Pathology of resolving polyomavirus-associated nephropathy.

Control of polyomavirus BK (BKV) is achieved by reducing immunosuppression allowing an effective BKV-specific T-cell response. The morphology of resolving BKV-associated nephropathy (PyVAN) has not been systematically investigated. Ninety-nine surveillance biopsies of 35 patients with BKV viremia treated exclusively by immunosuppression reduction were scored according to Banff criteria and grouped relative to BKV viremia as pre-, increasing, decreasing and post-BKV viremia. Thirty-four of 35 patients (97%) cleared BKV viremia after a median of 9 months posttransplantation. The tubulitis score, extent of tubules with intraepithelial lymphocytes, and interstitial inflammation significantly increased from the time of increasing to decreasing viremia. Tubulointerstitial inflammation, to a lower extent, persisted after clearance. The number of SV40+ tubules correlated with the BKV load in plasma, but SV40 immunohistochemistry was frequently negative (60%). During decreasing viremia, 31% of PyVAN cases were plasma cell-rich and 40% showed tubular HLA-DR expression. Compared to baseline 1 month posttransplantation, allograft function remained stable or improved in 29/35 patients (83%) after a median follow-up of 48 months. Within 1 year after clearance of BKV viremia, clinical rejection occurred in 2/35 patients (6%). Our data suggest that resolving PyVAN is typically characterized by a self-limiting acute interstitial nephritis, morphologically indistinguishable from interstitial rejection.

High incidence of rejection episodes and poor tolerance of sirolimus in a protocol with early steroid withdrawal and calcineurin inhibitor-free maintenance therapy in renal transplantation: experiences of a randomized prospective single-center study.

Immunosuppressive maintenance therapy after kidney transplantation leads to various undesired side effects such as calcineurin inhibitor (CNI)-associated nephrotoxicity or elevated cardiovascular risk due to posttransplantation diabetes and hypertension. These effects show negative impacts on long-term allograft function as well as patient morbidity and mortality. Therefore, we used an immunosuppressive regimen with early corticosteroid withdrawal (ESW), maintenance therapy containing tacrolimus, sirolimus (SRL), and mycophenolate sodium for 3 months followed by a prospective randomized trial comparing a CNI free versus a low-dose CNI therapy. The primary endpoint was 6-month graft function. Among 75 patients, ESW was performed after 4 days in 65 patients. Over the following 3 months before randomization to CNI-free maintenance therapy, we experienced a high number (25%) of SRL discontinuations due to adverse events, including leukopenia, anemia, arthritis, and pneumonitis. In addition there were significantly more allograft rejection episodes in the CNI-free group (P = .017) during the study period leading to a switch from SRL to a CNI. Despite the higher rate of rejection episodes in the CNI-free groups, glomerular filtration rates (GFR) at 6 months were comparable between the study groups (P = .25). After 1 year only 9.2% (6/65) of all patients treated with SRL remained on this drug. Conclusion, there was an unacceptably high rate of SRL intolerance using an ESW and CNI-free immunosuppressive regimen combined with a significantly higher rate of rejection episodes.

Polyomavirus nephropathy: a brief review with special emphasis on clinico-patholgical aspects.

From 1995 Polyomavirus (PyV) nephropathy (PVN) has played an important role in solid organ transplant recipients. The disease is caused by a DNA virus, usually the BK variant, more rarely JC virus. In immune incompetent patients either latent endogenous virus is reactivated, or donated virus can multiply. The frequency of PVN nephropathy (previously 10% or higher) is declining. The disease follows a stepwise course: viruria, viraemia, nephropathy. Nephropathy usually manifests itself during the first year after transplantation. The disease remains clinically silent for long periods, later progressive loss of renal function and renal failure occur. A major risk factor is therapy with potent immune suppressive agents. Morphologically, viral replication produces nuclear inclusions and necrosis, predominantly in the urothelium and tubular epithelium. Inflammation (T and B lymphocytes, monocytes/macrophages and granulocytes) accompanies necrosis. Progression is marked by tubular atrophy, interstitial fibrosis and transplant loss. The virus can be detected by the electron microscope and, better, by immunohistology (preferentially mAb against SV40 Large T antigen). It is often hard to differentiate PVN from an interstitial cellular rejection reaction (Banff 1 A/B). As no effective drug treatment exists, the disease must be diagnosed as early as possible and immune suppression reduced. Screening for polyomavirus reactivation is best done stepwise: search for urinary "Decoy cells" (PyV infected cells), PCR for PyV in the blood and in the case of reduced renal function, renal biopsy. Compliance with a stringent screening algorithm allows early detection and adequate treatment and prevents organ loss.

