Renal biopsy seminar


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Presentation on the various aspects of renal biopsy
Author: Vishal Golay, DM trainee in Nephrology, IPGMER, Kolkata

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  • In their initial report in 1951, Iverson and Brun described localization of the lower pole of the right kidney using intravenous pyelography and a lead mark on the skin
  • The result of the biopsy altered management in 86% of cases of nephrotic range proteinuria, 71% of cases of ARF, 45% of cases of CRF, 32% of cases with haematuria and proteinuria, 12% of cases with non-nephrotic proteinuria alone, and 3% of cases with haematuria alone.
  • Mean decsease in Hb after bx is 1g/dl. Transient microscopic hematuria is detected in all cases.
  • Incidence is 0.8-1% and is seen in post transplant kidney biopsies
  • The purpose of fixation is to preserve tissues permanently in as life-like a state as possible. Fixation should be carried out as soon as possible after removal of the tissues (in the case of surgical pathology) or soon after death (with autopsy) to prevent autolysis. Aldehydes include formaldehyde (formalin) and glutaraldehyde. Tissue is fixed by cross-linkages formed in the proteins, particularly between lysine residues. This cross-linkage does not harm the structure of proteins greatly, so that antigenicity is not lost. Therefore, formaldehyde is good for immunoperoxidase techniques
  • A product called paraplast contains added plasticizers that make the paraffin blocks easier for some technicians to cut.
  • IgA shows clumped staining with a branching, polygonal pattern typical of mesangial immune complex deposition. IgG shows bright, finely granular staining of capillary loop basement membranes, indicating the presence of capillary loop immune complex deposits
  • Positive stains deposit electron dense material on the area of interest, so that it stands out as a dark area on a light background. Negative stains penetrate and darken the interstices between areas of interest, which then appear light on a dark background.
  • Renal biopsy seminar

    1. 1. RENAL BIOPSY DR. VISHAL GOLAY 28-10-10 DM Seminar
    2. 2. History of renal biopsy <ul><li>Iverson and Brun (1951)- first renal biopsy description. Aspiration biopsy of kidney. Am J Med 11:324—330, 1951 </li></ul><ul><li>Kark and Muehrcke (1954)-blind prone biopsy. </li></ul><ul><li>Biopsy of kidney in prone position. Lancet 1:1047—1049, 1954 </li></ul><ul><li>New era - biopsy guns & imaging </li></ul>
    3. 3. Vim-Silverman needle
    4. 4. OVERVIEW <ul><li>Indications & contraindications </li></ul><ul><li>The procedure </li></ul><ul><li>Tissue processing and staining </li></ul><ul><li>Brief introduction to microscopy </li></ul><ul><li>Some clinical examples </li></ul>
    5. 5. Indications of renal biopsy <ul><li>Nephrotic syndrome: </li></ul><ul><ul><li>Adult NS </li></ul></ul><ul><ul><li>Children with atypical features </li></ul></ul><ul><li>Acute renal failure: </li></ul><ul><ul><li>Undiagnosed </li></ul></ul><ul><ul><li>Non resolving clinical ATN >3-4 weeks </li></ul></ul><ul><li>Systemic diseases with renal dysfunction </li></ul>
    6. 6. Indications of renal biopsy CONtd…. <ul><li>Sub-nephrotic proteinuria </li></ul><ul><ul><li>>2g/d in DM, early MGN, FSGS, IgAN </li></ul></ul><ul><ul><li><2g/d needs clinicians discretion </li></ul></ul><ul><li>Hematuria </li></ul><ul><ul><li>Isolated ( Q J Med 2004; 97:739–745) </li></ul></ul><ul><ul><li>Associated with proteinuria and abnormal urine sedimen t </li></ul></ul>
