The ORION study: comparison of two sirolimus-based regimens versus tacrolimus and mycophenolate mofetil in renal allograft recipients.

Safety and efficacy of two sirolimus (SRL)-based regimens were compared with tacrolimus (TAC) and mycophenolate mofetil (MMF). Renal transplantation recipients were randomized to Group 1 (SRL+TAC; week 13 TAC elimination [n = 152]), Group 2 (SRL + MMF [n = 152]) or Group 3 (TAC + MMF [n = 139]). Group 2, with higher-than-expected biopsy-confirmed acute rejections (BCARs), was sponsor-terminated; therefore, Group 2 two-year data were limited. At 1 and 2 years, respectively, graft (Group 1: 92.8%, 88.5%; Group 2: 90.6%, 89.9%; Group 3: 96.2%, 95.4%) and patient (Group 1: 97.3%, 94.4%; Group 2: 95.2%, 94.5%; Group 3: 97.0%, 97.0%) survival rates were similar. One- and 2-year BCAR incidence was: Group 1, 15.2%, 17.4%; Group 2, 31.3%, 32.8%; Group 3, 8.2%, 12.3% (Group 2 vs. 3, p < 0.001). Mean 1- and 2-year modified intent-to-treat glomerular filtration rates (mL/min) were similar. Primary reason for discontinuation was adverse events (Group 1, 34.2%; Group 2, 33.6%; Group 3, 22.3%; p < 0.05). In Groups 1 and 2, delayed wound healing and hyperlipidemia were more frequent. One-year post hoc analysis of new-onset diabetes posttransplantation was greater in TAC recipients (Groups 1 and 3 vs. 2, 17% vs. 6%; p = 0.004). Between-group malignancy rates were similar. The SRL-based regimens were not associated with improved outcomes for kidney transplantation patients.

Lack of benefit of early protocol biopsies in renal transplant patients receiving TAC and MMF: a randomized study.

We conducted a randomized, multicenter study to determine whether treatment of subclinical rejection with increased corticosteroids resulted in beneficial outcomes in renal transplant patients receiving tacrolimus (TAC), mycophenolate mofetil (MMF) and prednisone. One hundred and twenty-one patients were randomized to biopsies at 0,1,2,3 and 6 months (Biopsy arm), and 119 to biopsies at 0 and 6 months only (Control arm). The primary endpoint of the study was the prevalence of the sum of the interstitial and tubular scores (ci + ct)> 2 (Banff) at 6 months. Secondary endpoints included clinical and subclinical rejection and renal function. At 6 months, 34.8% of the Biopsy and 20.5% of the Control arm patients had a ci + ct score >or= 2 (p = 0.07). Between months 0 and 6, clinical rejection episodes were 12 in 10 Biopsy arm patients and 8 in 8 Control arm patients (p = 0.44). Overall prevalence of subclinical rejection in the Biopsy arm was 4.6%. Creatinine clearance at 6 months was 72.9 +/- 21.7 in the Biopsy and 68.90 mL/min +/- 18.35 mL/min in the Control arm patients (p = 0.18). In conclusion, we found no benefit to the procurement of early protocol biopsies in renal transplant patients receiving TAC, MMF and prednisone, at least in the short term. This is likely due to their low prevalence of subclinical rejection.

Transplant glomerulopathy, late antibody-mediated rejection and the ABCD tetrad in kidney allograft biopsies for cause.

To define the relative frequency of phenotypes of transplant glomerulopathy, we retrospectively reviewed the findings in 1036 biopsies for clinical indications from 1320 renal transplant patients followed in our clinics between 1997 and 2005. Transplant glomerulopathy, defined by double contours of glomerular basement membranes (D), was diagnosed in 53 biopsies (5.1%) from 41 patients (3.1%) at a median of 5.5 years post-transplant (range 3.8-381 months). In cases with D, we studied the frequency of circulating anti-HLA alloantibody (A), peritubular capillary basement membrane multilayering (B) and peritubular capillary C4d deposition (C). B was present in 48 (91%) of D biopsies. C4d staining by indirect immunofluorescence was detected in 18 of 50 D biopsies studied (36%). By Flow PRA Screening or ELISA, A was detected in 33 (70%) in 47 D cases with available sera, of which 28/33 or 85% were donor-specific. Class II (13/33) or class I and II (17/33) were more common than class I (3/33) antibodies. Thus 73% of transplant glomerulopathy has evidence of alloantibody-mediated injury (A and/or C), with ABCD and ABD being the common phenotypes in biopsies for cause. The remaining 27%, mostly BD, may be a different disease or a stage in which A and C are undetectable.

