Differential effects of donor-specific HLA antibodies in living versus deceased donor transplant.

The presence of donor-specific anti-HLA antibodies (DSAs) is associated with increased risk of graft failure after kidney transplant. We hypothesized that DSAs against HLA class I, class II, or both classes indicate a different risk for graft loss between deceased and living donor transplant. In this study, we investigated the impact of pretransplant DSAs, by using single antigen bead assays, on long-term graft survival in 3237 deceased and 1487 living donor kidney transplants with a negative complement-dependent crossmatch. In living donor transplants, we found a limited effect on graft survival of DSAs against class I or II antigens after transplant. Class I and II DSAs combined resulted in decreased 10-year graft survival (84% to 75%). In contrast, after deceased donor transplant, patients with class I or class II DSAs had a 10-year graft survival of 59% and 60%, respectively, both significantly lower than the survival for patients without DSAs (76%). The combination of class I and II DSAs resulted in a 10-year survival of 54% in deceased donor transplants. In conclusion, class I and II DSAs are a clear risk factor for graft loss in deceased donor transplants, while in living donor transplants, class I and II DSAs seem to be associated with an increased risk for graft failure, but this could not be assessed due to their low prevalence.

Urinary MicroRNA as Biomarker in Renal Transplantation.

Urine represents a noninvasive source in which proteins and nucleic acids can be assessed. Such analytes may function as biomarkers to monitor kidney graft pathology at every desired frequency, thereby providing a time window to prevent graft damage by therapeutic intervention. Recently, several proteins have been measured in urine as markers of graft injury. However, the specificity is limited, and measuring urinary proteins generally lacks the potential to predict early kidney graft damage. Currently, urinary mRNA and microRNA are being investigated to evaluate the prognostic value of changes in gene expression during the initial stages of graft damage. At such time point, a change in treatment regimen and dosage is expected to have maximum potency to minimize future decline in graft function. Both mRNA and microRNAs have shown promising results in both detection and prediction of graft injury. An advantage of microRNAs compared to mRNA molecules is their stability, a characteristic that is beneficial when working with urine samples. In this review, we provide the current state of urinary biomarkers in renal transplantation, with a focus on urinary microRNA. In addition, we discuss the methods used to study urinary microRNA expression.

Reactivation of Latent HPV Infections After Renal Transplantation.

Female renal transplant recipients (RTRs) have an increased risk for developing human papillomavirus (HPV)-related (pre)malignant lesions of the genital tract. This study aims to assess the genital prevalence of HPV before and after renal transplantation (RT). In female patients who were counseled for RT at the Radboud University Medical Center Nijmegen, the Netherlands, gynecological examination was performed at first visit, and 1 and 2 years later. HPV self-sampling and questionnaires on sexual behavior were performed every 3 months. In 65 patients who underwent RT, the high-risk human papillomavirus (hrHPV) prevalence as assessed with the highly sensitive SPF10 -LiPA25 test increased significantly from 19% before to 31% after RT (p = 0.045). Based upon the clinically validated Cobas 4800 HPV test, the hrHPV prevalence increased from 10% before to 14% after RT (p = 0.31). During follow-up, no changes in sexual behavior were reported. Thirty-three patients who did not undergo RT showed a hrHPV prevalence of 21% at study entry and of 27% after 12 months with the sensitive test, and a stable prevalence of 16% with the clinically validated test. The results of this study indicate that activation of latent HPV infections may contribute to the increased risk of HPV-related (pre)malignant lesions in female RTRs.

Pretransplant Numbers of CD16+ Monocytes as a Novel Biomarker to Predict Acute Rejection After Kidney Transplantation: A Pilot Study.