    7. 7. Indications of renal biopsy CONtd…. <ul><li>Post transplant </li></ul><ul><li>CKD- </li></ul><ul><ul><li>generally contraindicated </li></ul></ul><ul><ul><li>In moderate dysfunction-potential reversibility and basic disease </li></ul></ul><ul><li>Diabetes Mellitus </li></ul><ul><ul><li>Microscopic hematuria </li></ul></ul><ul><ul><li>Absence of retinopathy and neuropathy </li></ul></ul><ul><ul><li>Onset of proteinuria <5years from diagnosis </li></ul></ul><ul><ul><li>Acute worsening of renal function </li></ul></ul><ul><ul><li>Systemic features </li></ul></ul>
    8. 8. Renal biopsy; a necessary evil? <ul><li>276 native renal biopsies; management was altered in 42% of cases overall. Nephrol Dial Transplant. 1994;9(9):1255-9 </li></ul><ul><li>3 year prospective study involving 80 patients; Pre-biopsy predicted histologic diagnosis was changed in 44% of the patients as a result of the biopsy. Prognosis changed in 57% of the patients. Therapy changed in 31% of the patients. Clin Nephrol. 1986 Nov;26(5):217-21 </li></ul><ul><li>111 renal biopsies in childern; biopsy altered diagnosis in 24.5%. Post biopsy, treatment was changed in 22.6%. 18.9% had a change in both the diagnosis and treatment. Zhonghua Min Guo. 1998 Jan-Feb;39(1):43-7 </li></ul>
    9. 9. contraindications
    10. 10. Preparation of the patient <ul><li>Prerequisites </li></ul><ul><ul><li>Relevant history & examination </li></ul></ul><ul><ul><li>USG KUB </li></ul></ul><ul><ul><li>Hemogram and Hct </li></ul></ul><ul><ul><li>PT/aPTT </li></ul></ul><ul><ul><li>Urine RE/ME & culture </li></ul></ul><ul><ul><li>Bleeding time </li></ul></ul><ul><li>Current Opinion in Nephrology & Hypertension 1999;8: 715-718 </li></ul>
    11. 12. Biopsy gun
    12. 13. Biopsy gun choice <ul><li>14 G guns gives greater number of glomeruli per core than 18-G cores, but the rates of adequate biopsies were similar </li></ul><ul><li>Larger needle provided more tissue and glomeruli but were associated with more pain. </li></ul><ul><li>16-gauge needles are used as a compromise between the need of a sufficient size of tissue and the need of clinical safety. </li></ul><ul><li>Adequate quantity of tissue for diagnosis may be obtained with this needle size in 98.9 per cent of biopsies either in native or in transplanted kidneys. </li></ul><ul><li>OTCN </li></ul>
    13. 14. Procedure <ul><li>Informed consent </li></ul><ul><li>Patient in prone position with wedge or pillow below the abdomen </li></ul><ul><li>Light sedation </li></ul><ul><li>Local anesthesia with 1-2% lignocaine from the skin down to the capsule </li></ul><ul><li>Stab incision can be given to ease biopsy gun entry </li></ul><ul><li>Advance the biopsy gun, when the capsule is reached, instruct patient to take a deep breath and fire the gun </li></ul><ul><li>2-3 cores can be taken from the lower pole of the left kidney </li></ul><ul><li>Press on wound for 2-5 minutes </li></ul>
    14. 16. Procedure
    15. 17. Procedure
    16. 18. USG guided vs blind biopsies <ul><li>Worldwide the trend is towards USG guided biopsies </li></ul><ul><li>129 renal biopsies, </li></ul>Semin Dial. 2007 Jul-Aug;20(4):355-8 USG guided Blind P Mean number of glomeruli 18±9 11±9 0.0001 Repeat biopsy 0% 16% 0.0006 Hematoma requiring intervention 0% 11% 0.006 24 hour Hct 32±5% 30±4% 0.04
    17. 19. USG guide to nephrologist <ul><li>Safer and more efficient then blind biopsy </li></ul><ul><li>In one study of 101 biopsies nephrologists used a portable ultrasound machine for location and depth of the kidney. The results are similar to those of previous studies using automatic devices but under direct ultrasound guidance. </li></ul><ul><li>AJKD 1999;34:955-959 </li></ul>
    18. 20. Post procedure <ul><li>Hand held lens/dissecting microscope can be used to check for the presence of cortical tissue </li></ul>
