American Journal of Transplantation 2002; 2: 203–206 Blackwell Munksgaard
Copyright C Blackwell Munksgaard 2002 ISSN 1600-6135
Meeting Report
Fibrosis and Atrophy in the Renal Allograft: Interim Report and New Directions Lorraine C. Racusena, Kim Solezb and Robert Colvinc a
The Johns Hopkins Medical Institutions, Baltimore MD, USA b University of Alberta Health Science Center, Edmonton, Alberta, Canada c The Massachusetts General Hospital and Harvard Medical School, Boston MA, USA * Corresponding author: Lorraine C. Racusen,
[email protected] The topic of chronic allograft nephropathy/chronic rejection is reviewed, with focus on fibrosing/sclerosing changes and late loss of function in renal allografts. Discussion includes a review of the problem, pathological and clinical findings, and new directions. Emphasis is placed on definition of specific diagnostic entities in these allografts, and identification of ongoing/active processes in this setting that might be amenable to direct intervention. A schema for categorizing these cases is proposed. Key words: Allograft nephropathy, chronic rejection, kidney transplant Received 25 September 2001, revised and accepted for publication 15 January 2002 This document represents an invited summary report from a symposium focused on chronic allograft injury held at Banff, Canada, in April 2001. This symposium, and relevant background, summarized below, led to definition of the following goals for future efforts in this area: stimulation of scientific studies to define specific features enabling definition of etiologic factors producing fibrosis/sclerosis in individual grafts; and definition of early end-points, either functional or morphological, identifying those in early stage of graft deterioration and enabling early recognition of therapeutic efficacy in preventing progressive injury. A follow-up conference is planned for June 2003 in Banff, Scotland, to revisit advances in this area. Other initiatives are being undertaken as well by the National Institutes of Health, the Federal Drug Administration, and other agencies. ‘Chronic’ changes, that is fibrosing changes with tubular atrophy and often with arterial fibrointimal thickening due to previous and potentially ongoing injury, occur in the majority of renal allografts (1). The half-life of cadaveric renal transplants is 12–14 years, with longer survival in living-donor grafts. Pro-
gressive deterioration of function with fibrosing changes accounts for about 35–40% of all late allograft loss. A variety of factors can produce or contribute to fibrosis in the allograft, including ischemia, hypertension, drug toxicity, infection, increased ureteral pressures, de novo or recurrent glomerular disease, non-immune inflammatory processes, and chronic or ongoing allo-immune injury, i.e. true chronic rejection (CR) (2, 3). Since many of these conditions are difficult to differentiate morphologically, the nonspecific term chronic allograft nephropathy (CAN) has been used to denote fibrosing changes in the allograft (4). This term is intellectually preferable to labeling all fibrosing changes as ‘chronic rejection’, since rejection by definition implies injury due to inflammatory processes targeting allo-antigens. However, the term is a tacit admission that specific features to enable definition of specific pathogenic factors are often not present or not recognized. As reviewed by Dr Phil Halloran, five major factors repeatedly correlate with reduced graft function and survival in study after study. These are: donor age, brain death as the cause of donor demise, preservation injury, immune injury/rejection, and recipient factors. These factors impinge on the allograft in a time-dependent fashion (5). Of these, four are non-immune, and could be addressed by donor selection and organ allocation strategies and/or by strategies to reduce non-immune injury and stress. Ongoing immune injury, on the other hand, requires specific therapeutic strategies to prevent or adequately treat the alloimmune response. Likely reflecting the various factors impinging on the allograft, there are several kinetic patterns of progression to allograft dysfunction and loss. As demonstrated by Dr Larry Hunsicker, some kidney transplants show early injury with persistent functional loss followed by stabilization. Others display early injury with persistent functional loss followed by stabilization. Yet others display excellent early function followed by progression, and some remain stable with excellent function throughout the lifetime of the allograft. Studies designed to monitor progression and intervention to avoid allograft loss must be designed with recognition of these patterns, with the goal of achieving stabilization of the slope of progression in the individual patient. Dr J.M. Cecka discussed chronic allograft dysfunction (presumed ‘chronic rejection’) as a population phenomenon, based on large data-base analysis. He noted that, while there has been some increase in allograft half-life over the last 10 years, eventual allograft loss remains a major problem. In large data-bases [e.g. (6)], after censoring for patient death 203
Racusen et al.