Acute rejection is one of the major immunological determinants of kidney graft function and survival. Early biomarkers to predict rejection are lacking. Emerging evidence reveals a crucial role for the monocyte/macrophage lineage cells in the pathogenesis of rejection. We hypothesized that higher pretransplant numbers of proinflammatory CD16+ monocytes can predict rejection. The study cohort consisted of 104 kidney transplant recipients (58 with no rejection and 46 with biopsy-proven rejection) and 33 healthy persons. Posttransplant median follow-up time was 14.7 mo (interquartile range 0.3-34 mo). Pretransplantation blood samples were analyzed by flow cytometry for monocyte immunophenotypes. Groups were compared by Cox regression models for the occurrence of acute rejection. We documented a significantly increased absolute number of pretransplant CD16+ monocytes in patients who developed biopsy-proven rejection after transplantation compared with those with no rejection (hazard ratio [HR] 1.60, 95% CI 1.28-2.00, p < 0.001) and healthy persons (HR 1.47, 95% CI 1.18-1.82, p < 0.001). In parallel, significantly fewer absolute numbers of CD16- monocytes were observed at pretransplant time points in rejectors versus nonrejectors (HR 0.74, 95% CI 0.58-0.94, p < 0,014). A higher pretransplant number of CD16+ monocytes is significantly associated with a higher risk of acute rejection after kidney transplantation.

The Course and Predictors of Health-Related Quality of Life in Living Kidney Donors: A Systematic Review and Meta-Analysis.

A better understanding of the course and risk factors for impaired long-term health-related quality of life (HRQoL; ie, physical, psychological, and social-relational functioning) after kidney donation might help clinicians improve the care of live kidney donors. This systematic review and meta-analysis summarizes prospective studies about the course and predictors of HRQoL in living kidney donors. Studies indicate that shortly after donation, donors have lower HRQoL, with minor to moderate changes in psychological and social-relational functioning and major changes in physical functioning. At 3-12 months after donation, HRQoL returned to baseline or was slightly reduced, particularly for fatigue, but scores were still comparable to general population norms. Results were mainly robust across surgery techniques. A limited number of studies examined risk factors for impaired HRQoL, with low psychological functioning before donation as the most consistent predictor. Based on these results, clinicians can inform potential donors that, on average, kidney donors have high long-term HRQoL; however, donors with low psychological functioning at baseline are those most at risk of impaired long-term HRQoL. Future studies should focus on other potentially relevant predictors of postdonation HRQoL, including donor eligibility criteria and donor-recipient relationships, to optimize screening and interventions for donors at risk.

Rituximab as induction therapy after renal transplantation: a randomized, double-blind, placebo-controlled study of efficacy and safety.

We evaluated the efficacy and safety of rituximab as induction therapy in renal transplant patients. In a double-blind, placebo-controlled study, 280 adult renal transplant patients were randomized between a single dose of rituximab (375 mg/m(2)) or placebo during transplant surgery. Patients were stratified according to panel-reactive antibody (PRA) value and rank number of transplantation. Maintenance immunosuppression consisted of tacrolimus, mycophenolate mofetil and steroids. The primary endpoint was the incidence of biopsy proven acute rejection (BPAR) within 6 months after transplantation. The incidence of BPAR was comparable between rituximab-treated (23/138, 16.7%) and placebo-treated patients (30/142, 21.2%, p = 0.25). Immunologically high-risk patients (PRA >6% or re-transplant) not receiving rituximab had a significantly higher incidence of rejection (13/34, 38.2%) compared to other treatment groups (rituximab-treated immunologically high-risk patients, and rituximab- or placebo-treated immunologically low-risk (PRA ≤ 6% or first transplant) patients (17.9%, 16.4% and 15.7%, p = 0.004). Neutropenia (<1.5 × 10(9) /L) occurred more frequently in rituximab-treated patients (24.3% vs. 2.2%, p < 0.001). After 24 months, the cumulative incidence of infections and malignancies was comparable. A single dose of rituximab as induction therapy did not reduce the overall incidence of BPAR, but might be beneficial in immunologically high-risk patients. Treatment with rituximab was safe.