Reproducibility studies on arteriolar hyaline thickening scoring in calcineurin inhibitor-treated renal allograft recipients.

Arteriolar hyaline thickening (AH) is the most characteristic lesion of chronic calcineurin inhibitor nephrotoxicity. This study was performed to compare the inter-observer reproducibility of AH scoring using Banff criteria and a newly proposed criterion. Forty-five nonprotocol post-transplant biopsies from 38 patients immunosuppressed with tacrolimus or cyclosporine A (CsA) were included. The severity of AH was blindly scored by three observers. According to the new criteria, AH is graded based on circular vs. noncircular involvement and the number of arterioles involved. The kappa statistics were used to assess the inter-observer reproducibility. Twenty-seven (60%) biopsies showed AH. The AH grades by both criteria were correlated with serum creatinine at biopsy and inversely correlated with estimated glomerular filtration rate (GFR) (p < 0.05). The recent AH criteria improved the mean pairwise agreement (79.4% vs. 68%) and the overall kappa value (0.67 vs. 0.52) (p = 0.02) compared to Banff criteria. The mean inter-slide variation using Banff and the new criterion were 23% and 27.6%, respectively (p > 0.05). The new AH criterion results in better inter-observer reproducibility, and is clinically validated against serum creatinine and estimated GFR. There is substantial intra-biopsy variation, therefore, evaluation of more than one section is crucial to determine severity of arteriolar damage more accurately.

Identifying the patient at risk for post-transplant lymphoproliferative disorder.

Post-transplant lymphoproliferative disorders (PTLD) are a recognized complication of the immunosuppression required to prevent allograft rejection, occurring in 1-20% of recipients of solid organ transplants. Several factors greatly increase the risk of developing PTLD early post-transplant in any individual recipient. Epstein-Barr virus (EBV) infection is critical in the pathogenesis of the majority of these cases. Pre-transplant EBV seronegativity increases the incidence of PTLD 10- to 75-fold over that of EBV-seropositive recipients. Other risk factors include very young recipient age, cytomegalovirus infection or mismatching (donor positive-recipient negative), aggressive immunosuppression with conventional biologic agents, and the type of organ transplanted. In contrast, the risk of developing PTLD late in the post-transplant course does not appear to be influenced by the type of immunosuppressive agents employed, but rather by the duration of any immunosuppression. The role of EBV in late PTLD is also less certain, as a greater proportion of lesions are not associated with evidence of EBV infection. As the understanding of these risk factors has expanded, opportunities exist to target those populations at highest risk for the development of PTLD for aggressive monitoring and pre-emptive or prophylactic therapy. It is hoped that implementation of such strategies will render early PTLD a preventable complication of transplantation.

Manifestations and treatment of Schimke immuno-osseous dysplasia: 14 new cases and a review of the literature.

UNLABELLED: Schimke immuno-osseous dysplasia (SIOD) is a rare autosomal recessive spondylo-epiphyseal dysplasia. The characteristic features of SIOD include 1) short stature with hyperpigmented macules and an unusual facies, 2) proteinuria with progressive renal failure, 3) lymphopenia with recurrent infections, and 4) cerebral ischaemia. Although 25 patients have been reported with this disorder, the clinical course and phenotype of SIOD are not well characterized. This report summarizes the clinical findings, course and treatment of reported patients and includes 14 additional patients with SIOD. We emphasize the high incidence of cerebral ischaemia and ocular abnormalities, define the high incidence of thyroid dysfunction and blood cytopenia, and confirm the absence of effective and durable medical therapies. CONCLUSION: Schimke immuno-osseous dysplasia is a multi-system autosomal recessive disorder with variable expression that affects the skeletal, renal, immune, vascular, and haematopoietic systems. Medical therapy is limited especially for more severely affected individuals.

