+-ATPase and H+/ K+-ATPase is expressed (Pannabecker et al., 2002). In addition, Cl− channels are identified in both apical and basolateral membrane (Liu et al., 2002). The thin descending limb has significant permeability to urea, allowing rapid equilibrium of the tubular fluid and medullary interstitium.

Thick ascending limb of Henle The TALH is a major site of Na+Cl− reabsorption, accounting for approximately 20% of the filtered load. Cells of the TALH contain a high number of mitochondria, reflecting the significant activity of Na+/K+-ATPase. Sodium crosses the apical membrane of TALH

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Lumen

TALH

Interstitial fluid

ROMK K+

Na+ K+ 2Cl–

NKCC-2

K+

Na+

Na+

NHE2 NHE3 +

K+

H+ Na+ K+ Ca2+ Mg2+

Cl–

ClC-Kb

Cl– Barttin

ClC-Ka

K+ Cl–

KCC1

Fig. 20.6  Major transport pathways in the thick ascending limb of Henle.

cells by several routes (Fig. 20.6). The NHE3 and NHE2 exchangers but their contribution to transcellular Na+ flux is marginal (Shirley et al., 1998). More recently mRNA for a sodium-activated sodium channel has been found in the TALH. Expression is regulated by dietary sodium content and although it is hypothesized that this may be the elusive renal sodium sensor, its physiological function is unclear (Lara et al., 2012). The main route of entry is via NKCC2 (Fig. 20.6). This protein is inhibited by loop diuretics, which compete with Cl− at one of its binding sites. NKCC2 is encoded by a single gene, SLC12A1, mutations in which cause the salt-wasting Bartter syndrome type 1 (Seyberth and Schlingmann, 2011) (Fig. 20.7). Alternative splicing of SLC12A1 produces variants of NKCC2, which differ in their ion affinity and in their distribution along the TALH. Physiologically, the distribution is such that the affinity matches the concentration

of the transported ions in the tubular fluid, thereby permitting maximum rates of Na+Cl− reabsorption (Ares et al., 2011). Chloride exits the basolateral membrane via K+Cl− cotransport or through a heteromeric chloride channel assembled from CLC-Kb and Barttin (BSDN). Mutations in these subunits cause Bartter’s type 3 and 4, respectively (Fig. 20.7). Most of the potassium that enters the cell via NKCC2 diffuses back across the apical membrane through apical K+ channels ROMK/ KCNJ1, driven by the electrochemical gradient. This recycling of K+ has two consequences. First it ensures the continued availability of K+ to the cotransporter. Na+ and K+ must bind in a 1:1 stochiometry and the K+ concentration in the fluid delivered into the TALH is an order of magnitude lower than that of Na+. Second, it confers electrogenicity on a transporter that is intrinsically electroneutral. The charge separation caused by movement of K+ back into the tubule

Lumen

TALH

Interstitial fluid

Na+ Type 2

K+

K+ Type 1

K+ 2Cl– Na+

Na+ K+ Ca2+ Mg2+

K+ Cl–

Type 3

Cl– Type 4

Fig. 20.7  Mutations causing Bartter syndrome impair the transcellular reabsorption of Na+Cl−, leading to a diminution of the gradient for paracellular reabsorption of cations.

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fluid and Cl− out across the basolateral membrane generates a trans­ epithelial PD that is lumen positive (~ +10 mV). Theoretically, the transepithelial PD can drive paracellular reabsorption of cations or secretion of anions. The tight junctions of the TALH do have a high ionic conductance (and a very low hydraulic conductance) but are selective for cations. Approximately 50% of the Na+ reabsorbed in the TALH is via this voltage-driven paracellular shunt. This also provides a major route for the reabsorption of Ca2+ and Mg2+. The TALH reabsorbs approximately 25% of the filtered K+ load. Despite a significant K+ conductance in both apical and basolateral membrane, the majority of this reabsorption is paracellular and under conditions of high Na+ reabsorption, the net transcellular K+ flux can be secretory. Nevertheless the apical K+ channels are a vital for the efficient functioning of the TALH. Patch-clamp studies have identified two K+ channels in this membrane. The smaller conductance channel has the electrophysiological ‘fingerprint’ of the cloned ROMK channel, encoded by KCNJ1. Mutations in this gene cause Bartter syndrome type 2 and small molecule inhibitors of ROMK are in development as novel diuretics (Bhave et al., 2011). Although the molecular identity of the second channel is unknown, it is not expressed in the TALH of KCNJ1 knockout mice (Lu et al., 2004), suggesting that it is formed as a heteromeric complex in which ROMK is essential for physiological function.

The macula densa The macula densa, a small plaque of approximately 20 cells found at most distal segment of the cTALH, is the senor unit of the juxtaglomerular apparatus and influences the secretion of renin from granular cells of the afferent arteriole (Kurtz, 2011). Macula densa cells also exert a countervailing influence on GFR in the glomerulus of origin (and perhaps also in neighbouring nephrons) via vasoactive effects on the afferent arteriole and mesangial cells. This process, called tubule-glomerular feedback, serves to stabilize the moment-to-moment delivery of NaCl into the distal nephron and thereby optimize the fine-tuning of Na+ (and K+/H+) homeostasis by the kidney: aberrant tubuloglomerular feedback has been implicated in the progression of diseases such as diabetic nephropathy (Vallon and Thomson, 2012). Macula densa cells express the high-affinity isoform of NKCC2 in the apical membrane. The exit pathways for Na+ are unclear and intracellular Na+ concentration is not well buffered compared to other renal epithelial cells, rising and falling along with apical entry. As Cl− exits through channels, the membrane becomes depolarized, activating a maxi-anion channel through which ATP is released. The release of ATP, directly proportional to Na+ delivery to the apical membrane, is the critical effector of tubule-glomerular feedback, promoting vasoconstriction of the afferent arteriole either directly or following metabolism to adenosine (Bell et al., 2009). The TALH is also the site for the production and secretion of Tamm–Horsfall protein (aka uromodulin). This glycoprotein is the most abundant protein in normal urine, with up to 150 mg/ day being excreted. The physiological roles are poorly understood. Mutations in the encoding gene, UMOD, cause a cluster of rare autosomal dominant kidney diseases, characterized by progressive tubule-interstitial damage, hyperuricaemia, urinary concentrating defects, cyst formation, and progressive renal failure. Genome-wide association studies have linked common UMOD variants to chronic kidney disease, hypertension, and type 2 diabetes. Studies in umod knockout mice provide a more mechanistic

an overview of tubular function

link to physiological function. Tamm–Horsfall protein may guard against infection by inhibiting adherence of bacteria to tubular cells and act as a constitutive inhibitor of calcium crystallization in tubular fluid (Rampoldi et al., 2011).

