Acute cellular and antibody-mediated rejection of the pancreas allograft: incidence, risk factors and outcomes.

Antibody-mediated rejection (AMR) after pancreas transplantation is a recently identified entity. We describe the incidence of, risk factors for, and outcomes after AMR, and the correlation of C4d immunostaining and donor-specific antibody (DSA) in the diagnosis of AMR. We retrospectively analyzed 162 pancreas transplants in 159 patients who underwent 94 pancreas allograft biopsies between 2006 and 2009. Univariate and multivariate analyses were performed to evaluate risk factors for pancreas graft AMR. One-year rejection rates and survival after rejection were calculated by Kaplan-Meier methods. AMR occurred in 10% of patients by 1-year posttransplant. Multivariate risk factors identified for AMR include nonprimary simultaneous pancreas-kidney (SPK) transplant, primary solitary pancreas (PAN) transplant and race mismatch. After pancreas rejection, patient survival was 100% but 20% (8 of 41) of pancreas grafts failed within 1 year. Graft survival after acute cellular rejection (ACR), AMR and mixed rejection was similar. Of biopsies that stained >5% C4d, 80% were associated with increased Class I DSA. In summary, AMR occurs at a measurable rate after pancreas transplantation, and the diagnosis should be actively sought using C4d staining and DSA levels in patients with graft dysfunction, especially after nonprimary SPK and primary PAN transplantation.

Nox2 is a mediator of chronic CsA nephrotoxicity.

We hypothesized that Nox2, the classical phagocytic NADPH oxidase, plays an important role in calcineurin inhibitor (CNI)-induced renal fibrosis. We tested this hypothesis in vitro, in animal and in human studies. Cyclosporine A (CsA) and tacrolimus (TAC) were associated with greater levels of Nox2 mRNA and epithelial to mesenchymal transition (EMT) in NRK52E cells. CsA increased Nox2, α-SMA and phosphorylated-p38MAPK, Smad3 and NFκB proteins. Nox2 upregulation and EMT were inhibited in TGF-ß1 knockout cells suggesting that TGF-ß1 is required for Nox2 activation. Fisher344 rats treated with high dose CsA showed increased Nox2 in the tubulointerstitium and greater Nox2, α-SMA, phosphorylated Smad3 and nitrotyrosine by immunoblot analyses. Inhibition of Nox2 by coadministration of apocynin or diphenyleneiodonium was associated with reduced fibrogenesis. We validated these findings by treating wild type and Nox2 null (B6.129S-Cybb(Tm1Din)/J) mice with high dose CsA. Western blot analyses confirmed the absence of Nox2 and significantly lower levels of α-SMA and 4-hydroxynonenal (HNE) in CsA-treated knockout mice. These findings were clinically relevant since Nox2 and α-SMA were increased in the tubulointerstitium of kidneys from 15 liver transplant recipients with biopsy-confirmed chronic CsA or TAC nephrotoxicity. In conclusion, specific Nox2 inhibition strategies may improve chronic CNI nephrotoxicity in solid organ transplantation.

Guidelines for the diagnosis of antibody-mediated rejection in pancreas allografts-updated Banff grading schema.

The first Banff proposal for the diagnosis of pancreas rejection (Am J Transplant 2008; 8: 237) dealt primarily with the diagnosis of acute T-cell-mediated rejection (ACMR), while only tentatively addressing issues pertaining to antibody-mediated rejection (AMR). This document presents comprehensive guidelines for the diagnosis of AMR, first proposed at the 10th Banff Conference on Allograft Pathology and refined by a broad-based multidisciplinary panel. Pancreatic AMR is best identified by a combination of serological and immunohistopathological findings consisting of (i) identification of circulating donor-specific antibodies, and histopathological data including (ii) morphological evidence of microvascular tissue injury and (iii) C4d staining in interacinar capillaries. Acute AMR is diagnosed conclusively if these three elements are present, whereas a diagnosis of suspicious for AMR is rendered if only two elements are identified. The identification of only one diagnostic element is not sufficient for the diagnosis of AMR but should prompt heightened clinical vigilance. AMR and ACMR may coexist, and should be recognized and graded independently. This proposal is based on our current knowledge of the pathogenesis of pancreas rejection and currently available tools for diagnosis. A systematized clinicopathological approach to AMR is essential for the development and assessment of much needed therapeutic interventions.