    19. 21. Post procedure <ul><li>Bed rest is instructed for 18-24 hours </li></ul><ul><li>BP and pulse are monitored in the following way- </li></ul><ul><ul><ul><li>Every 15 mins for 1 hour f/b </li></ul></ul></ul><ul><ul><ul><li>Every 30 mins for 1 hour f/b </li></ul></ul></ul><ul><ul><ul><li>Every hour for 4 hours f/b </li></ul></ul></ul><ul><ul><ul><li>4 hourly for next remaining 24 hours </li></ul></ul></ul><ul><li>Save aliquots of each voided urine sample in clear specimen jars </li></ul><ul><li>Hct monitored 6-8 hours and 18-24 hours after biopsy </li></ul>
    20. 22. Complications of renal biopsy <ul><li>Complications of percutaneous renal biopsy: a review of 37 years experience. Clin Nephrol 1992;38:135-41 . </li></ul>
    21. 23. Complications of renal biopsy Ateriovenous fistula-post biopsy
    22. 24. Complications of renal biopsy <ul><li>Page kidney : compression of kidney by a hematoma leads to high renin hypertension. It is believed to result from microvascular ischemia and alteration of small-vessel hemodynamics from external compression. </li></ul>
    23. 25. Outpatient renal biopsy <ul><li>750 biopsies; Complications were identified 42% patients by 4 h, in 67% patients by 8 h, in 85% patients by 12 h, and in 89% patients at 24 h. After biopsy, an observation time of up to 24 h remains optimal. </li></ul><ul><li>J Am Soc Nephrol 15:142-147, 2004 </li></ul><ul><li>Maya and Allon et al; 100 renal biopsies. 8 hour observation is safe. </li></ul><ul><li>Semin. Dial. 20, 355–358 (2007) </li></ul>
    24. 26. Specimen processing <ul><li>Specimen division </li></ul><ul><li>>8mm - LM/IF/EM </li></ul><ul><li>4-8mm - EM/IF </li></ul><ul><li><4mm - EM </li></ul>
    25. 27. Specimen processing <ul><li>Adequacy of the sample: </li></ul><ul><ul><li>two biopsy cylinders with a minimal length of 1 cm and a diameter of at least 1.2 mm. </li></ul></ul><ul><ul><li>10–15 glomeruli are optimal; very often 6–10 glomeruli are sufficient </li></ul></ul><ul><ul><li>some cases even one glomerulus is enough </li></ul></ul><ul><ul><li>Nephrol Dial Transplant (2006) 21: 1157–1161 </li></ul></ul>
    26. 28. Biopsy tissue examination
    27. 29. Light microscopy <ul><li>1. Fixative: </li></ul><ul><ul><li>Buffered, 10% aqueous formaldehyde solution (formalin)-most commonly used </li></ul></ul><ul><ul><li>Bouin’s </li></ul></ul><ul><ul><li>Duboscq-Brasil </li></ul></ul><ul><ul><li>Zenker’s </li></ul></ul><ul><ul><li>4% paraformaldehyde </li></ul></ul><ul><ul><li>The material processed for LM can serve as reserve material for IHC or EM if either of these other modalities is found lacking glomeruli. </li></ul></ul>
    28. 30. Light microscopy <ul><li>2 . Dehydration: done with a series of alcohols, say 70% to 95% to 100%. </li></ul><ul><li>3. Clearing: xylene, toluene, chloroform, limolene </li></ul><ul><li>4. Embedding : </li></ul><ul><ul><ul><li>Paraffin </li></ul></ul></ul><ul><ul><ul><li>Paraplast </li></ul></ul></ul><ul><ul><ul><li>Plastics- methyl methacrylate, glycol methacrylate, araldite, and epon . </li></ul></ul></ul><ul><ul><li>5. Sectioning: manual/microtome. 2 µm sections are used </li></ul></ul>
    29. 31. Light microscopy <ul><li>Routine stains (paraffin sections) </li></ul><ul><ul><li>Haematoxylin and eosin (HE) </li></ul></ul><ul><ul><li>Periodic acid–Schiff’s (PAS) </li></ul></ul><ul><ul><li>Fibrous tissue stain (i.e. Sirius red, Trichrome, Ladewig, etc.) </li></ul></ul><ul><ul><li>Silver stain </li></ul></ul><ul><ul><li>Protein stain- acid fuchsin–Orange G stain (SFOG). </li></ul></ul><ul><li>Optional stains </li></ul><ul><ul><li>Kossa stain (calcifications) </li></ul></ul><ul><ul><li>Congo red stain (amyloid) </li></ul></ul>