Major control mechanisms As with the proximal tubule, the loop of Henle is well endowed with sympathetic nerve endings: the medullary thick limb is the most densely innervated of any tubule segment. Increased RNSA increases Na+ reabsorption in the loop of Henle as a whole and β-adrenergic stimulation in vitro stimulates Na+ flux in isolated TALH. In vivo bilateral denervation studies suggest that RNSA increases NKCC2 by a cyclic AMP-dependent mechanism (Torp et al., 2012). Aldosterone has a stimulatory effect on Na+ and K+ reabsorption in superficial loops of Henle perfused, although this has not been a consistent finding. NKCC2 does not seem to be an aldosterone-induced protein, but studies report stimulation of basolateral Na+/K+-ATPase (Grossman and Hebert, 1988), which would increase the driving force for Na+ transport reabsorption. The receptor through which these effects are mediated is undefined:  both mineralocorticoid and glucocorticoid receptors are expressed in the thick limb but 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2), the enzyme that governs ligand access, is absent (Ackermann et al., 2010). Circulating vasopressin stimulates Na+ reabsorption in the TALH through V2 receptor-mediated activation of adenylate cyclase and increased production of cAMP. This cascade culminates in a coordinated activation of key components in the transcellular reabsorptive pathway: apical K+ and basolateral Cl− channels are activated and there is increased trafficking of NKCC2 to the apical membrane Vasopressin also phosphorylates two threonine residues in the N-terminal of NKCC2, which may contribute to enhanced transporter activity (Ares et al., 2011). Other hormones that lead to increased intracellular cAMP such as parathyroid hormone (PTH), calcitonin, and glucagon increase NKCC2 activity and stimulate reabsorption in the TALH. Intraluminal factors inhibiting reabsorption in the loop of Henle include eicosanoids (e.g. prostaglandin E2), produced intrarenally from arachadonic acid metabolism by cyclooxygenase, endothelin-1 (ET-1) and extracellular ATP. The TALH is second only to the collecting duct in terms of ET-1 synthesis. ET-1, via ETB receptors, inhibits sodium transport via an effect on NKCC2 (Ramseyer et al., 2011). ATP is released by TALH cells in response to increased flow and inhibits NKCC2 activity via P2 receptors in both the apical and basolateral membranes. Nitric oxide production (through NOS3) may be the final inhibitory mediator for these paracrine factors (Garvin et al., 2011). Increased extracellular Ca2+ inhibits Na+ transport in the TALH, reducing the driving force for calcium reabsorption by the paracellular cation shunt pathway. The increased interstitial Ca2+ is detected by the G-protein coupled Ca2+-sensing receptor, localized on the basolateral membrane. Activation of the receptor increases the production of 20-HETE and thereby inhibits NKCC2, ROMK, and the basolateral Na+/K+-ATPase. The cognate ligand is Ca2+ but the receptor is activated by a number of other divalent and trivalent cations, including Mg2+ (Gamba and Friedman, 2009). Mutations in this receptor can cause a bartterlike syndrome, sometimes called Bartter type 5, but this nomenclature is not widely accepted.

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Distal tubule and collecting duct The distal tubule lies between the macula densa and the point of confluence with another distal tubule at which a collecting duct is formed. The distal tubule incorporates several subsections: the distal convoluted tubule (DCT), the connecting tubule (CNT), and a short section of the cortical collecting tubule, often referred to as the initial collecting tubule. The collecting duct is divided into the cortical (CCD), outer medullary (OMCD), and inner medullary collecting duct (IMCD). In addition to a cell type characteristic of each subsegment, intercalated cells are found in much of the distal nephron. In general, adjacent cells have a much deeper contact than in the proximal tubule and the tight junctions are much less ‘leaky’:  paracellular flux is not so common and the epithelia is able to maintain a larger transepithelial PD.

Distal convoluted tubule The DCT makes up approximately 50% of the total distal tubule. The first part (DCT1) consists exclusively of DCT cells while DCT2 also includes a small proportion of intercalated cells. The DCT has the highest density of mitochondria along the nephron. These cluster along the basolateral membrane, which has a considerably higher surface area than that of the apical membrane. The epithelial of the DCT is very plastic and will amplify or regress depending on sodium delivery. The exact cue to this is unknown but transepithelial flux of Na+ is likely to be a major factor. In the apical membrane, DCT cells express a Na+Cl− cotransporter (TSC/NCC) sensitive to thiazide diuretics binding to the chloride-binding site. The apical membrane also contains the epithelial calcium channel (ECaC or TRPV5), a ROMK-like potassium channel, a K+Cl− cotransporter and NHE2 (Fig. 20.8). In keeping with a central role for the DCT in Na+ homeostasis, basolateral Na+/K+-ATPase is greater than in any other nephron segment. The basolateral membrane also has a chloride exit channel (CLC-K2) and a Ca2+-ATPase. An inwardly rectifying potassium channel (Kir4.1) mediates K+ recycling across the basolateral membrane. The importance of this process is underscored by the description

DCT

of the salt wasting EAST syndrome that results from mutations in encoded by KCNJ10, the encoding gene (Bockenhauer et al., 2009). The major route for Na+Cl− transport across the apical membrane is NCC, coupled to exit across the basolateral membrane via Na+/K+-ATPase and ClC-K2 respectively. Loss of function mutations in the encoding gene, SLC12A3, cause Gitelman syndrome (Seyberth and Schlingmann, 2011), which presents as a milder form of Bartter syndrome. A point of differential diagnosis, however, is that Gitelman syndrome presents with hypocalciuria and hypomagnesaemia, while Bartter syndrome patients have normal or hypercalciuria and typically normal Mg2+ levels. The basis for these differences is explained in the section on Ca2+ transport. Pseudohypoaldosteronism type 2 (Gordon disease or familial hypertension with hyperkalaemia (FHH)) presents as a gain of function of NCC: hypertension in these patients is very sensitive to low-dose thiazide diuretics. Human genetic screening identified mutations in two members of a novel family of regulatory kinases, the with-no-lysine (WNK) kinases isoforms 1 and 4. The expression of WNK4 is limited to the aldosterone-sensitive distal nephron. In the DCT is normally exerts an inhibitory effect on thiazide-sensitive Na+ transport, partly by reducing expression of NCC at the apical membrane. This inhibitory effect is no longer observed in PHAII-WNK4 mutants, giving rise to dysregulated NCC activity. The physiological effects of WNK1 are not well understood. It is expressed along the nephron but there is no evidence to suggest a direct interaction with NCC and it is likely that WNK1 affects the activity of other regulatory kinases (Gamba, 2009). The DCT is not a major site of potassium secretion: the absence of electrogenic sodium reabsorption in DCT1 results in an apical membrane potential close to the K+ equilibrium potential and there is thus little driving force for ROMK-mediated section. DCT2 contains hybrid cells that also express the epithelial sodium channel (ENaC) in the apical membrane (Nesterov et  al., 2012). ENaC-mediated Na+ entry depolarizes the apical membrane, sustaining some K+ secretion. Potassium secretion can also occur via K+Cl− cotransport (KCC1) but only if the luminal chloride concentration is unusually low. This can happen if the delivery of HCO3− is high (Amorim et al., 2003), a situation that also favours

Lumen

Interstitial fluid

3Na+

Na+ NCC

Kir4.1 K+

K+

KCC1

Kv1.1

2K+

Cl–

Cl–

Cl–

K+

TRPV-5 Na+ ENaC

Fig. 20.8  Major transport pathways in the distal convoluted tubule.

Ca2+ Ca2+

ClC-Kb

Na+ Ca2+

NCX1

chapter 20 

NHE2-mediated NaHCO3 reabsorption in this segment (Bailey et al., 2004).

Major control mechanisms In order to be physiologically active, NCC must be phosphorylated on threonine and serine residues in a regulatory domain in the N-terminus (Rafiqi et al., 2010) and correctly trafficked to the apical membrane of the DCT. Recent studies indicate that angiotensin II, via activation of AT1R, can increase NCC phosphorylation (van der Lubbe et al., 2011) and stimulate thiazide-sensitive Na+ transport (Ashek et al., 2012). In vivo studies have shown that corticosteroids can increase NCC-mediated sodium transport in the DCT (Velazquez et  al., 1996). The exact mechanisms of action are not clear and both MR and GR are expressed in the DCT, but not 11β-HSD2 (Ackermann et al., 2010). Other hormones, such as vasopressin and insulin can phosphorylate NCC and increase apical expression. In the chronic setting, the kidney escapes from the Na+-retaining effects of aldosterone (aldosterone escape) by downregulation of NCC (Turban et al., 2003).

The connecting tubule The CNT consists of two cell types: CNT cells, the majority, and intercalated cells, the remainder (~ 30% of the total). Like DCT cells, CNT cells exhibit basolateral amplification and have mitochondria along this membrane. The majority of Na+ transport in the CNT is amiloride-sensitive. These cells express NHE2, which can be inhibited by amiloride analogues, but the major route for sodium reabsorption, is via the ENaC in the apical membrane, coupled to basolateral Na+/K+-ATPase activity. Notably, ENaC activity in the CNT does not seem to be regulated by aldosterone (Nesterov et al., 2012).