Cervicovaginal HPV infection in female renal transplant recipients: an observational, self-sampling based, cohort study.

Immunosuppressive treatment of organ transplant recipients is associated with an increase in the occurrence of human papillomavirus (HPV) related anogenital (pre)malignancies. This cohort study investigated the genotype-specific prevalence of HPV infections in a large cohort of female renal transplant recipients (RTRs). Participants self-collected a cervicovaginal sample for detection and genotyping of HPV. Besides, they completed a questionnaire regarding sociodemographic variables, medical data and sexual behavior. Anogenital screening was offered to all HPV-positive participants. A total number of 218 female RTRs was included. The prevalence of mucosal HPV infections was 27.1% and 17.4% for high risk HPV in particular. The studied cohort showed a broad range of HPV genotypes and multiple HPV genotypes were found in 27.1% of HPV-positive patients. Seven participants were identified with occult premalignant anogenital lesions. In conclusion, this study shows a high point-prevalence of HPV in female RTRs (age-matched West-European general population: 9-10%) with a shift in the distribution of genotypes as compared with the general population. Moreover, a substantial number of patients with occult premalignancies was identified. The introduction of self-sampling for HPV positivity can help in early detection of (pre)malignant anogenital lesions in this vulnerable population.

Limited sampling strategy for prolonged-release tacrolimus in renal transplant patients by use of the dried blood spot technique.

PURPOSE: The aim of this study was to develop a clinically applicable limited sampling strategy for ambulatory Caucasian kidney transplant patients to estimate area under the curve in a 24-h period (AUC0-24) of prolonged-release tacrolimus. METHODS: Twenty six kidney recipients, at least 6 months after transplantation, receiving prolonged-release tacrolimus, were enrolled. In each patient, seven blood samples were collected during a period of 24 h by use of the validated dried blood spot method. Best subset selection multiple linear regression was performed to derive limited sampling strategy (LSS). The equations were constrained to include a maximum of three samples collected within 4 h after the intake to maintain clinical applicability. To assess the predictive performance of LSS, residuals for each patient were calculated based on models fitted to a dataset where that patient was omitted. RESULTS: The prediction formula for the AUC(0-24) using the time points 0, 2, and 4 h after ingestion (C(0h)-C(2h)-C(4h)) provided the highest correlation with the AUC(0-24) (r(2) = 0.95): AUC0-24 = 44.9 + 8.9 × C(0h) + 2.1 × C(2h) + 7.6 × C(4h). Measures for bias and precision, i.e., median percentage prediction error (MPPE) and median absolute prediction error (MAPE), were 0.4 and 4.8%, respectively. For the same patients, the correlation between C(24h) and AUC0-24 was worse (r(2) = 0.77) while MPPE and MAPE were 6.2 and 7.2%, respectively. CONCLUSION: In the outpatient department, a LSS using C(0h)-C(2h)-C(4h) can be used for reliable estimation of the AUC(0-24) of prolonged-release tacrolimus.

Treatment of steroid-resistant acute renal allograft rejection with alemtuzumab.

Steroid-resistant renal allograft rejections are commonly treated with rabbit antithymocyte globulin (RATG), but alemtuzumab could be an effective, safe and more convenient alternative. Adult patients with steroid-resistant renal allograft rejection treated with alemtuzumab (15-30 mg s.c. on 2 subsequent days) from 2008 to 2012 (n = 11) were compared to patients treated with RATG (2.5-4.0 mg/kg bodyweight i.v. for 10-14 days; n = 20). We assessed treatment-failure (graft loss, lack of improvement of graft function or need for additional anti-rejection treatment), infections during the first 3 months after treatment and infusion-related side effects. In both groups, the median time-interval between rejection and transplantation was 2 weeks, and approximately 75% of rejections were classified as Banff-IIA or higher. Three alemtuzumab-treated patients (27%) experienced treatment failure, compared to eight RATG treated patients (40%, p = 0.70). There was no difference in the incidence of infections. There were mild infusion-related side-effects in three alemtuzumab-treated patients (27%), and more severe infusion-related side effects in 17 RATG-treated patients (85%, p = 0.013). Drug related costs of alemtuzumab-treatment were lower than of RATG-treatment (€1050 vs. €2024; p < 0.01). Alemtuzumab might be an effective therapy for steroid-resistant renal allograft rejections. In contrast to RATG, alemtuzumab is nearly devoid of infusion-related side-effects. These data warrant a prospective trial.