    30. 32. Light microscopy <ul><li>H&E: composition of the tissue (i.e. renal cortex vs medulla, number of glomeruli, cellular infiltration, etc.). </li></ul><ul><li>PAS: delineates in great detail </li></ul><ul><ul><li>glomerular cells, </li></ul></ul><ul><ul><li>mesangial matrix and potential expansion, as well as potential modifications of the composition of the matrix, </li></ul></ul><ul><ul><li>changes of the GBM, i.e. thickening, irregularities, doubling, rupture </li></ul></ul><ul><ul><li>fibrinoid necrosis of the glomerular tuft </li></ul></ul><ul><ul><li>alterations of the vessels, particularly arterial hyalinosis and fibrinoid necrosis. </li></ul></ul>
    31. 33. Light microscopy <ul><li>Protein stains : Immune deposits </li></ul><ul><li>Silver stain : permits the detection of changes of the GBM </li></ul><ul><li>Fibrous tissue stain : extent of fibrosis in the glomerulus or tubular interstitium </li></ul>
    32. 34. Light microscopy-normal
    33. 36. Immunohistochemistry-IF/IP <ul><li>Preparation : </li></ul><ul><ul><li>IF is best performed on unfixed, frozen sections. Tissues can be transported to the laboratory fresh on saline-soaked gauze or in Michel’s fixative. Serial sections are cut at 2–4 mm in a cryostat. </li></ul></ul><ul><ul><li>IP staining requires no special tissue preparation in that the same formalin-fixed, paraffin- embedded material used for LM is also used for IP. </li></ul></ul>
    34. 37. Immunohistochemistry-IF/IP <ul><li>Staining: The antigens that should be routinely examined include: </li></ul><ul><ul><li>immunoglobulins (primarily IgG, IgM and IgA), </li></ul></ul><ul><ul><li>complement components (primarily C3, C1q, and C4), </li></ul></ul><ul><ul><li>fibrin, </li></ul></ul><ul><ul><li>κ - and – λ light chains </li></ul></ul><ul><ul><li>collagen IV alpha chains </li></ul></ul><ul><ul><li>IgG subclasses, </li></ul></ul><ul><ul><li>virus identification, </li></ul></ul><ul><ul><li>lymphocyte </li></ul></ul><ul><ul><li>phenotyping in allografts in suspected cases of PTLD, </li></ul></ul><ul><ul><li>C4d in allograft biopsies </li></ul></ul>
    35. 38. Immunohistochemistry-IF/IP <ul><li>Reported as: </li></ul><ul><ul><li>reaction is positive , </li></ul></ul><ul><ul><li>pattern of staining, e.g. mesangial vs capillary staining pattern, </li></ul></ul><ul><ul><li>linear (or pseudolinear) vs granular staining. </li></ul></ul><ul><ul><li>describe where the deposits are located , e.g. in a subendothelial, intramembranous or subepithelial position </li></ul></ul>
    36. 39. Immunohistochemistry-IF/IP
    37. 40. IgG IgA Idiopathic MGN IgA Nephropathy
    38. 41. Electron Microscopy <ul><li>Fixative : 2-3% glutaraldehyde, 1-4 % paraformaldehyde. </li></ul><ul><li>Processing : </li></ul><ul><ul><li>Instead of the clearing fluid, “transitional fluid”- 1,2 Epoxypropane is used. </li></ul></ul><ul><ul><li>E mbedding done in epon. </li></ul></ul><ul><ul><li>Sections must be less than 80 nm thick in order to allow at least 50% of the electron beam to penetrate the sample. </li></ul></ul><ul><ul><li>Toluidine blue stained 1µm thick sections are used as an initial guide </li></ul></ul><ul><li>Stains : uranyl acetate (stains DNA), lead citrate, gold. </li></ul>
    39. 42. Electron Microscopy <ul><li>Electron microscopy permits assessment of the following: </li></ul><ul><ul><li>The presence and degree of cell proliferation (mesangial vs endothelial cell proliferation) </li></ul></ul><ul><ul><li>Changes in cell structure (i.e. podocyte foot process fusion or podocyte vacuolization) </li></ul></ul><ul><ul><li>Necrosis or apoptosis of cells </li></ul></ul><ul><ul><li>Changes of glomerular basement membrane (i.e. thickening, thinning, splicing, irregularities) </li></ul></ul><ul><ul><li>Localization of immunoglobulin deposits (i.e. mesangial, subendothelial or subepithelial) </li></ul></ul><ul><ul><li>In some renal diseases, specific morphological changes tubuloreticular structures. </li></ul></ul>