CCD

an overview of tubular function

There is functional evidence for thiazide-sensitive Na+ transport, but NCC expression is virtually zero. This may reflect a novel, pathway for Na+ reabsorption in the intercalated cell (see Fig. 20.14), in which Cl− entry through pendrin is recycled on a newly identified apical transporter, NDCBE (Na+-driven, Cl−, bicarbonate exchanger (SLC4A8)), in exchange for Na+ (Leviel et al.). This system is best described in the collecting duct and may explain the long-standing observation that a significant proportion of Na+ transport in the cortical collecting duct is sensitive to thiazides. The remainder is amiloride-sensitive and in the CNT, ENaC-mediated Na+ entry depolarizes the apical membrane with respect to the basolateral membrane, generating a large-lumen negative PD of 30–40 mV. The apical membrane also expresses two K+ channels in the apical membrane: the low conductance (~ 35 pS) K+ channel, ROMK (KCNN1) and a larger (~ 100 pS), big potassium (BK) (KCNMA1) channel (Malnic et  al., 2013). The high K+ permeability of the apical membrane favours K+ secretion and exerts a repolarizing influence on the apical membrane potential, thereby sustaining ENaC-mediated Na+ transport. The CNT is also an important site for Ca2+ reabsorption, accounting for approximately 15% of the filtered load and exerting the final regulation of urinary excretion under the control of PTH (see below). CNT is the most proximal site of vasopressin-regulated water transport, expressing both AQP2 water channels and the V2 receptor. The route of Cl− transport is not clear. A basolateral exit pathway (ClC-K2) is available and Cl− might enter the cell via NDCBE in the β-intercalated cell (Leviel et al.). Alternatively, chloride might be reabsorbed paracellularly.

The collecting duct The initial collecting duct is the final segment of the distal tubule and has the same structural makeup as the CNT, comprising mainly

Principal cell

Lumen

Interstitial fluid

Na+ Na+

ENaC

K+ K+ K+

ROMK

H2O AQP-2

Fig. 20.9 Na+ reabsorption in the principal cell of the connecting tubule and collecting duct.

AQP-3 or 4

179

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fluid, electrolyte, and renal tubular disorders Cortisol

11βHSD2

Aldosterone

MR

Transcription MR

Na+ ENaC

Cortisone Na+

Protein synthesis

Na+ K+

Fig. 20.10  Aldosterone specificity in the distal nephron is governed by the enzyme 11β-dehydroxysteroid dehydrogenase type 2 (11β-HSD2), which converts glucocorticoids into metabolites that are not able to activate the mineralocorticoid receptor (MR).

of principal cells with intercalated cells making up the remainder. The OMCD is much the same as the CCD, both in appearance and cell structure. There is a diminution in the proportion of intercalated cells during the transition from the outer to the inner stripe and are absent from the terminal IMCD. In this segment, structurally distinct IMCD cells replace principal cells. In the initial collecting tubule (ICT), CCD, and OMCD, the major entry pathway for Na+ is apical ENaC. Expression of this channel, and of the basolateral Na+/K+-ATPase, declines progressively along the collecting duct and the principal cells become less densely packed with mitochondria. ENaC is not found beyond the first third of the IMCD, being replaced by an amiloride-sensitive, non-selective cation channel. Mechanistically, Na+ reabsorption in the CCD proceeds via the same mechanism as in the CNT: entry through ENaC is electrophysiologically coupled to K+ secretion via ROMK/BK channels. The OMCD and IMCD do not express apical K+ channels: ENaC-mediated Na+ reabsorption is, therefore, self-limiting as depolarization of the apical membrane attenuates the electrochemical gradient for Na+ entry (Fig. 20.9). Under sodium replete conditions, ENaC reabsorbs approximately 1–2% of the filtered Na+ load (Ashek et al., 2012). Sodium transport in this segment is plastic and strongly influenced by the RAAS. In the short term (< 4 hours), aldosterone stimulates Na+/K+-ATPase activity and regulates ENaC trafficking, prolonging the half-life of the channel complex in the apical membrane. In the long term, MR activation increases the expression of the basolateral pump, ENaC and ROMK potassium channels. Constitutive activation of ENaC causes the hypertensive Liddle syndrome. ENaC is composed of three subunits (alpha, beta, and gamma), encoded by distinct genes (SCNN1a, SCNN1b, and SCNN1c). Genetically, Liddle syndrome arises from mutations in β or γ subunits that impair the removal of the channel complex from the apical membrane (Palmer et al., 2012). Conversely, the salt-wasting disorder of pseudohypoaldosteronism type 1 is caused by loss-of function mutations in the mineralocorticoid receptor (PHA1A) or by mutations in the ENaC genes (PHA1B), rendering the channel non-functional (Mullins et al., 2006). In mice, deletion of Scnn1a, Scnn1b, or Scnn1c causes perinatal lethality. More sophisticated genetic approaches have deleted specifically the αENaC subunit from cells of collecting duct lineage (Rubera et al., 2003) or from cells of the CNT (Christensen et al.,

2010). These studies strongly suggest that sodium balance, even under conditions of dietary sodium restriction, is critically dependent on ENaC in the late DCT and CNT.

Aldosterone action and 11β-hydroxysteroid dehydrogenase type 2 In vitro, MR can be activated with equal potency both by aldosterone and cortisol; in vivo, ligand access to MR is determined by co-localization with 11β-HSD2 (Fig. 20.10). By catalysing the rapid conversion of cortisol into cortisone, which does not activate MR, 11β-HSD2 confers upon MR the specificity to aldosterone that it inherently lacks (Funder et  al., 1988). MR and 11β-HSD2 have overlapping distributions in the collecting duct, helping to define the ‘aldosterone-sensitive distal nephron’. The DCT, however, does not express the enzyme (Ackermann et al., 2010) and the control by corticosteroids of sodium transport is not fully understood. In rats, pharmacological inhibition of 11β-HSD2 increased sodium reabsorption by the collecting duct (Bailey et al., 2001). In humans, excess intake of liquorice, which contains the 11β-HSD2 inhibitor glycyrrhetinic acid, causes a sodium-dependent hypertension. Inactivating mutations in the encoding gene (HSD11B2) cause the hypertensive syndrome of apparent mineralocorticoid excess (AME): a mouse model of this disorder presents with hypertension and severely hypokalaemia and impaired renal Na+ excretion due to activation of ENaC (Bailey et al., 2008). AME is an extreme phenotype and, like the other Mendelian blood pressure disorders, is very rare. Nevertheless, these disorders illustrate the fundamental role of renal Na+ transport in blood pressure. Moreover, mild mutations in these same genes may be prevalent in the essential hypertensive population (Wagner, 2008), particularly in those individuals with low-renin or salt-sensitive hypertension. For example, several association studies link polymorphisms in HSD11B2 to blood pressure and mice with 50% of the normal levels of enzyme salt-sensitive blood pressure due to an impaired ability regulate ENaC activity (Craigie et al., 2012).

Major control mechanisms The major regulator of Na+ reabsorption and K+ secretion in the distal nephron is aldosterone, which is secreted from the zona

chapter 20 

glomerulosa of the adrenal cortex in response to plasma angiotensin II (reflecting effective plasma volume) and K+ concentration. The glucocorticoid receptor is also expressed throughout the distal nephron and provides an alternative pathway for ENaC regulation, particularly when the hypothalamic–pituitary–adrenal axis is activated (Bailey et al., 2009). Vasopressin is emerging as a powerful regulator of Na+ reabsorption in the collecting duct, acting synergistically with aldosterone activating ENaC via V2 receptor activation (Stockand, 2010). Another neurohypophysial hormone, oxytocin, is natriuretic due to nitric oxide-dependent inhibition of tubular reabsorption and by stimulating the release of atrial natriuretic peptide (Gimpl and Fahrenholz, 2001). The natriuretic peptides are a complex family of peptides influencing cardiovascular and renal function. Atrial natriuretic peptide, released from atrial myocytes in response to stretch, increases sodium excretion through coordinated inhibition of the apical cation channel and basolateral Na+/K+-ATPase in the IMCD (Beltowski and Wojcicka, 2002). Natriuretic peptides also act as antagonists of the RAAS. A wide-range of paracrine factors influences collecting duct function. Extracellular nucleotides regulate Na+ and water transport through activation of P2X and P2Y receptors (Bailey and Shirley, 2009). Endothelins also influence ENaC activity (Kohan et al., 2011). These systems have complex levels of interaction and probably serve to modify the overall ‘tone’ set by the RAAS. Proteolytic cleavage of the γ subunit by serine proteases (e.g. prostatin, trypsin, chymotrypsin, and elastase) will increase ENaC activity (Rossier and Stutts, 2009). ENaC cleavage is regulated in vivo by aldosterone and may also underpin the sodium retention observed in proteinuric states such as nephrotic syndrome. Several of the key transporters in the distal nephron have marked circadian rhythm of expression (Stow and Gumz, 2012). ENaC, for

an overview of tubular function

example, is under control of the clock gene per1 and disruptions in circadian control have been linked to non-dipper patterns of blood pressure (nocturnal hypertension), a risk factor for cardiovascular and kidney disease.