with a functioning graft, ‘chronic rejection’ is the most common cause of graft failure, increasing as a percentage of causes of graft failure after 6 months post-transplant. While loosely labeled as ‘chronic rejection’, a term which should be reserved for injury triggered by allo-immune mechanisms, fibrosing/sclerosing allograft damage reflects non-immune and immune damage, including donor age, with associated nephron loss, underlying disease, and less effective repair processes; drug toxicity; and stress to the allograft, including size mismatch, hypertension, and proteinuria. Design of therapeutic interventions and assessment of therapeutic efficacy in preventing and/or treating ongoing alloimmune injury to the allograft ultimately depend on the ability to reliably distinguish CR from the other various causes of fibrosis/sclerosis and dysfunction. In particular, morphologic definition of ongoing alloimmune injury is potentially critical in determining the efficacy of newer immunosuppressive regimens in preventing or treating long-term alloimmune damage to the allograft. As discussed by Dr Michael Mihatsch, two morphologic features have been proposed as specific for ‘true’ CR (7, 8). The first is an arterial lesion characterized by intimal proliferation/thickening, with intimal lymphocytes, and splintering and disruption of the elastica with formation of neo-media and neo-intima. However, it is not clear whether this change is a consequence of previous damage or a reflection of ongoing damage, and the distinction needs to be sorted out if therapy is to be directed at this lesion. The second lesion readily detectable by light microscopy is duplication of basement membranes in glomerular capillaries, so-called chronic allograft glomerulopathy, which is associated with the proteinuria recognized as a feature of some kidneys with deteriorating function. Whether allograft glomerulopathy represents ongoing alloimmune injury, or another type of process, such as autoimmune injury, remains unresolved. There was no disagreement that these features are indeed quite reliable markers of immune/inflammatory damage to the allograft. Specific data were presented indicating that differentiation of fibrosis due specifically to chronic/ongoing alloimmune injury from fibrosis with other causes is clinically relevant. Dr Mihatsch reported that 30% of those with vascular changes as defined above with creatinine greater than 200 meq/L had decreased function or were on dialysis within 3 months of biopsy, compared to only 9% of those with elevated creatinine and CAN lacking these features on biopsy. Duplication of capillary lamina densa in peritubular capillaries analogous to changes in glomerular capillaries may also be potentially specific for CR, especially if moderate to severe; this feature is only reliably detectable by electron microscopy. Monga et al. initially demonstrated lamination of the peritubular capillary basement membrane in allograft kidneys (9, 10). This finding has been associated with chronic allograft glomerulopathy (11). As reviewed by Dr Robert Colvin, Mauiyyedi et al. recently analyzed biopsies from those with decreased function with or without proteinuria, measuring layers in peritubular capillaries (12). They found 4.7 ∫ 1.8 layers in those that stained positive for C4d, a late complement component which 204
binds covalently to vessel wall and appears to serve as a marker for earlier complement activation at the site, vs. 1.9 ∫ 1.2 in those that were C4d –, suggesting that capillary changes may be associated with ongoing alloimmune injury. Controls had 2.5 ∫ 0.9 layers in peritubular capillaries. Dr Colvin also reviewed a recent study of 38 cases of CR defined (by histologic criteria), 31 control allografts with chronic CsA toxicity or nonspecific fibrosis, and 15 native kidneys with chronic/fibrosing renal disease (13). Twenty-three (61%) of cases of CR had peritubular capillary staining for C4d in frozen tissue, a change which they (14, 15) and others (16) have reported as sensitive and specific for antibody-mediated allograft rejection. Of those C4d-positive cases tested, 