    40. 43. Electron Microscopy <ul><li>Immunohistology is essential for diagnosis in 21% of cases and Electron Microscopy is essential in the diagnosis of 8% of cases OTCN </li></ul>
    41. 44. Electron Microscopy
    42. 45. Reporting a Renal Biopsy <ul><li>The final report of a kidney biopsy should include information on: </li></ul><ul><ul><li>The adequacy of the specimen </li></ul></ul><ul><ul><li>A description of the morphological changes in a systematic fashion for each of the compartments of interest (glomeruli, tubules, interstititum, vessels </li></ul></ul><ul><ul><li>The results of immunofluorescence /immunohistochemical studies. </li></ul></ul><ul><ul><li>The results of the electron microscopy </li></ul></ul><ul><li>It is useful to give two types of diagnosis: </li></ul><ul><li>Descriptive </li></ul><ul><li>Final </li></ul>
    43. 46. Reporting a Renal Biopsy
    44. 47. Reporting a Renal Biopsy
    45. 48. Reporting a Renal Biopsy
    46. 49. Reporting a Renal Biopsy
    47. 50. Biopsy in special situations <ul><li>Pediatric age group : </li></ul><ul><ul><li>Sedation is used (ketamine, midazolam, promethazine, BZDs) </li></ul></ul><ul><ul><li>18-G gun is used </li></ul></ul><ul><li>Pregnancy : Indications of biopsy </li></ul><ul><ul><li>Sudden unexplained deterioration of renal function before 30-32 weeks POG </li></ul></ul><ul><ul><li>Symptomatic NS before 30-32 weeks POG </li></ul></ul><ul><ul><li>Active urinary sediments, proteinuria and borderline renal function </li></ul></ul><ul><li>renal biopsy in pregnancy is safe before 30 weeks of pregnancy. </li></ul><ul><li>Acta Obstetricia Gynecologica Scandinavica 2001;80:888–893 </li></ul>
    48. 51. Biopsy in special situations <ul><li>Allograft biopsy : </li></ul><ul><li>Indications- </li></ul><ul><ul><li>Failure of the graft to function in the first 7-10 days after surgery </li></ul></ul><ul><ul><li>Rapid deterioration of renal function of unknown cause after initial good function </li></ul></ul><ul><ul><li>Absence of response to antirejection therapy </li></ul></ul><ul><ul><li>Unexplained nephrotic range proteinuria </li></ul></ul><ul><ul><ul><li>Biopsy technique is the same </li></ul></ul></ul><ul><ul><ul><li>Supine position </li></ul></ul></ul><ul><ul><ul><li>USG guidance is necessary-for position and localisation, and also to exclude presence of intestinal loops, fluid collections </li></ul></ul></ul><ul><ul><ul><li>2 cores of tissue is recommended- the sensitivity of a single core for rejection is 91%; the addition of the second core improves the sensitivity to about 99% </li></ul></ul></ul><ul><ul><ul><li>Half the tissue is used for frozen section </li></ul></ul></ul>
    49. 53. <ul><li>CLINICAL EXAMPLES </li></ul>
    50. 54. <ul><li>SHEIKH ABDUL HALIM </li></ul><ul><li>Kala Azar with Nephrotic Syndrome </li></ul>
    51. 56. Sushmita Das (SLE Class IV) Wire loops
    52. 57. Cellular crescent Fibrinoid necrosis Global proliferation with neutrophilic infiltration
    53. 58. <ul><li>Naima Khatoon </li></ul><ul><li>Lupus nephritis Class IV G( A/C) </li></ul><ul><li>Activity index: 19/24 </li></ul><ul><li>Chronicity index: 4/12 </li></ul>
    54. 59. Kalpana Das (FSGS)
    55. 60. Amal Panja (Crescentic GN)
    56. 61. Sabita Majumdar (Pauci-immune CrGN)
    57. 62. Sukumar Bodhak (IgA Nephropathy) IgA
    58. 63. <ul><li>THANK YOU </li></ul>