A single dose of rituximab does not deplete B cells in secondary lymphoid organs but alters phenotype and function.

A single dose of the anti-CD20 monoclonal antibody rituximab induces a nearly complete B cell depletion in peripheral blood, but not in secondary lymphoid organs. Modulation of this remaining B cell population due to rituximab treatment may contribute to the therapeutic effects of rituximab. To assess the in vivo effects of rituximab we used lymph nodes (LNs) collected during renal transplant surgery in patients who had received rituximab 4 weeks earlier in preparation for an ABO-incompatible transplantation. Rituximab treatment resulted in a lower percentage of naïve (IgD(+)CD27(-)) and a higher percentage of switched memory (IgD(-)CD27(+)) B cells. Remarkably, transitional (CD24(++)CD38(++)) B cells were virtually lacking in the LNs of rituximab-treated patients. Moreover, LN-derived B cells from rituximab-treated patients produced different amounts of various Ig-subclasses after anti-CD40/IL-21 stimulation ex vivo. Finally, after stimulation of allogeneic T cells with LN-derived B cells from rituximab-treated patients, the proliferated T cells showed a decreased production of IL-17. In conclusion, after treatment with rituximab there remains a B cell population with different functional capacities. Consequently, the effect of rituximab on the immune response will not only be determined by the extent of B cell depletion, but also by the functional properties of the remaining B cells.

In vitro effects of rituximab on the proliferation, activation and differentiation of human B cells.

Rituximab is a chimeric anti-CD20 monoclonal antibody (mAb) used in B-cell malignancies, various autoimmune disorders and organ transplantation. Although administration of a single dose of rituximab results in full B-cell depletion in peripheral blood, there remains a residual B-cell population in secondary lymphoid organs. These nondepleted B cells might be altered by exposure to rituximab with subsequent immunomodulatory effects. Therefore, we analyzed in vitro the effects of rituximab on proliferation, activation and differentiation of CD19(+) B cells by means of carboxyfluorescein succinimidyl ester (CFSE)-based multiparameter flow cytometry. Rituximab inhibited the proliferation of CD27(-) naïve, but not of CD27(+) memory B cells. Interestingly, upon stimulation with anti-CD40 mAb and interleukin-21 in the presence of rituximab there was an enrichment of B cells that underwent only one or two cell divisions and displayed an activated naïve phenotype (CD27(-)IgD(+)CD38(-/+)). The potency of prestimulated B cells to induce T-cell proliferation was increased by exposure of the B cells to rituximab. Of note, after stimulation with rituximab-treated B cells, proliferated T cells displayed a more Th2-like phenotype. Overall, these results demonstrate that rituximab can affect human B-cell phenotype and function, resulting in an altered outcome of B-T cell interaction.

Synchronized turbo apoptosis induced by cold-shock.