88% had demonstrable anti-donor HLA antibody; only 2% of the controls had C4d staining, and none of the C4d-negative patients had anti-donor antibody. C4d-positive cases were histologically similar to those without C4d, but 1-year graft survival was higher (62% vs. 25%) in the C4d π group. More C4d-positive cases had had a previous transplant. Treatment of a recent small cohort of C4d π cases with mycophenolate mofetil plus (MMF π) tacrolimus resulted in 100% 1-year graft survival, which probably contributed to the improved survival of the C4d-positive group overall. These data suggest that a significant percentage of cases of CR are antibodymediated, and that C4d may be a useful marker for CR that can be used to guide successful therapy. Some discussants clearly remained concerned about the specificity of C4d staining in settings of early and late allograft dysfunction. However, while the significance of C4d staining, particularly in late biopsies, remains to be defined, this is clearly an important area for further investigation. Participants agreed that the concomitant demonstration of anti-donor antibody in most cases which were C4d π was compelling. As demonstrated by these studies, identifying and defining ongoing/active immune injury in the setting of allograft fibrosis/sclerosis is important, potentially leading to therapy which may provide at least some improvement in function. Identification of apparent late/ongoing antibody-mediated capillary injury with C4d staining may enable institution of successful therapy. Similarly, kidneys with significant tubulitis or infiltrating lymphocytes in a fibrosing intima, both markers of ongoing cell-mediated rejection, in an allograft with deteriorating function may also respond to increased immunosuppression. Indeed, as suggested by Rush et al. (17), treatment of even clinically silent active inflammatory infiltrates may have a long-term impact on graft survival. Another application of immunohistology in the allograft biopsy is the demonstration of vascular cell adhesion molecule-1 (VCAM-1) on peritubular capillaries, which is apparently a specific marker of true CR, when other potential causes of fibrosis are excluded (18). Up-regulation of adhesion molecules on peritubular capillaries, which appear to be damaged in ongoing alloimmune injury, would certainly be consistent with known mechanisms of inflammatory tissue injury. American Journal of Transplantation 2002; 2: 203–206
Fibrosis in the Renal Allograft
Recognition of the earliest signs of later graft deterioration could lead to early therapy to prevent graft loss. Early prediction, in turn, may be possible using both clinical and morphological criteria. Dr Hunsicker reviewed patterns of renal function, calculated using the Cockcroft-Gault equation, that can be defined post-transplant. Clearance at 6 months was related to donor age, donor type and HLA mismatch, and occurrence of acute rejection. He found no relationship between glomerular filtration rate (GFR) at 6 months and subsequent slope of functional loss, and suggested defining an intercept at 6 months, an intercept which would be impacted by early events. Following early graft events, including preexisting donor-related injury to the allograft, a new and stable functional slope is achieved, which generally defines the ultimate outcome of the allograft. This slope was improved by 0 mismatch (but not other degrees of HLA mismatch), nonblack donor and recipient race, and young donor age. Ultimate renal scarring is a function of both initial damage and later/ongoing damage to the allograft. Interventions which benefit the allograft could potentially be identified by positive alteration in the functional slope of the individual allograft. Turning to histologic features which may be useful predictors, Isoniemi et al. were among the first to define early fibrosing changes in allografts with stable function as a marker for later dysfunction (19). They assessed protocol biopsies taken at 2 years from allograft with normal/stable function, using 8 morphological features of sclerosis and inflammation. They found that a significant