Histological Evolution of BK Virus-Associated Nephropathy: Importance of Integrating Clinical and Pathological Findings.

Long-term clinicopathological studies of BK-associated nephropathy (PyVAN) are not available. We studied 206 biopsies (71 patients), followed 3.09 ± 1.46 years after immunosuppression reduction. The biopsy features (% immunostain for PyV large T ag + staining and inflammation ± acute rejection) were correlated with viral load dynamics and serum creatinine to define the clinicopathological status (PyVCPS). Incidence of acute rejection was 28% in the second biopsy and 50% subsequently (25% mixed T cell-mediated allograft rejection (TCMR) + antibody-mediated allograft rejection (AMR); rejection overall affected 38% of patients (>50% AMR). Graft loss was 15.4% (0.8-5.3 years after PyVAN); 76% had complete viral clearance (mean 28 weeks). The only predictors of graft loss were acute rejection (TCMR p = 0.008, any type p = 0.07), and increased "t" and "ci" in the second biopsy (p = 0.006 and 0.048). Higher peak viremia correlated with poorer viral clearance (p = 0.002). Presumptive and proven PyVAN had similar presentation, evolution, and outcome. Late PyVAN (>2 years, 9.8%) justifies BK viremia evaluation at any point with graft dysfunction and/or biopsy evaluation. This study describes the histological evolution of PyVAN and corresponding clinicopathological correlations. Although the pathological features overall reflect the viral and immunological interactions, the PyVAN course remains difficult to predict based on any single feature. Appropriate clinical management requires repeat biopsies and determination of the PyVCPS at relevant time points, for corresponding personalized immunosuppression adjustment.

BK Polyomavirus Infection and Renourinary Tumorigenesis.

BK polyomavirus (BKPyV) infection represents a major problem in transplantation, particularly for renal recipients developing polyomavirus-associated nephropathy (PyVAN). The possibility that BKPyV may also be oncogenic is not routinely considered. Twenty high-grade renourinary tumors expressing polyomavirus large T antigen in the entirety of the neoplasm in 19 cases, including the metastases in six, have been reported in transplant recipients with a history of PyVAN or evidence of BKPyV infection. Morphological and phenotypical features consistent with inactivation of the tumor suppressors pRB and p53 were found in the bladder tumors, suggesting a carcinogenesis mechanism involving the BKPyV large tumor oncoprotein/antigen. The pathogenesis of these tumors is unclear, but given the generally long interval between transplantation and tumor development, the risk for neoplasms after BKPyV infections may well be multifactorial. Other elements potentially implicated include exposure to additional exogenous carcinogens, further viral mutations, and cell genomic instability secondary to viral integration, as occurs with the Merkel cell PyV-associated carcinoma. The still scarce but increasingly reported association between longstanding PyVAN and renourinary neoplasms requires a concerted effort from the transplant community to better understand, diagnose, and treat the putative association between the BKPyV and these neoplasms.

Concurrent cytomegalovirus glomerulitis and BK polyomavirus-associated nephropathy in a kidney allograft biopsy.

A 58-year-old renal transplant recipient underwent biopsy 11 weeks post transplantation for increasing creatinine. The biopsy showed cytomegalovirus (CMV) glomerulitis together with BK polyomavirus (BKPyV)-associated nephropathy (PVAN). Treatment with intravenous ganciclovir and overall reduction in maintenance immunosuppression resulted in prompt resolution of the CMV glomerulitis, but with persistence of PVAN in a follow-up biopsy 4 weeks later. Stable creatinine and BKPyV viral clearance were observed at the last clinical visit 15 months post transplantation. This case exemplifies infectious glomerulitis, which requires differentiation from the more common glomerulitis caused by antibody-mediated allograft rejection. The morphological similarities and differences between BKPyV and CMV infections are discussed.

Islet Amyloid in Whole Pancreas Transplants for Type 1 Diabetes Mellitus (DM): Possible Role of Type 2 DM for Graft Failure.