Urine concentration and dilution (See Chapter 22.) The loop of Henle reabsorbs a considerable amount of sodium, potassium, and water. In addition to these transepithelial fluxes, a major function of the loop of Henle is the generation and maintenance of the interstitial osmotic gradient that increases from approximately 290 mOsm/kg in the renal cortex to approximately 1200 mOsm/kg the tip of the medulla. The driving force for this is the reabsorption in the TALH of solute without water, which generates a ‘horizontal’ osmotic gradient of approximately 200 mOsm/kg at any point between the tubule fluid and interstitium. This ‘single osmotic effect’ also exists at any given level between the ascending and descending limb because the latter has high osmotic permeability and is in equilibrium with the surrounding interstitium. The hairpin structure of the loop, in which flow in the ascending limb is in the opposite direction to that in the descending limb, multiplies the single effect (countercurrent multiplication), creating a much larger ‘vertical’ or corticomedullary gradient (Fig. 20.11). The highest degree of urine concentration is found in mammals with the longest renal papilla. This partly reflects the increased length of the loop of Henle, which in species adapted to arid climates can multiply the single effect to 11 Osmol/L/kg at the papillary tip. The countercurrent multiplier function of the loop of Henle dilutes the tubule fluid (fluid exiting the TALH has an osmolarity of ~ 100mOsm/kg) and concentrates the medullary interstitium. This creates the potential to produce hypotonic or hypertonic urine

Na+ H2O

Na+ Isotonic

290

H2O

290

Hypertonic

Na+

Na+ 100

290

Na+ 600

600

Na+ 400

H2O

900

Na+

Inner medulla 700

900

Hypertonic 1200

Outer medulla

600

H2O

900

Cortex

1200

Fig. 20.11  Countercurrent multiplication of the ‘single osmotic effect’ in the loop of Henle.

1200

181

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in response to homeostatic requirements, by varying the permeability to water of the collecting duct system. The key hormone is vasopressin (antidiuretic hormone), released from the hypothalamus, via the posterior pituitary, in response to increased plasma osmolarity or decreased circulating volume. Vasopressin binds the V2 receptor, activating a signalling cascade that leads ultimately to insertion of AQP2 into the apical membrane of the principal cell. The basolateral membranes of these cells constitutively express AQP3 and AQP4: apical insertion of AQP2 is the rate-limiting step for transepithelial water reabsorption. When plasma vasopressin is high, the hypotonic fluid delivered into distal tubule becomes isotonic by the ICT and progressively hypertonic as it descends through the MCD. In conditions of hydration, when vasopressin is low, little water is extracted during passage through the distal nephron and the final urine can be further diluted by the continued reabsorption of sodium chloride.

The role of urea The thin limbs of the loop of Henle are permeable to urea (ascending > descending), the TALH and distal nephron are impermeable up to the terminal section of the IMCD. In this part of the nephron, vasopressin-dependent water reabsorption has led to a high urea concentration within the tubule fluid. Vasopressin also determines the urea permeability of the terminal IMCD: V2 receptor activation increases the expression of urea transporters (UT-A1 and UT-A3) in the apical and basolateral membrane respectively, facilitating the passive reabsorption of urea into the inner medullary interstitium (Fenton and Knepper, 2007). Some of this urea enters the vasa recta and some the S3 segment of the proximal tubule and the descending and ascending thin limbs of the loop of Henle. This is then returned to the IMCD to be reabsorbed. The net result of this urea ‘recycling’ process is to add urea to the inner medullary interstitium, thereby increasing interstitial osmolality. The high urea concentration within the MCD is rendered osmotically ineffective since it is balanced by a similarly high urea concentration in the medullary interstitium. This allows large quantities of urea to be excreted without obligate osmotic diuresis. Moreover, the concentration of urea in the medullary interstitium increases water abstraction from the thin descending limbs of deep nephrons, raising the intraluminal Na+ concentration within these structures. Thus urea recycling also creates a concentration gradient that in theory could account for passive Na+ reabsorption from the thin ascending limbs, which lack Na+/K+-ATPase. However, this concept is challenged by studies in mice with genetic deletion of UT-A1 and UT-A3, which have reduced urea concentration in the inner medullary interstitium but a normal interstitial Na+Cl− gradient (Fenton and Knepper, 2007). Thus, the mechanisms responsible for the inner medullary electrolyte gradient are still undefined. An interesting theory is that peristaltic contractions observed in the renal pelvis compress rhythmically the hyaluronic acid matrix in the inner medulla, generating a hydrostatic pressure gradient to create the single effect: paralysis of the papillary wall reduces the osmolarity of the inner medulla (Pruitt et al., 2006). It is worth emphasizing, however, that the ultimate driving force for countercurrent multiplication is active Na+ reabsorption in the TALH. This is underscored by the disruption of the osmotic gradient when loop diuretics are given.

Countercurrent exchange in the vasa recta If the capillaries supplying the renal medulla had the usual anatomical arrangement of a capillary network, then medullary blood would rapidly dissipate the medullary osmotic gradient as the hypertonic interstitium equilibrated with isotonic capillary blood. This does not happen to any appreciable extent, because the vasa recta also have a special anatomical arrangement, embracing the epithelial structures in a U-shape. The blood does indeed equilibrate with the neighbouring interstitium but solute entry and water loss in the descending vasa recta are offset by solute loss and water entry in the ascending vasa recta. Although countercurrent exchange is a passive process, contractile cells, called pericytes, control vasa recta flow. This epithelial vascular cross-talk is modulated by a variety of autocrine/paracrine agents (e.g. nitric oxide, eicosanoids, adenosine ATP), and helps to match blood flow to transport in the TALH. Countercurrent exchange applies also to oxygen, which diffuses from descending to ascending vasa recta. This ‘shunting,’ combined with ongoing energy-dependent salt transport in the TALH, renders medullary tissue relatively hypoxic. Nevertheless, medullary cells are adapted to this hostile environment. They have a higher capacity for glycolysis than do cells in the cortex and hypoxia and hyperosmolarity induce, via the transcription factor TonEBP (NFAT5), the expression of proteins that protect the cell and inhibit apoptosis (Burg et al., 2007).

Renal H+/HCO3− transport (acid–base balance)

In individuals on a typical acid-producing diet, the kidneys must reabsorb essentially all the filtered HCO3− (> 4000  mmol/day) and add sufficient extra HCO3− to the plasma to regenerate the buffer anions consumed in buffering the daily acid load (normally ~ 50 mmol/day).

Acid–base transport (See Chapter 24.)