Interferon-gamma gene expression during acute graft-versus-host disease: relationship to MHC induction and tissue injury.

The pathogenetic role of interferon-gamma (IFN-gamma) in acute graft-versus-host disease (GVHD) was examined in a murine model. IFN-gamma gene expression was evaluated by northern blotting and mRNA in situ hybridization. The temporal and tissue specific patterns of IFN-gamma gene expression were related to the patterns of major histocompatibility complex (MHC) antigen induction and of tissue injury. Markedly increased levels of IFN-gamma transcripts were seen in the spleen during the early lymphoproliferative phase and coincided with widespread MHC induction in non-lymphoid tissues. Increased IFN-gamma transcripts were also found in the non-lymphoid target tissues during the phase of subsequent tissue injury. These findings support a role for IFN-gamma in leading to widespread MHC induction during acute GVHD and suggest that IFN-gamma may also contribute to target tissue injury during acute GVHD.

On the influence of sample size on the prognostic accuracy and reproducibility of renal transplant biopsy.

INTRODUCTION: The minimal specimen size necessary for accurate interpretation of a renal biopsy has not been identified. We attempted such a determination by three different analyses of a collection of biopsies performed in renal transplants. METHODS: First, we studied the influence of three lesions (glomerulosclerosis, arteriolar hyalinosis, interstitial fibrosis/tubular atrophy) in 199 baseline biopsies, obtained at time of transplantation, on transplant outcome. Secondly, we compared the results from the three lesions in baseline biopsy with those from 114 subsequent core biopsies in the same patients. Thirdly, we compared the two baseline biopsies obtained in 118 paired kidneys in cadaver transplantation where both kidneys were used. RESULTS: For statistically significant prediction of outcome from glomerulosclerosis, we found that a specimen containing at least 25 glomeruli was needed in the baseline biopsy. Arteriolar hyalinosis predicted outcome independent of sample size, but became less important than percentage glomerulosclerosis in predicting outcome if only samples containing more than 25 glomeruli were considered. Interstitial fibrosis/tubular atrophy did not predict the outcome of a kidney, independent of sample size. When comparing baseline with subsequent core biopsies, or with paired baseline biopsies, at least 14 glomeruli were necessary to allow even moderate reproducibility of glomerulosclerosis (Cohen's kappa > 0.25) and to allow statistical significance (P < 0.05). The reproducibility of arteriolar hyalinosis was not dependent on sample size but was reproducible in 80% of paired baseline biopsies, and in 67% of the comparison of the baseline with core biopsy. Both precision and significance was lost if sample numbers were reduced by including only larger samples. There was no reproducibility in any study of interstitial fibrosis/tubular atrophy when comparing either baseline with subsequent biopsy, or paired baseline biopsies. SUMMARY: Much larger biopsy samples are necessary than has generally been assumed in order for glomerulosclerosis rates to be reproducible or predictive of outcome. Arteriolar hyalinosis is prognostically important and shows good reproducibility independent of sample size. Interstitial fibrosis/tubular atrophy appear useless as predictors, being of no prognostic importance and lacking reproducibility. Our finding clarifies some of the discrepancies found by different investigators regarding the importance of renal biopsy in predicting prognosis. Preliminary, our data indicate that samples containing fewer than 25 glomeruli are unreliable in determining outcome based on glomerulosclerosis. The importance of our findings which are based only on chronic lesions, with respect to acute changes, is unknown.

Pathologic features of acute renal allograft rejection associated with donor-specific antibody, Analysis using the Banff grading schema.