Long-term results with whole pancreas (WPTx) and islet transplantation (IT) continue to be suboptimal. Graft failure with undetectable C-peptide level is attributed to graft sclerosis (chronic rejection), recurrence of Type 1 diabetes mellitus (DM), or insufficient islet mass. In contrast, graft failure with measurable C-peptide has overlapping clinical features with Type 2 DM (suggesting persistent but insufficient ß cell function), but is poorly understood. In general, the morphological substrate for islet failure is unclear because grafted islets are not routinely evaluated. We present two patients with graft failure at 5 and 8 years after successful WPTx for Type 1 DM, presenting with preserved C-peptide levels. On histopathology, the islets had preserved both α and ß cell populations but also prominent accumulation of islet amyloid (IA), the morphological hallmark of Type 2 DM. IA previously reported in IT, represents fibrillary aggregates of islet amyloid polypeptide, a hormone normally cosecreted with insulin. Accumulation of IA correlates quantitatively with the development of hyperglycemia and is known to cause ß cell dysfunction and loss. Accumulation of IA and development of Type 2 DM should be considered and studied as a potential cause of long-term islet failure in IT and WPTx.

Bartonella henselae infection-associated vasculitis and crescentic glomerulonephritis leading to renal allograft loss.

Bartonella henselae (BH) is the main cause of cat scratch disease (CSD), which more typically presents as a self-limited localized suppurative lymphadenopathy in immunocompetent individuals. In contrast, immunocompromised patients commonly have systemic disease with life-threatening complications. In addition to the angioproliferative lesions, such as bacillary angiomatosis, an increasing number of immune post-infectious complications are being recognized with BH infections, including glomerulonephritis, vasculitis, hemophagocytic syndrome, and neurological problems. We report the case of a renal transplant recipient who developed CSD in the second year post transplantation. In addition to prolonged fever and generalized lymphadenopathy and splenomegaly requiring differentiation from a post-transplant lymphoproliferative disorder, the course was complicated by the development of dermal leukocytoclastic vasculitis and pauci-immune necrotizing and crescentic glomerulonephritis, which led to failure of the renal graft. Glomerulonephritis as a complication of CSD has never been described in a kidney allograft, to our knowledge. Awareness of the diverse clinical symptoms associated with BH, including granulomatous/suppurative lesions and other less common complications can lead to more rapid and accurate diagnosis. Also, as recommended by the current guidelines, a thorough history of pet ownership should be part of the clinical evaluation before and after transplantation for all transplant recipients.

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.

Banff '09 meeting report: antibody mediated graft deterioration and implementation of Banff working groups.

The 10th Banff Conference on Allograft Pathology was held in Banff, Canada from August 9 to 14, 2009. A total of 263 transplant clinicians, pathologists, surgeons, immunologists and researchers discussed several aspects of solid organ transplants with a special focus on antibody mediated graft injury. The willingness of the Banff process to adapt continuously in response to new research and improve potential weaknesses, led to the implementation of six working groups on the following areas: isolated v-lesion, fibrosis scoring, glomerular lesions, molecular pathology, polyomavirus nephropathy and quality assurance. Banff working groups will conduct multicenter trials to evaluate the clinical relevance, practical feasibility and reproducibility of potential changes to the Banff classification. There were also sessions on quality improvement in biopsy reading and utilization of virtual microscopy for maintaining competence in transplant biopsy interpretation. In addition, compelling molecular research data led to the discussion of incorporation of omics-technologies and discovery of new tissue markers with the goal of combining histopathology and molecular parameters within the Banff working classification in the near future.

The Maryland aggregate pathology index: a deceased donor kidney biopsy scoring system for predicting graft failure.

Despite the common use of diagnostic pretransplant deceased donor kidney biopsy, there is no consensus on the prognostic significance of the pathologic findings. In order to assist clinicians with interpretation we analyzed 371 pretransplant biopsies and correlated the findings with graft failure. Glomerular pathology was assessed with percent glomerulosclerosis (GS), glomerular size and periglomerular fibrosis (PGF); vascular pathology with arterial wall-to-lumen ratio (WLR) and arteriolar hyalinosis and interstitial pathology with measurement of cumulative fibrosis and presence of scar. Using two-thirds of the study population as a model-development cohort, we found that biopsy features independently associated with an increased risk of graft failure were GS > or =15%, interlobular arterial WLR > or =0.5 and the presence of PGF, arteriolar hyalinosis or scar. The Maryland Aggregate Pathology Index (MAPI), was developed from these parameters and validated on the remaining one-third of the population. Five-year actuarial graft survival was 90% for kidneys with MAPI scores between 0 and 7, 63% for scores from 8 to 11 and 53% for scores from 12 to 15 (p < 0.001). We conclude MAPI may help transplant physicians estimate graft survival from the preimplantation biopsy findings, in clinical situations similar to this study population (cold ischemia over 24 h, GS < 25%).

Banff 07 classification of renal allograft pathology: updates and future directions.