Bicarbonate reabsorption The bulk of filtered HCO3− (~ 80%) is reabsorbed in the proximal tubule, largely in the S1 and S2 segments. About half of the remainder is reabsorbed in the loop of Henle, the rest in the distal tubule and collecting duct. HCO3− reabsorption is indirect: H+ and HCO3− ions are generated in tubular cells (facilitated by intracellular carbonic anhydrase (type II)); the H+ ions are secreted into the lumen, whereas the HCO3− ions enter the plasma. In the proximal tubule, H+ is secreted mainly via apical NHE3 but also via apical H+-ATPase. The secreted H+ ions combine with filtered HCO3− ions to form H2CO3, which is rapidly converted to CO2 and H2O in the presence of apical carbonic anhydrase (type IV); CO2 and H2O diffuse into the cell; both moieties are able to use AQP1 water channels (Endeward et al., 2006). The HCO3− ions generated within the cell enter the interstitial fluid (and thence plasma) via the basolateral Na+HCO3− cotransporter (NBC-1, SLC4A4), which carries three HCO3− ions to one Na+ ion. The net result of these processes is that a filtered HCO3− ion is removed while another one replaces it in plasma (Fig. 20.12). Bicarbonate reabsorption in the loop of Henle takes place mainly in the TALH. Secretion of H+ into the lumen is largely via NHE3, although H+−ATPase makes a modest contribution. There is no apical carbonic anhydrase in the loop of Henle. Intracellularly

chapter 20 

Lumen

NHE3

Interstitial fluid 3Na+

Na+

2K+

H+

H+ H2CO3 CA IV CO2 + H2O AQP-1

K+

HCO3–

H+

HCO3– + H+

an overview of tubular function

3HCO3–

H2CO3

Na+ NBC1A

CAII CO2 + H2O

Fig. 20.12 HCO3− reabsorption in the proximal tubule is dependent on carbonic anhydrase (CA), types 2 and 4.

generated HCO3− enters the interstitial fluid via a basolateral Na-3 HCO3 cotransporter, as in the proximal tubule, and possibly a Cl/ HCO3 anion exchanger (AE2; SLC4A2). In the early DCT, there is evidence for some H+ secretion through apical NHE2 and H+-ATPase (Bailey et  al., 2004), but the cells responsible for H+ handling beyond the early distal tubule DCT2 to IMCD are the intercalated cells. As their name implies, these are interspersed among the majority cell types in each segment. Intercalated cells comprise approximately 30% of the cells in the distal nephron and come in three varieties, all of which contain carbonic anhydrase (type II) and generate H+ and HCO3− ions: type A (the predominant form), type B (present only in the distal tubule and CCD), and non-A, non-B cells, which might be able to switch their function between the other two types according to the prevailing acid–base status. Type A  intercalated cells have an apical H+-ATPase, which secretes H+ into the lumen; HCO3− crosses the basolateral membrane using a Cl−/HCO3− exchanger (AE1; SLC4A1A). The Cl− ions entering the cell by this route are recycled largely through Cl− channels (CLCNKB) and partly through a K+Cl− cotransporter (KCC4; SLC12A7). Although the H+-ATPase is the major player, the apical membrane contains a second type of proton pump: H+/ K+-ATPase, whose activity is upregulated during potassium depletion (Fig. 20.13). Type B intercalated cells are essentially the reverse of type A intercalated cells: H+ ions are pumped across the basolateral membrane via H+-ATPase, and HCO3− ions enter the lumen via an anion exchanger, in this case pendrin (Fig. 20.14). Although intercalated cells have generally been regarded as being concerned solely with acid–base balance, the Cl− ions entering via pendrin can exit the basolateral membrane through Cl− channels, thus providing a mechanism for transepithelial Cl− reabsorption. Moreover, as described above, recent studies suggest the existence of a thiazide-sensitive, Na+-driven Cl−/HCO3− exchanger (Leviel et al., 2010).

Lumen

Interstitial fluid Na+

NH4+ 2Cl–

H+

NKCC-1 AE-1

Cl–

H+ K+

K+ K+ Cl– K+

NH3 RhCG

NH3 RhCG

Fig. 20.13  An acid-secreting (type A) intercalated cell.

This is achieved by the generation of H+ and HCO3− in tubular cells, by the same mechanism as for HCO3− reabsorption, and the addition of HCO3− to the peritubular plasma. The problem then becomes how to deal with the extra H+ ions generated simultaneously with the extra HCO3− ions. There are two principal means of doing so.

Titratable acid excretion Some of the extra H+ ions are secreted into the lumen (via Na+/ H+ exchange and H+-ATPase), where they react with buffer anions (principally filtered HPO42−); any buffer that escapes reabsorption effectively eliminates H+ ions in the urine. The quantity of H+ lost in this way, determined by back-titrating the urine with strong base to pH 7.4 (hence the term ‘titratable acid’), normally amounts to approximately one-third of overall acid excretion. (A few free H+ ions appear as such in the urine, but these are quantitatively insignificant since the minimum urine pH is ~ 4.5.)

Addition of extra bicarbonate to plasma

Ammonium excretion

As indicated above, the kidneys not only reabsorb virtually all filtered HCO3−, but also add further HCO3− to the renal circulation.

The other means of dealing with the extra H+ ions requires the production of NH4+ ions in the proximal tubule. Proximal tubular cells

183

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fluid, electrolyte, and renal tubular disorders

CD

Lumen

Interstitial fluid H+

HCO3–

Pendrin

Cl– Cl–

NDCBE

Cl–

Na2HCO3–

ClC-KB

Fig. 20.14  A base-secreting (type B) intercalated cell, which also expresses the thiazide sensitive Na+-driven chloride-bicarbonate exchanger (NDCBE).

take up the amino acid glutamine and deaminate it, through reactions catalysed by two enzymes (glutaminase and glutamate dehydrogenase), to NH4+ and α-ketoglutarate (Fig. 20.15). The NH4+ ions are secreted into the tubular lumen largely by substituting for H+ on NHE3 (although some intracellular NH4+ dissociates into NH3, which diffuses across the apical membrane to be ‘trapped’ in the lumen by recombining with secreted H+). The α-ketoglutarate is largely metabolized to glucose, through a series of reactions that consume H+ ions. Most NH4+ ions secreted into the proximal tubule and delivered to the loop of Henle are reabsorbed in the TALH. This reabsorption is partly paracellular, driven by the lumen-positive transepithelial PD, and partly transcellular. Apical uptake of NH4+ is mainly via NKCC2, on which NH4+ can substitute for K+, and basolateral exit is largely via a Na/H exchanger (NHE4; SLC9A4), on which NH4+ can substitute for H+ (Weiner and Verlander, 2011). As a result of events in the TALH, NH4+, in equilibrium with NH3, accumulates in the medullary interstitium. The final, and critical, stage of NH4+ excretion is the transference of this interstitial

NH4+/NH3 to the lumen of the collecting duct. Until recently, this was thought to occur by simple diffusion of NH3 across the basolateral and apical membranes, with ‘diffusion trapping’ of NH4+ in the lumen as a result of H+ secretion. However, it is now clear that simple diffusion cannot account for the NH3 movement; instead, specific transport proteins are involved, notably the rhesus glycoprotein RhCG (Weiner and Verlander, 2011). Genetically engineered mice lacking RhCG have a much reduced capacity to excrete NH4+, owing to reduced permeability to NH3 of apical and basolateral membranes in the CD (Fig. 20.13). Moreover, increased expression of RhCG is seen in the OMCD (in both intercalated and principal cells) of rats subjected to metabolic acidosis, though as yet the mechanism of this apparent adaptation is not known (Wagner et  al., 2011). Although rhesus glycoproteins are the major players, other transporters in the CD may have a subsidiary role in mediating NH4+ excretion. The basolateral membrane of A-type intercalated cells contains a Na+-K+-2Cl−- cotransporter (NKCC1; SLC12A2), on which NH4+ can compete with K+; similarly, NH4+ can compete with K+ on a basolateral Na+/K+-ATPase,

Lumen NHE3

Interstitial fluid

Na+ NH4

+

3Na+ 2K+

+

NH4

K+ Glutamine

α-ketoglutarate2– glucose H+

HCO3– H2CO3

CAII CO2 + H2O

Fig. 20.15  Ammonium secretion by the proximal tubule.

3HCO3– Na+

NBC1A

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but the physiological importance of these transporters in relation to acid–base balance is probably minor (Weiner and Verlander, 2011).