Alloantibody frequently appears during the immune response to alloantigens in renal transplant recipients. We studied whether the presence of antibody against donor class I antigens correlated with the clinical and pathologic features of acute rejection episodes. We identified patients who had (1) clinical evidence of acute rejection, (2) a renal biopsy showing pathologic features of acute rejection, defined by the Banff criteria, and (3) pre- and posttransplant sera screened against donor T cells. We divided these patients into those with or without donor-specific alloantibody reactive with donor T cells. Of 44 patients with biopsy-proven rejection, 20 were antibody negative (Ab-R) and 24 were antibody positive (Ab+R). The biopsies from Ab+R patients had a higher incidence of severe vasculitis (P=0.0009) and glomerulitis (P=0.01). Fibrin thrombi in the glomeruli and/or vessels, fibrinoid necrosis, and dilatation of peritubular capillaries were also more frequent in the Ab+R group. Infarction was present in biopsy specimens from 9/24 Ab+R patients versus none in the Ab-R group (P=0.002). The Ab+R biopsy specimens more often had polymorphonuclear leukocytes in the peritubular capillaries (P=0.003). In contrast, specimens of Ab-R patients showed tubulitis more often than the specimens of Ab+R patients: moderate and severe tubulitis was present in 19/20 (95%) Ab-R patients versus 12/24 (50%) Ab+R patients (P=0.002). Graft loss was increased in Ab+R patients, particularly in the first 3 months (12/24 compared with 3/20, P=0.025). Thus, during biopsy-proven acute rejection episodes, anti-class I antibody correlates with severe vascular lesions, glomerulitis, and infarction, whereas more severe tubulitis predominates in rejection episodes without antibody.

Endocapillary glomerulitis in the renal allograft.

Endocapillary glomerulitis is characterized by an increase in number of mononuclear cells in the glomerular capillary lumina. This lesion has been described in the early posttransplant period, but its pathogenesis, relation to conventional rejection, and prognostic impact is not well known. Using the definitions, scorings, and gradings of the Banff system for classification and grading of histopathologic changes in the renal allograft, we have analyzed 444 consecutive renal allograft biopsies from the first 90 days posttransplant. Moderate or severe glomerulitis occurred in 13.5% of the biopsies. There was a strong tendency toward clustering of glomerulitis: if one biopsy from a patient had glomerulitis, there was a high probability that it occurred in other biopsies from the same patient. There was some correlation with conventional acute rejection, but 40% of all biopsies with glomerulitis had no rejection and 53% of all biopsies with rejection had no glomerulitis. Graft function at biopsy was nil or decreased in many patients, but this could largely be explained by the independent presence of primary graft dysfunction or conventional rejection, these conditions being a frequent indication for performing a graft biopsy. Moderate or even severe glomerulitis was, however, compatible with a functioning graft. No correlation between glomerulitis and active CMV infection was found. The one-year graft survival of grafts with early posttransplant glomerulitis was 66%. If early conventional acute rejection is taken into consideration, graft survival does not seem to be influenced by the presence of glomerulitis. Early posttransplant endocapillary glomerulitis may be a peculiar pattern of rejection with a pathogenesis different from that of conventional rejection, but the present investigation does not demonstrate any adverse effects on graft function or graft prognosis.

Post-transplant lymphoproliferative disorder in renal allograft recipients. Clinical experience and risk factor analysis in a single center.

Post-transplant lymphoproliferative disorder (PTLD) is a well-recognized complication of solid organ transplantation. The University of Alberta Renal Transplant Program had not experienced a case of PTLD occurring in the early post-transplant period until March 1989. Since then, 4 patients have developed this complication. To identify the major risk factors for the recent appearance of PTLD, a retrospective analysis was carried out on 162 cadaveric renal transplants performed between July 1987 and December 1990. Four cases of polymorphic PTLD were seen. Two patients presented with fatal disseminated disease. Two others developed PTLD confined to the renal allograft; both are disease free at > 24 months of follow-up. Seventy-two (44.4%) of the cadaveric transplant recipients had received Minnesota antilymphocyte globulin (MALG) induction therapy during the study period. Twenty-four of these also received OKT3 for steroid-resistant rejection. Of the 4 patients with PTLD, 3 had received both MALG induction and OKT3; the remaining patient had received MALG induction only. The incidence of PTLD in the MALG/OKT3 group was 12.5%, which is significantly higher than that of patients receiving other immunosuppressive regimes (0.7%, P = 0.015). The incidence of PTLD was also significantly greater in the 13 patients at risk for primary EBV infection compared to the EBV seropositive patients (23.1 vs. 0.7%, P = 0.002). Only 2 seronegative patients received sequential MALG/OKT3; both developed PTLD. Thus, the population most at risk is that receiving potent antilymphocyte preparations in the setting of primary EBV infection. Allograft involvement with PTLD must be considered in the differential diagnosis of allograft dysfunction, as early diagnosis may permit the successful management of this complication.