The 9th Banff Conference on Allograft Pathology was held in La Coruna, Spain on June 23-29, 2007. A total of 235 pathologists, clinicians and scientists met to address unsolved issues in transplantation and adapt the Banff schema for renal allograft rejection in response to emerging data and technologies. The outcome of the consensus discussions on renal pathology is provided in this article. Major updates from the 2007 Banff Conference were: inclusion of peritubular capillaritis grading, C4d scoring, interpretation of C4d deposition without morphological evidence of active rejection, application of the Banff criteria to zero-time and protocol biopsies and introduction of a new scoring for total interstitial inflammation (ti-score). In addition, emerging research data led to the establishment of collaborative working groups addressing issues like isolated 'v' lesion and incorporation of omics-technologies, paving the way for future combination of graft biopsy and molecular parameters within the Banff process.

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.

Role of perforin in controlling B-cell hyperactivity and humoral autoimmunity.

To determine the role of perforin-mediated cytotoxic T lymphocyte (CTL) effector function in immune regulation, we studied a well-characterized mouse model of graft-versus-host disease (GVHD). Induction of acute GVHD using perforin-deficient donor T cells (pfp-->F1) initially resulted in features of acute GVHD, e.g., engraftment of both donor CD4(+) and CD8(+) T cells, upregulation of Fas and FasL, production of antihost CTL, and secretion of both Th1 and Th2 cytokines. Despite fully functional FasL activity, pfp donor cells failed to totally eliminate host B cells, and, by 4 weeks of disease, cytokine production in pfp-->F1 mice had polarized to a Th2 response. Pfp-->F1 mice eventually developed features of chronic GVHD, such as increased numbers of B cells, persistence of donor CD4 T cells, autoantibody production, and lupuslike renal disease. We conclude that in the setting of B- and T-cell activation, perforin plays an important immunoregulatory role in the prevention of humoral autoimmunity through the elimination of both autoreactive B cells and ag-specific T cells. Moreover, an ineffective initial CTL response can evolve into a persistent antibody-mediated response and, with it, the potential for sustained humoral autoimmunity.

Spectrum of histopathological changes in pancreas allograft biopsies and relationship to graft loss.

Histological evaluation of pancreas allografts through the use of needle biopsies is of paramount importance for the determination of the etiology of graft dysfunction. In addition, pathological assessment of the overall status of the exocrine, endocrine, and vascular components provides invaluable information with regards to the prognosis of the graft. Pancreas allograft failure results from a variety of causes, highly dependent on the time posttransplantation, but after the first 6 months' posttransplantation the most common cause of graft loss is chronic rejection. The main histological manifestations of chronic rejection are progressive graft sclerosis (increasing fibrosis and proportional atrophy of the glandular components), secondarily leading to endocrine failure. Evaluation of serial biopsies in patients with graft failure has shown that the most important histological predictors of chronic rejection/graft sclerosis are diffuse acinar inflammation and acute and chronic vascular injury in the form of intimal arteritis and proliferative transplant arteriopathy, respectively.

Spontaneous formation of germinal centers in autoimmune mice.

The mechanisms of autoantibody production are not well understood. Germinal centers (GC) may be important sites of immune disregulation in autoimmune diseases. In this study, we document the presence of spontaneous GC formation in the spleens of several autoimmune mouse strains that spontaneously develop autoimmune Type I diabetes and a lupus-like disease. In contrast, mouse strains that do not develop lupus did not exhibit spontaneous formation of GC. In all of the autoimmune strains studied, GC were present at 1-2 months of age, a time that closely parallels the appearance of autoantibodies. Like the GC that develop after purposeful immunization, GC in autoimmune mice contained B220(+), PNA(+), and GL-7(+) B cells, and FDC-M1(+) follicular dendritic cells. In addition, spontaneously formed GC in autoimmunity and those caused by immunization were abrogated in a similar way by a short-term treatment with anti-CD40 ligand antibody. These data indicate that spontaneously forming GC in autoimmunity are similar to those appearing after purposeful immunization.

Inhibitors of membrane lipid metabolism enhance complement-mediated nucleated cell killing through distinct mechanisms.