Is monitoring of FOXP3 Treg cells in renal transplants during acute cellular rejection episodes useful?

BACKGROUND: The FOXP3 (forkhead Box p3) transcription factor is a marker for T regulatory cells (Treg). During cellular immune responses, Treg are expected to increase in number to ultimately control and limit this response. In renal transplants massive infiltration by T cells is often seen during rejection crises. This prompted us to examine changes in the numbers of FOXP3 positive T cells accompanying acute cellular rejection events. METHODS: A total of 32 transplant biopsies from 23 patients were studied retrospectively, these 16 protocol biopsies and 16 biopsies taken during rejection episodes included 9 serial pairs (protocol-rejection). To quantify FOXP3 positive T cells, frozen sections were double immunostained with anti-CD3 and anti-FOXP3 antibodies. Areas revealing T cell infiltrates were measured morphometrically and the number of FOXP3 positive cells per 1,000 µm2 of CD3 positive cells was taken as an FOXP3 index. RESULTS: This index was 0.46 (median, range 0.00-1.00) in the 16 protocol biopsies and 0.48 (median, range 0.16-2.31) in rejection episode biopsies. The highest values were seen during rejection crises, exceeding 1.00 in 6/16 biopsies, whereas no protocol biopsies had values greater than 1.00 (0/16) (difference significant p<0.02). In serial biopsies no consistent behavior was observed; the FOXP3 index remained unchanged, fell slightly or rose to a maximum of 13 fold. Expression levels of FOXP3 could vary within weeks. No correlations were found between donor type, initial therapy, therapy at biopsy, serum creatinine at the time of biopsy, at 3 months or 1 year later, and any of the morphometric parameters (CD3 and FOXP3) studied. CONCLUSIONS: During rejection of renal allografts the fraction of FOXP3+ Treg cells within the infiltrating T-cell population can increase transiently. This phenomenon was not consistently seen in acute cellular rejection and the information does not appear to be of value for individual patient management in such cases.

Reducing immunosuppression preserves allograft function in presumptive and definitive polyomavirus-associated nephropathy.

Early detection of polyomavirus BK (BKV) viremia and reduction of immunosuppression is recommended for preventing polyomavirus-associated nephropathy (PyVAN), but systematic histological evaluations were not performed in previous studies. We routinely screen for decoy cells and, if positive, measure plasma BKV-loads. In a cohort of 203 consecutive renal transplantations performed from 2005-2008, 38 patients (19%) developed BKV-viremia and were treated with reduction of immunosuppression. Based on subsequent allograft biopsy results and peak BKV-viremia, patients were assigned to three groups: (i) definitive PyVAN (n = 13), (ii) presumptive PyVAN defined by plasma BKV-loads of ≥ 4 log(10) copies/ml (n = 17) and (iii) low BKV-viremia (n = 8). Clearance of BKV-viremia was achieved in 35/38 patients (92%) and subsequent clinical rejection occurred in 3/35 patients (8.6%), both without any difference among the groups. Patients with definitive PyVAN had higher peak plasma BKV-loads and required longer time for clearance (8.8 vs. 4.6 vs. 2.9 months; p = 0.001). However, allograft function remained stable from baseline to last follow-up at 34 months (range 18-60) in all three groups with median serum creatinine of 1.6 mg/dl, 1.6 mg/dl and 1.3 mg/dl, respectively. We conclude that screening for BKV-replication and reduction of immunosuppression is an effective strategy to preserve medium-term allograft function even in patients developing definitive PyVAN.