Major control mechanisms Chronic acidosis (of non-renal origin) enhances H+ secretion and HCO3− addition to the plasma, while chronic alkalosis results in HCO3− excretion. A  variety of mechanisms are involved. First, changes in systemic pH are paralleled by changes in intracellular pH, affecting H+ secretion directly. Second, NHE3 is directly upregulated by chronic acidosis. Third, chronic acidosis increases the trafficking of H+-ATPase to the apical membrane and that of AE1 to the basolateral membrane, whilst the activity of β-intercalated cells is downregulated. In chronic alkalosis, the opposite occurs. Finally, chronic acidosis will increase ammoniagenesis by the proximal tubule. In addition to the factors described above, acid–base disorders of respiratory origin affect intracellular PCO2 and thereby influence the generation of protons and bicarbonate within tubular cells. Stimulation of the RAAS by acidosis increases H+ secretion by both the proximal and distal nephron. Direct actions of angiotensin II on NHE3 activity are described. Aldosterone can increases H+-ATPase activity, both directly and due to increased lumen negative PD secondary to ENaC activity. The metabolic alkalosis resulting from hypokalaemia is due to the combined action of several factors. First, hypokalaemia causes a compensatory loss of K+ across the basolateral membrane and a reciprocal movement of H+ into the cell. Second, hypokalaemia stimulates renal ammoniagenesis; third, hypokalaemia increases insertion of H+-ATPase into the apical membrane of the α-intercalated cell (Bailey et al., 1998) and activates H+/K+-ATPase.

Renal handling of some specific solutes Phosphate transport (See Chapter 25.) The kidney plays a central role in phosphate homeostasis. Inorganic phosphate is filtered as HPO42− and H2PO4−, normally in the ratio 4:1. Micropuncture studies in superficial nephrons indicate that approximately 80% of the filtered phosphate is usually reclaimed in the proximal tubule. The urinary excretion normally amounts to approximately 10% of the filtered load, suggesting that a small proportion is reabsorbed beyond the proximal tubule. There is some evidence for phosphate reabsorption in the distal tubule but this is controversial and an alternative possibility is that the proximal tubules of deep nephrons that are inaccessible to micropuncture have high reabsorption rates. Proximal tubular reabsorption of phosphate is transcellular (Fig. 20.2A). Entry across the apical membrane uses one of two Na+-phosphate cotransporters, NPT2a (SLC34A1) and NPT2c (SLC34A3):  NPT2a is the major player, expression being subject to physiological regulation (Prie et al., 2011). Transport across the basolateral membrane is not defined at the molecular level, although there is functional evidence for a Na+-phosphate cotransporter, a phosphate/anion exchanger, and possibly a phosphate channel.

Control of phosphate reabsorption Dietary phosphate is a major factor in the control of absorption. A high phosphate intake lowers, and a low phosphate intake raises, the number of NPT2a cotransporters in the apical membrane.

an overview of tubular function

PTH and glucocorticoids reduce the number of NPT2a cotransporters in the apical membrane and thereby increases phosphate excretion (Biber et al., 2009). Phosphotonins are a group of phosphaturic factors (MEPE, SFRP-4, FGF-23) that inhibit reabsorption in the proximal tubule (Shirley et al., 2010) by reducing the abundance of NPT2a cotransporters in the apical membrane. The intracellular signalling events are not defined. The active metabolite of vitamin D, calcitriol (1.25 dihydroxycholecalciferol) has profound effects on phosphate homeostasis. Evidence supports both stimulation and inhibition of tubular phosphate reabsorption. Inhibitory actions might be indirect, via phosphotonin; stimulatory effects might be direct via a response element in the NPT2a promotor (Biber et al., 2009). Disturbances of acid–base balance affect phosphate excretion:  alkalosis stimulates, whilst chronic acidosis inhibits, apical Na+/phosphate co transporters, causing corresponding changes in excretion rates.

Calcium transport (See Chapter 26.) Around 60% of the filtered Ca²+ is reabsorbed in the proximal tubules, mainly in the PCT but also in the pars recta. No significant Ca²+ transport occurs in the thin descending or thin ascending limbs of Henle, owing to their low permeability to Ca²+: the TALH reabsorbs approximately 25% of the filtered load. The remaining Ca²+ reabsorption takes place in the distal tubule (~ 10% of the filtered load); very little is reabsorbed in the collecting duct (Lambers et al., 2006a). Usually 1–2% of the filtered load of Ca²+ is excreted, the actual figure being closely regulated by the requirements for overall Ca²+ balance.

Proximal tubule In the S1 segment the intratubular Ca²+ concentration increases slightly (by 10–20%), creating a small concentration gradient across the S2 epithelium. Together with the small lumen-positive transepithelial PD, this gradient is sufficient to drive passive paracellular Ca²+ reabsorption; a small proportion may be reabsorbed by solvent drag. A small component of proximal Ca²+ reabsorption is active and transcellular, but little information is available on the molecular mechanisms.

Thick ascending limb of Henle At least half the Ca²+ reabsorption in the TALH is passive and paracellular, driven by the lumen-positive transepithelial PD. Loop diuretics or Bartter syndrome abolish this gradient and are calciuric. The remainder is transcellular, most likely due to passive entry through as-yet-unidentified apical Ca²+ channels, coupled with active exit across the basolateral membrane via Ca²+-ATPase.

Distal tubule Ca²+ is reabsorbed in both the DCT and CNT exclusively through a transcellular route (Fig. 20.8). These segments express the Na+/ Ca²+ exchanger (NCX1; SLC8A1) and a Ca²+-ATPase in the basolateral membrane but the epithelial Ca2+ channel (ECaC or transient receptor potential vanilloid 5 (TRPV5) channel) in the apical membrane is rate-limiting (de Groot et al., 2008). DCT cells exhibit the highest Ca²+-ATPase activity of any nephron segment, and in the DCT region it is the sole mode of basolateral Ca²+ efflux,

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whereas in DCT and in CNT cells the basolateral membrane also contains Na+/Ca²+ exchangers. Transcellular Ca²+ reabsorption is facilitated by calbindin D28k, an intracellular Ca²+ binding protein expressed predominantly in the DCT and CNT. By binding Ca²+, calbindins help to maintain the extremely favourable electrochemical gradient for apical entry; they are also thought to help ‘shuttle’ Ca²+ from apical to basolateral membrane via a direct interaction with TRPV5 (Lambers et al., 2006b).

Regulation of calcium reabsorption Ca²+ reabsorption in the proximal tubule is essentially unregulated; physiological control of Ca²+ excretion is exerted at the TALH and the distal tubule. The principal hormone involved in the regulation of Ca²+ reabsorption is PTH. Although PTH may have a small inhibitory effect in the proximal tubule, it more than compensates for this by stimulating Ca²+ reabsorption in the TALH (at least in the cortical segment) and distal tubule. PTH appears to exert its effect on transcellular, rather than paracellular, reabsorption by stimulating apical Ca²+ uptake and basolateral Na+/Ca+ exchange activity. The effects of the hormone calcitonin on renal Ca²+ reabsorption are somewhat paradoxical. Despite its generally hypocalcaemic action, calcitonin stimulates Ca²+ reabsorption in the TALH and distal tubule by cAMP-dependent mechanisms. Calcitriol (1.25-dihydroxycholecalciferol) targets mainly intestine and bone, but may also affect Ca²+ handling in the kidney: there is evidence that it can stimulate Ca²+ reabsorption in the distal tubule, either directly or by potentiating the effect of PTH. A Ca²+/Mg²+ sensing receptor (see above) in the basolateral membrane of the TALH and distal tubule is activated by increased plasma concentrations and inhibits reabsorption of these cations. Renal Ca²+ excretion is influenced by acid–base status: acidosis increases, and alkalosis reduces, Ca²+ excretion rates. Although this can be attributed partly to changes in the filtered load of Ca²+ (acidosis reduces the proportion of plasma calcium bound to albumin and thereby increases ultrafilterable Ca²+), and partly to non-specific changes in proximal tubular reabsorption, these effects cannot account fully for the phenomenon. A specific inhibitory effect of acidosis on Ca²+ reabsorption in the distal tubule has been documented, and it is thought that the mechanism might involve pH sensitivity of the distal tubular apical Ca²+channel.