The ability of nucleated cells to survive limited complement attack has been attributed to metabolic processes unique to these cells, such as rapid elimination of terminal complement complexes (TCC) from their surfaces. The biochemical processes activated by complement channels responsible for cell defense remain poorly defined. Metabolic inhibitors affecting membrane lipid turnover have been shown to increase the complement-mediated cell death. Whether these metabolic inhibitors increase lytic susceptibility of target cells by reducing the rate of TCC elimination has not been previously evaluated. In the present study, inhibitors of membrane lipid transmethylation and lysolecithin reacylation were evaluated in view of the observations that TCC concurrently increase lipid transmethylation and inhibit lysolecithin reacylation, and the inhibition of lipid transmethylation correlates with increased complement-mediated cell death. We have measured the formation as well as the elimination of C5b-9 on the target membrane that affect the outcome of cell death. Our results in the present communication indicated that inhibitors of transmethylation and lysolecithin reacylation increased TCC-mediated cell death through distinct pathways, the former by allowing more efficient deposition of TCC, and the latter by impairing TCC elimination.

Effect of osmotic protection on nucleated cell killing by C5b-9: cell death is not affected by the prevention of cell swelling.

Formation of C5b-9 channels in the plasma membrane can lead to erythrocyte lysis or nucleated cell death. Lysis of erythrocytes by complement occurs as a result of colloid osmotic swelling and rupture of the plasma membrane, due to the unregulated flux of ions and water through C5b-9 channels. This colloid osmotic mechanism of lysis is largely based on the evidence that the extent of hemolysis is reduced, when macromolecules are placed in the medium to balance the osmotic gradient created by intracellular macromolecules, which are too large to diffuse through complement channels. The role of colloid osmotic deregulation, as a cause of nucleated cell killing by C5b-9, however, has been recently questioned [Kim S., Carney D. F. and Shin M. L. J. Immun. 138, 1530 (1987)]. In the present study, we investigated the effect of osmotic protection, with an 81,000 mol. wt dextran or bovine serum albumin, on Ehrlich cell killing by complement channels. The results indicated that prevention of cell swelling by dextran did not reduce the extent or rate of nucleated cell killing by either small (C5b-9l), or large (C5b-9m), complement channels when assessed by vital dye stain. The release of cytoplasmic lactate dehydrogenase as an alternative measure of cell death, however, was retarded and/or reduced, in the presence of dextran or albumin, at concns that prevented cell swelling. These results indicate that C5b-9 can kill nucleated cells effectively, in the absence of colloidal osmotic cell swelling, and that release of cytoplasmic macromolecules may not be a reliable indicator of cell death, when osmotic protectants are employed.

Effects of Ca2+ deregulation on mitochondrial membrane potential and cell viability in nucleated cells following lytic complement attack.

We have previously shown [Papadimitriou JC. Ramm LE. Drachenberg CB. Trump BF. Shin ML. (1991) J. Immunol., 147, 212-217] that formation of lytic C5b-9 channels on Ehrlich ascites tumor cells induced rapid depletion of adenine nucleotides associated with prelytic leakage preceding cell death. Extracellular Ca2+ concentration ([Ca2+]e) reduction by chelation markedly delayed the onset of cell death, although the adenine nucleotide leakage was enhanced. In the present study, we examined the temporal relationships between ionized cytosolic Ca2+ ([Ca2+]i), mitochondrial membrane potential (delta psi m) and cell death in individual cells by digital imaging fluorescence microscopy (DIFM), during the earliest phase of C5b-9 attack. The results showed an immediate, > 20-fold rise in [Ca2+]i, rapidly followed by dissipation of delta psi m and subsequent acute cell death. These events were markedly delayed by chelation of Ca2+e, but not by nominally Ca2+ free medium. Differing from previous reports indicating propidium iodide labeling of viable cells bearing C5b-9 channels, with DIFM we observed nuclear fluorescence with that marker only in association with cell death. These findings indicate that Ca2+ influx through lytic C5b-9 channels is responsible for the massive increase in [Ca2+]i, as well as for the rapid loss of delta psi m, followed by acute cell death. When this [Ca2+]i increase is prevented, the cell death is probably related to metabolic depletion.

Differential diagnosis between monomorphic clear cell adenocarcinoma of salivary glands and renal (clear) cell carcinoma.