In our research on the role of apoptosis in the pathogenesis of the autoimmune disease systemic lupus erythematosus (SLE), we aim to evaluate the effects of early and late apoptotic cells and blebs on antigen presenting cells. This requires the in vitro generation of sufficiently large and homogeneous populations of early and late apoptotic cells. Here, we present a quick method encountered by serendipity that results in highly reproducible synchronized homogeneous apoptotic cell populations. In brief, granulocytic 32Dcl3 cells are incubated on ice for 2 h and subsequently rewarmed at 37°C. After 30-90 min at 37°C more than 80-90% of the cells become early apoptotic (Annexin V positive/propidium iodide negative). After 24 h of rewarming at 37°C 98% of the cells were late apoptotic (secondary necrotic; Annexin V positive/propidium iodide positive). Cells already formed apoptotic blebs at their cell surface after approximately 20 min at 37°C. Inter-nucleosomal chromatin cleavage and caspase activation were other characteristics of this cold-shock-induced process of apoptosis. Consequently, apoptosis could be inhibited by a caspase inhibitor. Finally, SLE-derived anti-chromatin autoantibodies showed a high affinity for apoptotic blebs generated by cold-shock. Overall, cold-shock induced apoptosis is achieved without the addition of toxic compounds or antibodies, and quickly leads to synchronized homogeneous apoptotic cell populations, which can be applied for various research questions addressing apoptosis.

Reduced CXCL1 production by endogenous IL-37 expressing dendritic cells does not affect T cell activation.

The dendritic cell (DC)-derived cytokine profile contributes to naive T cell differentiation, thereby directing the immune response. IL-37 is a cytokine with anti-inflammatory characteristics that has been demonstrated to induce tolerogenic properties in DC. In this study we aimed to evaluate the influence of IL-37 on DC-T cell interaction, with a special focus on the role of the chemokine CXCL1. DC were cultured from bone marrow of human IL-37 transgenic (hIL-37Tg) or WT mice. The phenotype of unstimulated and LPS-stimulated DC was analyzed (co-stimulatory molecules and MHCII by flow cytometry, cytokine profile by RT-PCR and ELISA), and T cell stimulatory capacity was assessed in mixed lymphocyte reaction. The role of CXCL1 in T cell activation was analyzed in T cell stimulation assays with anti-CD3 or allogeneic DC. The expression of the co-stimulatory molecules CD40, CD80 and CD86, and of MHCII in LPS-stimulated DC was not affected by endogenous expression of IL-37, whereas LPS-stimulated hIL-37Tg DC produced less CXCL1 compared to LPS-stimulated WT DC. T cell stimulatory capacity of LPS-matured hIL-37Tg DC was comparable to that of WT DC. Recombinant mouse CXCL1 did not increase T cell proliferation either alone or in combination with anti-CD3 or allogeneic DC, nor did CXCL1 affect the T cell production of interferon-γ and IL-17. Endogenous IL-37 expression does not affect mouse DC phenotype or subsequent T cell stimulatory capacity, despite a reduced CXCL1 production. In addition, we did not observe an effect of CXCL1 in T cell proliferation or differentiation.

Cumulative pemetrexed dose increases the risk of nephrotoxicity.

INTRODUCTION: Pemetrexed is a pharmacotherapeutic cornerstone in the treatment of non-small cell lung cancer. As it is primarily eliminated by renal excretion, adequate renal function is essential to prevent toxic exposure. There is growing evidence for the nephrotoxic potential of pemetrexed, which even becomes a greater issue now combined immuno-chemotherapy prolongs survival. Therefore, the aim of this study was to describe the incidence of nephrotoxicity and related treatment consequences during pemetrexed-based treatment. METHODS: A retrospective cohort study was conducted in the Jeroen Bosch Hospital, Den Bosch, the Netherlands. All patients that received at least 1 cycle of pemetrexed based therapy were included in the dataset. The primary outcome was defined as a ≥25 % reduction in eGFR. Additionally, the treatment consequences of decreased renal function were assessed. Logistic regression was used to identify risk factors for nephrotoxicity during treatment with pemetrexed. RESULTS: Of the 359 patients included in this analysis, 21 % patients had a clinically relevant decline in renal function after treatment and 8.1 % of patients discontinued treatment due to nephrotoxicity. Cumulative dose (≥10 cycles of pemetrexed based therapy) was identified as a risk factor for the primary outcome measure (adjusted OR 5.66 (CI 1.73-18.54)). CONCLUSION: This study shows that patients on pemetrexed-based treatment are at risk of developing renal impairment. Risk significantly increases with prolonged treatment. Renal impairment is expected to become an even greater issue now that pemetrexed-based immuno-chemotherapy results in longer survival and thus longer treatment duration.