Magnesium transport (See Chapter 27.) Mg²+ is largely an intracellular ion, found mostly in bone; only approximately 1% is in extracellular fluid. Typically, approximately 10 mmol of Mg2+ is consumed per day, with approximately 6 mmol being lost in the faeces and the remaining approximately 4% in the urine. In contrast to the situation with Ca2+, bone does not appear to play a role in the control of Mg2+ levels, and there is little evidence for control of intestinal uptake, so Mg2+ balance depends entirely on the regulation of urinary excretion. Approximately 75% of serum Mg2+ is ultrafilterable, and overall tubular Mg2+ reabsorption usually amounts to approximately 97% of the filtered load. Intriguingly, the major site of reabsorption is the loop of Henle: only 15–20% of filtered Mg2+ is reabsorbed in the proximal tubule, whereas up to 70% is reabsorbed in the loop, mainly in the TALH; the remaining 10–15% is reabsorbed in the distal tubule.

There is no evidence for transcellular Mg2+ transport in either the proximal tubule or the TALH. In the proximal tubule S2 segment, both concentration and electrical gradients favour paracellular Mg2+ reabsorption, but the permeability is low. Mg²+ reabsorption is driven by the lumen-positive transepithelial voltage in the TALH (Fig. 20.6). Recent evidence indicates that the integrity of the cation-selective paracellular pathway is dependent on the interaction of at least two tight-junction proteins: claudin-16 and claudin-19 (Hou and Goodenough, 2010). Mutations in the gene for either protein result in the syndrome of familial hypomagnesaemia with hypercalciuria and nephrocalcinosis (Haisch et al., 2011). The final nephron segment in which Mg2+ is reabsorbed is the distal tubule, where its transport is transcellular (Ferre et al., 2011). Entry across the apical membrane may be mediated by the transient receptor potential channel melastatin (TRPM), members 6 and 7, but the concentration gradient for entry is much smaller than that for calcium. It is not yet known whether an intracellular binding protein exists for Mg2+; nor has the mechanism of exit across the basolateral membrane been identified, though it is usually assumed to be a Na+/Mg²+ exchanger (San-Cristobal et al., 2010). Clearly, anything inhibiting normal reabsorptive processes in the TALH (e.g. loop diuretics) is likely to be magnesiuric, and anything stimulating reabsorption in the TALH (e.g. peptide hormones) antimagnesiuric, but the factors controlling Mg2+ excretion are ill understood. It appears that epidermal growth factor (EGF), released locally, has a stimulatory effect on TRPM6 activity, though how (or if) this is regulated remains to be determined. Other factors that can influence Mg2+ reabsorption in the distal tubule include the apical K+ channel Kv1.1 and the basolateral K+ channel Kir4.1 (KCNJ10), the latter often found as a heteromeric complex with Kir5.1. Mutations in the gene encoding Kv1.1 are associated with inappropriately high Mg2+ excretion. This channel normally hyperpolarizes the apical membrane, so its dysfunction reduces the driving force for Mg2+ entry (San-Cristobal et al., 2010). Mutations that disable the basolateral Kir4.1/5.1 channel, by preventing K+ recycling, reduce the activity of the basolateral Na+/K+-ATPase and also result in hypomagnesaemia, though in this case the explanation is elusive.

References Ackermann, D., Gresko, N., Carrel, M., et al. (2010). In vivo nuclear translocation of mineralocorticoid and glucocorticoid receptors in rat kidney: differential effect of corticosteroids along the distal tubule. Am J Physiol Renal Physiol, 299, F1473–85. Amorim, J. B., Bailey, M. A., Musa-Aziz, R., et al. (2003). Role of luminal anion and pH in distal tubule potassium secretion. Am J Physiol Renal Physiol, 284, F381–8. Ares, G. R., Caceres, P. S., and Ortiz, P. A. (2011). Molecular regulation of NKCC2 in the thick ascending limb. Am J Physiol Renal Physiol, 301, F1143–59. Aronson, P. S. (2006). Essential roles of CFEX-mediated Cl(-)-oxalate exchange in proximal tubule NaCl transport and prevention of urolithiasis. Kidney Int, 70, 1207–13. Ashek, A., Menzies, R. I., Mullins, L. J., et al. (2012). Activation of thiazide-sensitive co-transport by angiotensin II in the cyp1a1-Ren2 hypertensive rat. PLoS One, 7, e36311. Bailey, M. A., Fletcher, R. M., Woodrow, D. F., et al. (1998). Upregulation of H+-ATPase in the distal nephron during potassium depletion: structural and functional evidence. Am J Physiol, 275, F878–84.

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CHAPTER 21

Sodium transport and balance: a key role for the distal nephron Laurent Schild Introduction (Na+)

Sodium is the most important contributor to the osmolality of the extracellular fluid (ECF), and hence is a major determinant of the ECF volume. The kidneys constantly maintain the ECF volume by regulating urinary Na+ excretion. The urinary Na+ excretion has to precisely balance the daily Na+ intake to avoid changes in ECF volume. The intravascular volume of the ECF is critically dependent on changes in body Na+ content and represents an important determinant of systemic blood pressure. The arterial pressure of a normal adult is kept within a narrow range and rarely deviates by > 10–20%, even between diurnal and nocturnal periods. A number of pressure control systems are necessary to maintain such constancy in blood pressure; they include baroreceptors and neural reflex systems that respond within seconds or minutes to abrupt changes in blood pressure. This short-term regulation of blood pressure relies mainly on the heart, the blood vessels, and the adrenal medulla. The kidney maintains blood pressure within hours or days by controlling the ECF volume. A.  C. Guyton first described this renal control system for the long-term regulation of blood pressure, and postulated the existence of a unique mean arterial blood pressure called the equilibrium pressure, the pressure at which Na+ intake and output are in balance (Guyton, 1992). If arterial pressure rises above the equilibrium pressure, then the urinary Na+ excretion becomes greater than the net Na+ intake, and circulating volume decreases until pressure returns to equilibrium. This pressure control never stops functioning to balance Na+ intake and output to maintain blood pressure at equilibrium. Of course, this leaves unexplained the reasons and the mechanisms that cause blood pressure to rise in the majority of the hypertensive patients. Elevated blood pressure may result from a high salt intake that exceeds the ability of the kidneys to eliminate Na+. The prolonged ingestion of large quantities of salt increases blood pressure in the dog, rabbit, baboon, and chimpanzee. In humans, a relation between salt intake and blood pressure is well documented from epidemiological studies (Denton et  al., 1995; Elliott et  al., 1996; Meneton et al., 2005). The effect of Na+ on blood pressure is certainly complex and may include factors other than the Na+ handling by the kidney. However, this chapter will focus mainly on regulated Na+ absorption along the nephron, which influences Na+ homeostasis and the maintenance of blood pressure.

Sodium handling by the nephron (See Chapter 20.) More than 99% of the filtered load of Na+ is reabsorbed along the nephron and in the collecting tubule. The major fraction (90%) of the filtered Na+ is reabsorbed along the proximal tubule and in the thick ascending limb (TAL). The remaining 8–10% of the filtered Na+ is reabsorbed in the distal convoluted tubule (DCT), the connecting tubule (CNT), and the collecting duct; the Na+ absorption in the distal nephron and the collecting duct is tightly regulated by aldosterone and vasopressin. In the proximal tubule, the electroneutral Na+/hydrogen (H+) exchanger links Na+ reabsorption to that of bicarbonate. In addition, Na+ absorption is coupled with the uptake of solutes such as glucose, amino acids, phosphate, sulphate, and lactate by different cotransporter systems. The thin descending limb of Henle is impermeable to Na+ ions, but the TAL contributes to the reabsorption of 20–30% of the filtered Na+. Two major transport pathways contribute to the Na+ absorption in this segment, the electroneutral Na-K-2Cl cotransporter and the Na+/H+ exchanger (Fig. 21.1). A particularity of the TAL is the presence of a lumen positive electrical potential generated by the recycling of potassium (K+) across the apical membrane; because of the selective cationic permeability of the tight junctions in the TAL, this lumen positive potential contributes to approximately 50% of Na+ absorption along a favourable driving force for diffusion of Na+ and divalent cations through the paracellular pathway. In the DCT, Na+ absorption is mediated by the electroneutral Na-Cl cotransporter (NCC) that is specifically expressed in the apical membrane of this nephron segment. Further downstream, the epithelial Na+ channel (ENaC) is responsible for electrogenic Na+ absorption. In the late portion of the distal DCT (DCT2), the expression of NCC and ENaC overlap. Further downstream in the CNT, the cortical (CCD) and medullary portions of the collecting duct (MCD), ENaC is found without NCC, and its expression follows a decreasing axial gradient from the CNT down to the MCD (Loffing and Kaissling, 2003). ENaC allows the electrogenic entry of Na+ into the cell along a favourable electrochemical gradient (Fig. 21.1). The resulting depolarization of the apical membrane provides the driving force for K+ secretion through the K+ channels (ROMK) that co-localize with ENaC at the apical membrane of principal cells (Loffing and Kaissling, 2003; Loffing and Korbmacher, 2009). Thus, in the CNT and CCD, the electrogenic Na+ absorption mediated by