percentage of these allografts had fibrosing changes, and that their chronic allograft damage index score predicted creatinine at 3, 4, 5, and 6 years posttransplant (and perhaps correlated with function at 2 years as well). This study highlighted the potential for defining morphological changes in the allograft that could be used as early surrogate markers of later functional deterioration in the allograft. Some of the subsequent morphological studies performed on (protocol) biopsies at even earlier time points have identified morphological features that were also predictive, including fibrosis, vascular changes, and total chronic histological score (20–22). Interstitial fibrosis and tubular atrophy and loss are generally highly correlated. Primary tubular injury can result in alterations in tubular phenotype with expression of neo-antigens, including chemotactants for monocyte/macrophages (23). The ‘nonspecific’ mononuclear inflammation around and within atrophying tubules in native kidney may reflect an essentially immune response that may trigger fibrosis, a process to which tubular cells may actively contribute. Conversely, inflammation in the interstitium with capillary injury may lead to ischemia with tubular atrophy and fibrosis. The process of fibrosis is a dynamic one, progressing through several stages before dense organized scar develops. Dr Alison Eddy has divided the process into four phases in rat models (24): (i) cellular activation and injury phase; (ii) the fibrogenic signaling phase; (iii) fibrogenic phase; and (iv) renal destruction, with obliteration of tubules and capillaries. The first three are active phases involving inflammatory and American Journal of Transplantation 2002; 2: 203–206
parenchymal cells, and soluble factors including growth factors, cytokines and matrix proteins. Recent studies suggest that at least in the earlier stages of sclerosing processes, successful intervention to stabilize and even reverse chronic damage is possible (25). This is an exciting avenue for further research, with the potential to prolong allograft survival by preventing irreversible fibrosis. For the consensus discussion at the end of the symposium, a draft schema for chronic allograft dysfunction classification was proposed by Dr Kim Solez that would include all patients, with or without biopsy: 1. Progressive allograft dysfunction – biopsy unavailable or uninformative 2. Chronic/fibrosing allograft rejection (biopsy diagnosis) O Active – with evidence of immunological activity (active infiltrates, C4d staining) O Inactive – characteristic late lesions – arterial, capillary – without activity 3. Specific other diseases (biopsy diagnosis) O E.g. Chronic calcineurin inhibitor toxicity, chronic polyoma virus infection 4. Chronic allograft nephropathy (biopsy diagnosis) O Fibrosing lesions with no etiologically specific lesions Points raised following formal presentations included recognition that C4d staining late post transplant needs more study. The suggestion was made that fibrosis per se should be given primacy (especially as quantitated with newer techniques), recognizing that fibrosis may take place in the allograft without change in creatinine, so that a definition of progressive allograft damage based on function may miss cases in an early phase of fibrosis. Dr Agnes Fogo stressed that criteria for ‘activity’ on biopsy should be expanded from alloimmune/inflammatory activity to include abnormal cell turnover and matrix production in the allograft, with a view to future intervention with anti-fibrosis therapy as well as immunosuppressive therapy. The proposal will serve to stimulate further study, and will be revisited at subsequent consensus conferences, including the Banff Conference to be held near Banff, Scotland in June, 2003. Further information on the proceedings of the Seventh Banff Conference can be found at http://cnserver0.nkf.med.ualberta.ca/Banff/2001.
References 1. Solez K, Vincenti F, Filo RS. Histopathologic findings from 2-year protocol biopsies from a US multicenter kidney transplant trial comparing
205
Racusen et al.
2.
3.
4. 5.
6.
7.
8. 9.
10.
11.
12.