Efficacy of induction therapy with ATG and intravenous immunoglobulins in patients with low-level donor-specific HLA-antibodies.

Low-level donor-specific HLA-antibodies (HLA-DSA) (i.e. detectable by single-antigen flow beads, but negative by complement-dependent cytotoxicity crossmatch) represent a risk factor for early allograft rejection. The short-term efficacy of an induction regimen consisting of polyclonal anti-T-lymphocyte globulin (ATG) and intravenous immunoglobulins (IvIg) in patients with low-level HLA-DSA is unknown. In this study, we compared 67 patients with low-level HLA-DSA not having received ATG/IvIg induction (historic control) with 37 patients, who received ATG/IvIg induction. The two groups were equal regarding retransplants, HLA-matches, number and class of HLA-DSA. The overall incidence of clinical/subclinical antibody-mediated rejection (AMR) was lower in the ATG/IvIg than in the historic control group (38% vs. 55%; p = 0.03). This was driven by a significantly lower rate of clinical AMR (11% vs. 46%; p = 0.0002). Clinical T-cell-mediated rejection (TCR) was significantly lower in the ATG/IvIg than in the historic control group (0% vs. 50%; p < 0.0001). Within the first year, allograft loss due to AMR occurred in 7.5% in the historic control and in 0% in the ATG/IvIg group. We conclude that in patients with low-level HLA-DSA, ATG/IvIg induction significantly reduces TCR and the severity of AMR, but the high rate of subclinical AMR suggests an insufficient control of the humoral immune response.

Individual effect of renin-angiotensin system inhibition and corticosteroid therapy in IgA glomerulonephritis: results of a pilot study.

BACKGROUND: The impact of different therapy modalities on the outcome of Immunoglobulin A glomerulonephritis (IgAGN) in individual patient is not clear. We present preliminary results from the sequential application of renin-angiotensin system (RAS) inhibition and corticosteroids to discriminate the individual effect of both therapies. METHODS: Regardless of the degree of proteinuria, renal function and histological grading, patients with biopsy-proven IgAGN were treated with a standardized protocol. RAS inhibition was performed for 3 months. Thereafter, immunosuppressive therapy with prednisone (0.5 mg/kg body weight) on alternate days for 6 months was started. The primary endpoint was a maximal reduction of proteinuria (spot urine protein/ creatinine ratio (uPCR)), by RAS inhibition and by the combination of RAS inhibition and steroids. RESULTS: 10 patients were treated according to the protocol. During a median follow-up of 18 months, uPCR decreased from initial 230 mg/mmol (2 g/g) (median, interquartile range (IQR) 146 - 396) to 154 mg/mmol (1.4 g/g) (IQR 88 - 190) at 3 months during the RAS inhibition period (33% reduction, p = 0.01) and further to 31 mg/mmol (0.3 g/g) (IQR 21 - 71) until end of the steroid period at 9 months (80% reduction compared to uPCR at 3 month, p < 0.001). At the last F/U, uPCR (median) remained stable at 41 mg/mmol (0.4 g/g). The estimated glomerular filtration rate was stable during the whole observation period. CONCLUSIONS: Sequential RAS inhibition and steroid treatment leads to a continuous decrease in proteinuria, beyond the decrease produced by isolated RAS inhibition. Our data suggest independent effects of both, RAS inhibition and steroids, on the reduction of proteinuria in a small, non selected group of patients with IgAGN. The hypothesis that patients with IgAGN, regardless of the degree of proteinuria, renal function and histological grading, may benefit from combination therapy with maximal RAS inhibition and low dose corticosteroids now has to be confirmed in a randomized study.