Section 2  

fluid, electrolyte, and renal tubular disorders The distal convoluted tubule Thiazides

Na+

+ K+ K

CI–

MR

P

Na+

CI–

The cortical collecting duct The thick ascending limb Furosemide

P

Amiloride

CI–

Spironolactone

ENaC

Na+ ROMK

K+

+ + K+

ROMK

+

Na+

Ca++ Mg++

MR

K+

P

Na+

K+

–

K+

CI –

Na+

Na+ 2 CI– K+

K+ K+

K+

190

CI–

Fig. 21.1  Cellular mechanisms contributing to Na+ absorption in the thick ascending limb (red) the distal convoluted tubule (grey), the connecting tubule, and the collecting duct (yellow). In the thick ascending limb, Na+ absorption is essentially mediated by the Na-K-2Cl co-transporter (NKCC); this cotransport system serves Na+ absorption and K+ recycling across the apical membrane. In the distal convoluted tubule (DCT), the electroneutral Na-Cl cotransporter (NCC) is responsible for Na+ absorption. In the principal cells of the connecting tubule and the collecting duct, Na+ absorption is electrogenic via the epithelial sodium channel (ENaC). In these segments, the negative electrical potential in the lumen provides a favourable driving force for K+ secretion. The transporters NKCC, NCC, and ENaC are the target of the diuretics furosemide, thiazides and amiloride respectively. Spironolactone is an antagonist of the mineralocorticoid receptor (MR).

ENaC is coupled with K+ secretion, and is critically dependent on whole-body K+ status (Frindt et al., 2011). Although it represents only a small fraction of the filtered load, the Na+ absorption in the distal nephron and the collecting tubule is under the tight control of the renin–angiotensin–aldosterone system (RAAS). Following a reduction of the effective circulating volume and/or a dietary salt restriction, the RAAS is activated, leading to a decrease in urinary Na+ excretion. Aldosterone increases Na+ absorption in the DCT, CNT, and CCD, where it increases the abundance of both transporters NCC and ENaC at the apical membrane of principal cells (Velazquez et al., 1996; Kim et al., 1998). Angiotensin II also enhances Na+ absorption in the DCT (Wang and Giebisch, 1996). In addition, arginine vasopressin (AVP) stimulates Na+ absorption in the DCT, CNT, and CCD (Ecelbarger et al., 2000; Pedersen et al., 2010). These nephron segments share in common the expression of the mineralocorticoid receptor (MR), the vasopressin receptor V2R, and the enzyme 11-β hydroxysteroid dehydrogenase type 2 (Bostanjoglo et al., 1998; Bachmann et al., 1999; Mutig et al., 2007). The understanding of the detailed molecular and cellular mechanisms involved in the regulation of Na+ excretion by the kidney has greatly progressed with the identification of the genetic basis of

Mendelian disorders featuring alterations in Na+ homeostasis and elevated blood pressure.

Human diseases The importance of the RAAS in controlling renal Na+ excretion, Na+ balance, and blood pressure is largely supported by use of drugs that lower blood pressure such as diuretics, angiotensin converting enzyme inhibitors, or angiotensin II receptor antagonists. Beside this pharmacological evidence, recent genetic studies have identified renal and adrenal genes responsible for monogenic forms of hypertension. The glucocorticoid-remediable aldosteronism (GRA) and the syndrome of apparent mineralocorticoid excess (AME) are two examples of Mendelian forms of elevated blood pressure (Lifton, 1996). GRA is a defect of the regulated synthesis of aldosterone associated with severe hypertension and elevated plasma aldosterone levels. The genetic defect is a gene duplication of the aldosterone synthase and 11β-hydroxylase, generating a novel gene that places the synthesis of aldosterone under the control of corticotrophin (ACTH), instead of its physiological secretagogue angiotensin II. The high plasma levels of aldosterone can be normalized by suppression of ACTH release with glucocorticoids.

chapter 21 

AME is an autosomal recessive disorder due unphysiological stimulation of the MR by cortisol. This disorder is characterized by a moderate to severe hypertension with very low plasma aldosterone levels, but sensitive to MR antagonists such as spironolactone. The genetic defects of the AME syndrome are loss of function mutations in 11β-hydroxysteroid-dehydrogenase, a key enzyme present in aldosterone-sensitive kidney cells that protects the MR from stimulation by cortisol (Stewart et al., 1987). The MR has a high affinity for cortisol, which is present in the plasma at a higher concentration than aldosterone. Other Mendelian disorders have established the critical role of the NCC or ENaC in the maintenance of salt balance. Pseudohypoaldosteronism type II (PHA-II also named familial hyperkalaemic hypertension or Gordon syndrome) is characterized by a salt-sensitive hypertension exquisitely sensitive to thiazide diuretics, associated with hyperkalaemia, and hyperchloraemic acidosis (Wilson et al., 2001). These clinical features strongly suggest that this disorder is due to an overactivity of the NCC in the DCT. Genome-wide linkage studies showed that PHA-II is genetically heterogeneous: they identified overlapping deletions in chromosome 1 located in the first intron of a gene encoding a serine-threonine protein kinase WNK1. Expression studies of WNK1 in lymphocytes of affected patients showed a fivefold increase in WNK1 expression compared with unaffected patients, suggesting a gain-of-function mutation of WNK1 (Wilson et al., 2001). The WNK1 gene encodes two WNK1 isoforms, a ubiquitous long isoform L-WNK1, and a kidney-specific KS-WNK1, expressed predominantly in the DCT, and missing most of the kinase domain. In another PHA-II locus on chromosome 17, mutations were found in a gene encoding a WNK isoform, WNK4; these mutations target a highly conserved segment of negatively charged residues distal to the kinase domain. Although strong genetic evidence supports the notion that the mutations in the WNK4 gene are the cause in PHA-II, there is no clear evidence that favours loss- or gain-of-function mutations of WNK4 kinase. WNK4 is localized in the aldosterone-sensitive distal nephron at the tight junctions and at the subapical membrane region of the DCT, in the cytoplasm of CNT and CCD (Wilson et al., 2001; Kahle et al., 2004). These genetic studies strongly suggest that the WNK1 and WNK4 kinases are important regulators of NCC in the distal nephron. Liddle syndrome (or pseudoaldosteronism) is an autosomal dominant form of salt-sensitive hypertension associated with low plasma aldosterone, low plasma renin activity, hypokalaemia, and metabolic alkalosis. In the early 1960s, G. W. Liddle reported a case of pseudoaldosteronism, and described the syndrome as ‘a disorder in which the renal tubules transport ions with such abnormal facility that the end result simulates that of a mineralocorticoid excess’ (Liddle et al., 1963). Blood pressure in these patients could be normalized with amiloride and dietary salt restriction, but spironolactone was not effective. The genetic defects are mutations in the last exon of the SCNN1B and SCNN1G genes encoding the β and γ ENaC subunits that delete a conserved proline-rich motif in the cytosolic C-terminus (Shimkets et al., 1964). In vitro experiments could establish that these mutations are gain-of-function mutations, as postulated by G.W. Liddle, leading to an increase in the abundance and in the activity of ENaC at the cell surface (Schild et  al., 1995; Firsov et  al., 1996). The important role of NCC and ENaC in Na+ homeostasis is further demonstrated by Mendelian disorders with mirror images of Liddle syndrome or PHA-II.