Regulation of IFN-gamma and tumor necrosis factor-alpha expression in vivo. Effects of cycloheximide and cyclosporine in normal and lipopolysaccharide-treated mice.

Despite accumulating information about cytokine expression in vitro, relatively little is known about the regulation and biologic relevance of these mediators in vivo. In order to study the effects of inhibition of protein synthesis and cyclosporine in vivo, we made use of systemically administered LPS, which induces the expression of a variety of cytokines. The expression of IFN-gamma and TNF-alpha mRNA in normal and LPS-treated mice was examined by Northern blot analysis and amplification using the polymerase chain reaction. IFN-gamma activity was monitored using the biologic end point of MHC induction. TNF-alpha activity in serum was assessed using a L929 cytotoxicity assay. Messenger RNA for IFN-gamma and TNF-alpha could not be reliably detected by Northern analysis in spleens or kidneys of normal mice. After treatment with cycloheximide, a protein synthesis inhibitor, IFN-gamma and TNF-alpha mRNA could be detected in both sites in otherwise normal mice. The level of both IFN-gamma and TNF-alpha mRNA increased after LPS, although the temporal patterns of expression were different. The concurrent administration of cycloheximide led to marked superinduction of both cytokine mRNA levels. Similar effects were seen in T cell-deficient nude mice, suggesting that these responses are T cell independent. Cyclosporin A blocked induction of IFN-gamma in a dose dependent manner, but failed to significantly inhibit TNF-alpha mRNA or protein expression. Thus at least part of the immunosuppressive effect of cyclosporin A in vivo may be caused by its ability to inhibit the expression of certain cytokine genes, as has been found in vitro systems. However, the cellular target for this effect may extend to cell populations other than T cells.

Local T cell responses induce widespread MHC expression. Evidence that IFN-gamma induces its own expression in remote sites.

Local inflammation induces increased expression of MHC and other genes in the affected tissue because of the paracrine effects of cytokines such as IFN-gamma. We previously reported that one such process--local allograft rejection--was accompanied by increased expression of MHC in a remote tissue, namely kidney. To explore how local inflammation affects gene expression in remote tissues, we studied MHC, beta 2-microglobulin, and IFN-gamma expression in mice undergoing either of two T cell-dependent localized inflammatory processes: rejection of an ascites tumor allograft, and skin sensitization by oxazalone. As assessed by binding of radiolabeled mAb and by immunohistology, each stimulus increased MHC expression in many remote tissues, including liver, heart, pancreas, and kidney. This was associated with increases in steady state mRNA for class I, class II, and beta 2-microglobulin. MHC induction was inhibited by the in vivo administration of cyclosporine or anti-IFN-gamma mAb and did not occur in nude mice, confirming the key role of IFN-gamma released from T cells. When we examined tissues of mice with these localized inflammatory lesions for IFN-gamma mRNA levels by polymerase chain reaction, we found that IFN-gamma steady state mRNA levels were increased in the spleen and, more surprisingly, in the kidney, and in uninvolved skin. Moreover, anti-IFN-gamma inhibited the induction of IFN-gamma mRNA in the kidney, suggesting that IFN-gamma expression was induced by IFN-gamma in an autoregulatory fashion. Thus the systemic MHC induction accompanying local T cell-mediated inflammation reflects the release of IFN-gamma from the site of inflammation, but may be amplified by the ability of IFN-gamma to induce its own expression in remote tissues. This self-amplification of IFN-gamma may contribute to the ability of local inflammation to induce extensive systemic effects.

A view from the groove: peptide binding by MHC molecules and the implications for regional immune responses.

A significant event in immunology occurred in 1987 with the publication of the three dimensional structure of a class I major histocompatibility complex (MHC) product as determined by X ray crystallography. The crystal structure revealed a groove created by the first and second domains of the molecule which could be the antigen (Ag) binding site. Subsequent analyses have suggested that a similar groove exists in the class II molecule. The present view will focus on the implications of this new knowledge for understanding regional immune responses. In particular, we shall discuss how the occupation of the groove by different peptides in different tissues implies that a component of T cell recognition of MHC may be tissue specific.