Expression of carcinoembryonic antigen-related cell adhesion molecule 1 in acute rejection of human renal allografts.

BACKGROUND: Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is expressed on various cell types and mediates homophilic cell adhesion. CEACAM1 plays an important role in cell morphogenesis and angiogenesis. Furthermore, CEACAM1 regulates adhesive activity of immune-competent cells, suggesting an additional role in inflammatory processes. METHODS: Therefore, in the present study the expression of CEACAM1 was analysed retrospectively in renal biopsies from kidney transplant recipients (stable graft [Ctr; n = 18], acute vascular rejection [AVR; n = 14], acute tubulointerstitial rejection [AIR; n = 9], and combined vascular and interstitial rejection [AVIR; n = 7]). Expression patterns of CEACAM1 were determined using immunohistochemistry and quantitative morphometry. RESULTS: All biopsy specimens from patients with stable grafts showed low CEACAM1 levels, suggesting a constitutive expression in renal transplants. In patients with acute rejection, CEACAM1 was markedly up-regulated. AVR revealed the highest tubular CEACAM1 levels (4.9 +/- 0.5% [AVR] vs 2.2 +/- 0.3% [Ctr] of tubular area; P < .05), whereas interstitial rejections showed the highest glomerular expressions (4.5 +/- 0.5% [AIR] vs 0.9 +/- 0.1% [Ctr] of glomerular area; P < .05). CONCLUSIONS: An up-regulated expression of CEACAM1 in tubular and/or glomerular cells is an indicator of acute inflammatory processes in biopsy specimens from patients with acute renal allograft rejections and, therefore, might be used as a new clinical marker.

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.

Hereditary systemic angiopathy (HSA) with cerebral calcifications, retinopathy, progressive nephropathy, and hepatopathy.

Several hereditary conditions affecting cerebral, retinal and systemic microvessels have recently been described. They include CADASIL, CRV, and HERNS. We here report on a variant form of a hereditary systemic angiopathy (HSA) affecting two generations of a Caucasian family. Clinical symptoms of HSA appear in the mid-forties and are characterized by visual impairment, migraine-like headache, skin rash, epileptic seizures, progressive motor paresis and cognitive decline. Late symptoms include hepatic and renal failure. Retinal capillary microaneurysms and arteriolar tortuosity are associated with marked optic disc atrophy. Radiological hallmarks consist of multiple cerebral calcifications and tumor-like subcortical white matter lesions. Brain, peripheral nerve, muscle, kidney and colon biopsies have revealed a multi organ small vessel involvement with partly altered endothelium, perivascular inflammation and thrombotic microangiopathy. No curative therapeutic options are known for hereditary cerebral vasculopathies. The use of cyclophosphamide, azathioprine and methotrexate was of no benefit in our cases of HSA. Early diagnosis of hereditary systemic angiopathies is important in order to prevent patients from repetitive invasive diagnostic measures and to avoid the use of inappropriate and potentially harmful drugs.

Banff '05 Meeting Report: differential diagnosis of chronic allograft injury and elimination of chronic allograft nephropathy ('CAN').

The 8th Banff Conference on Allograft Pathology was held in Edmonton, Canada, 15-21 July 2005. Major outcomes included the elimination of the non-specific term "chronic allograft nephropathy" (CAN) from the Banff classification for kidney allograft pathology, and the recognition of the entity of chronic antibody-mediated rejection. Participation of B cells in allograft rejection and genomics markers of rejection were also major subjects addressed by the conference.

Pretransplant risk assessment in renal allograft recipients using virtual crossmatching.