Clear cell adenocarcinoma of salivary glands (CCASG) is a relatively rare tumor, composed entirely of clear cells of putative ductal origin. It bears striking morphologic similarities to renal cell carcinoma (RCC) of clear cell type on hematoxylin and eosin stains. Differentiation between CCASG and metastatic RCC to the salivary glands has been considered problematic or even impossible on morphologic grounds. We examined three cases of CCASG and 12 cases of RCC (6 primary and 6 metastatic) by hematoxylin and eosin staining, immunohistochemistry, and electron microscopy. Two distinctive immunohistochemical and ultrastructural patterns emerged from this analysis. CCASG showed positivity for high molecular weight cytokeratin and carcinoembryonic antigen and ultrastructurally showed prominent squamoid differentiation, glycogen pools, and absence of lipid. In contrast, RCC was characterized by positivity for vimentin and complete absence of staining for high molecular weight cytokeratin and carcinoembryonic antigen. On ultrastructural studies, RCC lacked any squamoid differentiation, and the tumor cells contained abundant cytoplasmic lipid in addition to glycogen. Thus, based on the consistent differences on the immunohistochemical staining patterns and their characteristic subcellular morphology, CCASG and RCC can be distinguished on pathologic evaluation. The different direction of differentiation of the cells in CCASG and RCC (i.e., ductal in the former and renal tubular and mesodermal in the latter) results in their distinctive immunophenotypical and ultrastructural features.

CMV allograft pancreatitis: diagnosis, treatment, and histological features.

BACKGROUND: Cytomegalovirus (CMV) infection is a common problem in solid organ transplant recipients. CMV infection of pancreas allografts is not, however, well described. METHODS: We report the clinical presentation, histologic findings, treatment, and outcome in four patients with CMV allograft pancreatitis. These patients presented 18 weeks to 44 months after transplantation with elevated serum amylase and lipase and were suspected to have acute rejection. Percutaneous pancreas allograft biopsy specimens showed evidence of tissue invasive CMV infection. One patient had simultaneous CMV infection and acute rejection. RESULTS: Prolonged treatment with ganciclovir resulted in clinical and histologic resolution of the CMV disease. Rejection was successfully treated. Primary CMV infection in seronegative recipients seemed to be a risk factor. Three patients maintain normal allograft function; one patient lost function due to chronic rejection. The histology of tissue-invasive CMV pancreas allograft infection and its differentiation from acute rejection is described. CONCLUSION: Prompt diagnosis and prolonged therapy with antiviral agents can result in maintenance of allograft function.

Histologic grading of acute allograft rejection in pancreas needle biopsy: correlation to serum enzymes, glycemia, and response to immunosuppressive treatment.

BACKGROUND: Allograft rejection continues to be the most common cause of graft failure in technically successful pancreas transplants. Early diagnosis and treatment of rejection is essential for long-term graft survival. Pancreas graft biopsies are now used routinely for the diagnosis of acute allograft rejection. The correlation between clinical evidence of graft dysfunction (increased serum enzymes and glucose), severity of acute rejection on biopsy (rejection grade), and response to treatment has not been previously studied. METHODS: A total of 151 pancreas transplant needle biopsy specimens from 57 patients were evaluated. Statistical correlation was done between the histologic rejection grade (O-V) and the peak level of enzymes in serum, glycemia, type of antirejection treatment instituted, and response to treatment. Differentiation between grades was also evaluated statistically. RESULTS: Response to antirejection treatment was 25%, 40%, 88%, 78%, 50%, and 17% for grades O-V, respectively. The response for grades II and III was better than for grades 0-I and IV-V (P=0.0003 and 0.0008, respectively). The response to corticosteroids alone was 36%, 86%, 68%, and 0% for grades I, II, III, and IV, respectively. The response to antilymphocyte regimen was 50%, 89%, 85%, 71%, and 17% for grades I, II, III, IV, and V, respectively. Overall correlation between the mean levels of enzymes and rejection grade was seen; the increase of lipase was statistically significant (r=0.24, P=0.012). Amylase and lipase correlated very well with each other (r=0.84, P=0.0001). No correlation was found in the mean values of blood glucose with the serum enzyme increase and with severity of rejection. Hyperglycemia was present in 12 patients; this abnormality in patients with grades II-IV responded promptly to treatment, whereas in patients with grade V, hyperglycemia persisted despite antirejection treatment. Other causes of increased enzymes were found in patients with biopsy specimens showing no rejection (grades 0 and I, 43% and 31%, respectively). CONCLUSIONS: Increased serum enzymes, particularly lipase, correlate with the grade of acute rejection, but their lack of specificity precludes their use as sole markers of acute rejection. Glucose levels are not a sensitive marker for acute rejection. Rejection grades II and III are the most responsive to treatment, and a significant proportion of these cases respond to treatment with corticosteroids only. The higher rejection grades (IV and V) require treatment with antilymphocytic regimens, and their overall response to treatment is moderate to poor, respectively.