Pretransplant immune-regulation predicts allograft tolerance.

CD4⁺ Tregs specific for noninherited maternal antigens (NIMA(d) ) are detectable in some but not all B6 × BDF1 backcross, H-2(b) homozygous offspring, and their presence is strongly correlated with extent of maternal (BDF1) microchimerism. We hypothesized that the level of pretransplant donor antigen-specific Tregs could predict allograft tolerance. To test this idea, mice were screened for bystander suppression in a DTH assay, followed 1 week later by DBA/2 heterotopic heart transplantation. NIMA(d) -exposed, H-2(b) offspring that failed to suppress DTH uniformly rejected heart allografts (12/12) by d15. In contrast, 5/6 NIMA(d) -exposed DTH 'regulators' accepted their allografts >100 days. The defect in 'nonregulator" offspring could be corrected by transfer of CD4⁺CD25⁺, but not CD4⁺ CD25(neg) or CD8⁺ T cells from transplant acceptor mice. In conclusion, donor-specific T reg screening of F1 backcross offspring correctly predicted which recipients would accept a heart allograft. If translated to the clinic, similar pretransplant Treg screening could greatly enhance the effectiveness of tolerance as a clinical strategy in transplantation between family members.

Early and limited use of tacrolimus to avoid rejection in an alemtuzumab and sirolimus regimen for kidney transplantation: clinical results and immune monitoring.

Alemtuzumab induction with 60 days of tacrolimus treatment and continuous sirolimus treatment prevented acute rejection in nine of 10 consecutive renal allograft recipients. All patients are alive with a functioning kidney graft at 27-39 months of follow-up. Extensive immune monitoring was performed in all patients. Alloantibody detection, cytokine kinetics assay (CKA), and trans vivo delayed-type hypersensitivity (DTH) assay were performed every 6 months showing correlation with clinical evolution. Despite alloantibody presence in five patients, eight patients remain without the need for specific treatment and only sirolimus monotherapy in decreasing dosage. Four patients take only 1 mg sirolimus daily with levels of 3-4 ng/mL. One patient showed clinical signs of rejection at month 9 post-transplant, with slow increase in serum creatinine and histological signs of mixed cellular (endarteritis) and humoral rejection (C4d positivity in peritubular capillaries and donor-specific antibody (DSA)). In summary, the addition of tacrolimus therapy for 2 months to a steroid-free, alemtuzumab induction and sirolimus maintenance protocol limited the previously shown acute rejection development. Nevertheless, alloantibody was present in serum and/or C4d present on 1-year biopsy in half the patients. The combination of CKA and DSA monitoring or the performance of transvivo DTH correlated with immune status of the patients.

Banff schema for grading pancreas allograft rejection: working proposal by a multi-disciplinary international consensus panel.

Accurate diagnosis and grading of rejection and other pathological processes are of paramount importance to guide therapeutic interventions in patients with pancreas allograft dysfunction. A multi-disciplinary panel of pathologists, surgeons and nephrologists was convened for the purpose of developing a consensus document delineating the histopathological features for diagnosis and grading of rejection in pancreas transplant biopsies. Based on the available published data and the collective experience, criteria for the diagnosis of acute cell-mediated allograft rejection (ACMR) were established. Three severity grades (I/mild, II/moderate and III/severe) were defined based on lesions known to be more or less responsive to treatment and associated with better- or worse-graft outcomes, respectively. The features of chronic rejection/graft sclerosis were reassessed, and three histological stages were established. Tentative criteria for the diagnosis of antibody-mediated rejection were also characterized, in anticipation of future studies that ought to provide more information on this process. Criteria for needle core biopsy adequacy and guidelines for pathology reporting were also defined. The availability of a simple, reproducible, clinically relevant and internationally accepted schema for grading rejection should improve the level of diagnostic accuracy and facilitate communication between all parties involved in the care of pancreas transplant recipients.