Preformed donor-specific HLA-antibodies antibodies (DSA) are a major risk for early antibody-mediated rejection (AMR). This prospective study evaluated the accuracy of pretransplant risk assessment using virtual crossmatching (virtualXM) (i.e. comparing HLA-typing of the donor with the recipient's HLA-antibody specificities determined by flow-beads). Sixty-five consecutive patients were stratified according to virtualXM results: patients without DSA (n= 56) were considered low risk and received standard immunosuppression; patients with DSA (n= 9) were considered high risk and received additional induction with anti-T-lymphocyte-globulin (ATG) and intravenous immunoglobulins. Despite induction therapy 4 of 9 patients with DSA (44%) had clinical/subclinical AMR, whereas only 2 of 56 patients without DSA (4%) (p = 0.002). Notably, one of these two patients had early AMR likely induced by non-HLA-antibodies; the other had subclinical AMR at month 6 consistent with de novo DSA. The results of virtualXM and retrospectively obtained flow-cytometric crossmatches (FCXM) (n= 59) were concordant in 51 patients (86%), four patients (7%) were virtualXM-/FCXM+ and none had AMR, four patients (7%) were virtualXM+/FCXM- and one had AMR. VirtualXM can accurately define absence or presence of DSA and may become an invaluable tool for organ allocation and pretransplant risk assessment. However, further studies need to address whether all HLA-antibodies detected by flow-beads are clinically relevant.

Protocol biopsies in renal transplantation: insights into patient management and pathogenesis.

A 1-day symposium on the application of protocol biopsies in renal transplantation was held in Boston, 21 July 2006. Representatives from centers with extensive experience in the use of protocol biopsies for routine patient care and research reported results on the pathological findings and their value in patient management. The consensus was that protocol biopsies, in experienced hands, are a safe and valuable means of detecting subclinical disease that can benefit from modification of therapy. Furthermore, molecular studies reveal evidence of activity or progression not readily appreciated by histological techniques. Wider application is expected in multicenter clinical trials to predict and validate outcomes. The principal barrier to wider use of protocol biopsies is knowledge of the benefits of intervention.

KCNMA1 gene amplification promotes tumor cell proliferation in human prostate cancer.

Molecular mechanisms of prostate cancer progression are poorly understood. Here, we studied gene amplification of the large conductance calcium-activated potassium channel alpha subunit (KCNMA1), which is located at the chromosomal region 10q22. Fluorescence in situ hybridization (FISH) revealed KCNMA1 amplification in 16% of 119 late-stage human prostate cancers and in the hormone-insensitive prostate cancer cell line PC-3. In contrast, KCNMA1 amplification was absent in 33 benign controls, 32 precursor lesions and in 105 clinically organ-confined prostate cancers. Amplification was associated with mRNA and protein overexpression as well as increased density of BK channel protein and beta-estradiol-insensitive BK currents in PC-3 cells as compared to non-amplified control cell lines. Specific blockade of BK channels by iberiotoxin or RNA(i) significantly inhibited K(+) currents and growth of PC-3 cells. The data demonstrate that 10q22 amplification drives KCNMA1 expression and cell proliferation. Thus, KCNMA1 qualifies as a promising diagnostic and therapeutic target in patients with prostate cancer.

[Urinary cytology after renal transplantation]. / Urindiagnostik bei nierentransplantierten Patienten.

BK polyomavirusnephropathy (BKN) is a serious complication of the potent immunosuppressive therapies in renal allograft recipients. BKN occurs in 5% of the patients and leads to allograft loss in approximately 50% of those affected. Early detection of BKN improves the chances to retain the allograft by allowing immediate intervention. Urine cytology is a simple, cheap and reliable method for early identification of patients at risk for BKN, together with BK DNA analysis in the serum. BK virus causes a spectrum of morphologic changes of tubular and urothelial cells that can be viewed online at http://vmic.unibas.ch/patho/seminar/index.html. These BK infected cells are called "decoy cells" as they can mimic the features of bladder carcinoma cells. In addition, certain morphologic features of the urinary sediment including tubular cells and lymphocytes can anticipate renal rejection. Historically, urinary cytology was important to screen patients with allograft for analgetic nephropathy for malignant cells, since they were at greatly increased risk of urothelial carcinoma.

Reverse diastolic intrarenal flow due to calcineurin inhibitor (CNI) toxicity.