tacrolimus versus cyclosporine: a report of the FK506 Kidney Transplant Study Group. Transplantation 1998; 66: 1736–1740. Monaco AP, Burke JF Jr, Ferguson RM et al. Current thinking on chronic renal allograft rejection: issues, concerns, and recommendations from a 1997 roundtable discussion. Am J Kidney Dis 1999; 33: 150–160. Azuma H, Tilney NL. Immune and nonimmune mechanisms of chronic rejection of kidney allografts. J Heart Lung Transplant 1995; 14: S136–S142. Racusen LC, Solez K, Colvin RB et al. The Banff97 working classification of renal allograft pathology. Kidney Int 1999; 55: 713–723. Prommool S, Jhangri GS, Cockfield SM, Halloran PF. Time dependency of factors. affecting renal allograft survival. J Am Soc Nephrol 2000; 11: 563–573. Takemoto S, Chow YW, Gjerston DW. Transplant risks. In Cecka JM, Terasaki PI, eds. Clinical Transplants 1999. Los Angeles: UCLA Typing Laboratory; 1999. Mihatsch JM, Ryffel B, Gudat F. Morphological criteria of chronic rejection: differential diagnosis, including cyclosporine nephropathy. Transplant Proc 1993; 25: 2031–2037. Mihatsch MJ, Nickeleit V, Gudat F. Morphologic criteria of chronic renal allograft rejection. Transplant Proc 1999; 31: 1295–1297. Monga G, Mazzucco G, Messina M, Motta M, Quaranta S, Novara R. Intertubular capillary changes in kidney allografts: a morphologic investigation of 61 renal specimens. Mod Pathol 1992; 5: 125–130. Mazzucco G, Motta M, Segoloni G, Monga G. Intertubular capillary changes in the cortex and medulla of transplanted kidneys and their relationship with transplant glomerulopathy: an ultrastructural study of 12 transplantectomies. Ultrastruct Pathol 1994; 18: 533–537. Morozumi K, Oikawa T, Fukuda M et al. Electron-microscopic peritubular capillary lesion is a specific and useful diagnostic indicator for chronic rejection of renal allografts showing less specific morphologic lesions in the cyclosporine era. Transplant Proc 1997; 29: 89–92. Mauiyyedi S, Nelson C, Tolkoff-Rubin N, Cosimi AB, Schneeberger EE, Colvin RB. Peritubular capillary lamination. A marker of antibody mediated chronic rejection of renal allografts. Mod Pathol 2000; 13: 176A.
206
13. Mauiyyedi S, Pelle PD, Saidman S et al. Chronic humoral rejection. Identification of antibody-mediated chronic renal allograft rejection by C4d deposits in peritubular capillaries. J Am Soc Nephrol 2001; 12: 574–582. 14. Collins AB, Schneeberger EE, Pascual M et al. Deposition of C4d in peritubular capillaries is a marker of acute humoral renal allograft rejection. J Am Soc Nephrol 1999; 10: 2208–2214. 15. Crespo M, Pascuarl M, Toldoff-Rubin N et al. Acute humoral rejection in renal allograft recipients: I. Incidence, serology and clinical characteristics. Transplantation 2001; 71: 652. 16. Lederer SR, Kluth-Pepper B, Schneeberger H, Albert E, Land W, Feucht HE. Impact of humoral alloreactivity early after transplantation on the long-term survival of renal allografts. Kidney Int 2001; 59: 334. 17. Rush D, Nickerson P, Gough J et al. Beneficial effects of treatment of early: subclinical rejection: a randomized study. JASN 1998; 9: 2129– 2134. 18. Solez K, Racusen LC, Abdulkareem F, Keneny E, Von Willebrande E, Truong LD. Adhesion molecules and rejection of renal allografts. Kidney Int 1997; 51: 1476–1480. 19. Isoniemi HM, Taskinen E, Hayry P. Histopathologic chronic allograft damage index accurately predicts chronic renal allograft rejection. Transplantation 1994; 58: 1195–1198. 20. Nickerson P, Jeffery J, Gough J et al. Identification of clinical and histopathologic risk factor for diminished renal function 2 years posttransplant. JASN 1998; 9: 482. 21. Nicholson ML, McCulloch TA, Harper SJ et al. Early measurement of interstitial fibrosis predicts long-term renal function and graft survival in renal transplantation. Br J Surg 1996; 83: 1082. 22. Seron D, Morso F, Bover J et al. Early protocol renal allograft biopsies and graft outcome. Kidney Int 1997; 51: 310. 23. Grandaliano G, Gewualdo L, Ranieri E et al. Monocyte chemotactic peptide-1 expression and monocyte infiltration in acute renal transplant rejection. Transplantation 1997; 63: 414–420. 24. Eddy AA. Molecular basis of fibrosis. Pediatr Nephrol 2000; 15: 290– 301. 25. Fogo AB. Progression and potential regression of glomerulosclerosis. Kidney Int 2001; 59: 804–819.
American Journal of Transplantation 2002; 2: 203–206