Renal calcineurin inhibitor (CNI) toxicity is a frequent side effect of immunosuppression with CNIs in solid organ transplantation, leading to acute and chronic renal failure. Acute CNI toxicity is due to vasoconstriction of the vasa afferens and efferens and vacuolization of smooth muscle cells with medial hyalinosis, leading to vessel lumen narrowing. Our case had an acute renal failure 8 months after deceased donor kidney transplantation under treatment with tacrolimus, sirolimus and prednisolone. In Doppler sonography, we observed reverse diastolic intrarenal blood flow, reflecting intense vessel narrowing. There were histological signs of acute CNI toxicity. Within days of reducing the tacrolimus trough level, renal function improved markedly and Doppler sonography showed orthograde intrarenal blood flow. This is the first case of functional, Doppler sonographic evidence for CNI-induced, rapidly reversible narrowing of intrarenal vessels. This case illustrates the potential role of tacrolimus and sirolimus dosing in combination therapy to produce severe intrarenal vasoconstriction.

Bilateral renal vein thrombosis in a twin newborn without known risk factors.

Neonatal renal vein thrombosis (RVT) is associated with neonatal stress, catheters and genetic prothrombotic risk factors. In an unusual case of bilateral RVT a twin newborn showed initial good adaptation at birth (weight 2,720 g). The placenta was monochorionic, diamnionic. The infant (gestational week 37) exhibited a severe macrohematuria within 24 hours after birth. Sonography of the kidneys showed a dense cortical parenchyma, loss of cortico-medullary differentiation and negative diastolic flow in both renal arteries and veins, while no thrombus in the main renal veins could be detected. No prothrombotic blood parameters and positive infection serology were detected. Because of acute renal failure peritoneal dialysis was necessary for 6 weeks. The patient was treated by heparinization for 5 days. Interestingly, it was kidney biopsy which confirmed the diagnosis of RVT in addition to the clinical presentation, whereas sonography was unspecific. Histology exhibited the picture of an ischemic contracted kidney with numerous siderophages. At present (age 19 months), the patient suffers from chronic renal failure (calculated glomerular filtration rate according to Schwartz 12 ml/min/1.73 m2). In conclusion, our case teaches that, despite the lack of a clinically obvious shock event, absence of known risk factors and indirect ultrasound findings, renal vein thrombosis should be considered in a macrohematuric newborn with renal failure. For clinical suspicion of RVT correct therapy was initiated, however, the diagnosis remained unclear until a renal biopsy was performed.

Acute and chronic vascular rejection in nonhuman primate kidney transplantation.

A nonhuman primate (NHP) study was designed to evaluate in nonlife-supporting kidney allografts the progression from acute rejection with transplant endarteritis (TXA) to chronic rejection (CR) with sclerosing vasculopathy. Group G1 (n = 6) received high cyclosporine A (CsA) immunosuppression and showed neither TXA nor CR during 90 days post-transplantation. Group G2 (n = 6) received suboptimal CsA immunosuppression and showed severe TXA with graft loss within 46 days (median). Arterial intimal changes included infiltration of macrophages and T lymphocytes (CD3, CD4, CD8) with few myofibroblasts, abundant fibronectin/collagen IV, scant collagens I/III, high rate of cellular proliferation and no C4d accumulation along peritubular capillaries. Group G3 (n = 12) received suboptimal CsA and anti-rejection therapy (rabbit ATG + methylprednisolone + CsA) of TXA. Animals developed CR and lost grafts within 65 days (median). As compared to G2, the arterial intimal changes showed less macrophages and T lymphocytes, an increased number of myofibroblasts, abundant fibronectin/collagen IV and scar collagens I/III, C4d deposition along capillaries in 60% of animals and transplant glomerulopathy in 80% of animals. In conclusion, CR is an immune stimulated process initiated during TXA with the accumulation and proliferation of myofibroblasts, and progressive deposition of collagens in the intima. Our experimental design appears well suited to study events leading to CR.