Encapsulating peritoneal sclerosis in patients on peritoneal dialysis.

Encapsulating peritoneal sclerosis (EPS) is an uncommon but one of the most serious complications in patients on long-term peritoneal dialysis. EPS is characterised by a diffuse thickening and/or sclerosis of the peritoneal membrane which leads to a decreased ultrafiltration and ultimately to bowel obstruction. We present four cases of EPS and discuss the clinical manifestations, multifactorial aetiology, diagnosis, treatment, prognosis, and prevention. We end with a proposal for the development of an EPS prevention guideline.

Rapamycin, not cyclosporine, permits thymic generation and peripheral preservation of CD4+ CD25+ FoxP3+ T cells.

Graft-versus-host-disease (GVHD) is the most common cause of poor outcome after allogeneic stem cell transplantation (SCT). Of late, exploitation of FOXP3(+) regulatory T-cell (T(REG)) function is emerging as a promising strategy in suppression of GVHD, while preserving graft-versus-leukemia (GVL). Cyclosporine and rapamycin reduce the expansion of effector T cells by blocking interleukin (IL)-2, but signaling by IL-2 is pivotal for T(REG) homeostasis. The resolution of GVHD is critically dependent on thymus-dependent reconstitution of the immunoregulatory system. Thus, there has been concern about the impact of blocking IL-2 signaling by immunosuppressive agents on T(REG) homeostasis. Here we demonstrate in a mouse model that in contrast to rapamycin, cyclosporine compromises not only the thymic generation of CD4(+)CD25(+)FoxP3(+) T cells but also their homeostatic behavior in peripheral immune compartments. Treatment with cyclosporine resulted in a sharp reduction of peripheral CD25(+)FoxP3(+) T cells in all immune compartments studied. Prolonged rapamycin treatment allowed for thymic generation of CD4(+)FoxP3(+) T cells, whereas treatment with cyclosporine led to a reduced generation of these cells. In conclusion, cyclosporine and rapamycin differentially affect homeostasis of CD4(+)FoxP3(+) T(REG) in vivo. As peripheral tolerance induction is a prerequisite for successful treatment outcome after allogeneic SCT, these findings are of potential clinical relevance.

Allostimulatory Effects of Dendritic Cells with Characteristic Features of a Regulatory Phenotype.

INTRODUCTION: Tolerogenic dendritic cells (DCs) have the potential to prolong graft survival after transplantation. Tolerogenic DCs are in general characterized by a low expression of co-stimulatory molecule and a high IL-10:IL-12 production ratio. Based on promising results with earlier used alternatively activated DCs, we aimed to generate in culture potentially tolerogenic DC by simultaneously blocking GSK3 by lithium chloride (LiCl) and stimulating TLR2 by PAM3CysSerLys4. MATERIALS AND METHODS: Bone marrow-derived LiClPAM3 DCs were generated by the addition of LiCl 24 hours before harvesting, and one hour later PAM3CysSerLys4. The phenotype of the DCs was assessed by determining the expression of co-stimulatory molecules in flow cytometry and cytokine production in ELISA, whereas their functional properties were tested in a mixed lymphocyte reaction. A fully MHC mismatched heterotopic heart transplant preceded by infusion of donor-derived LiClPAM3 DC was performed to assess the tolerogenic potential of LiClPAM3 DCs in vivo. RESULTS: LiClPAM3 DCs displayed a tolerogenic phenotype accompanied with a low expression of co-stimulatory molecules and a high IL-10:IL-12 production ratio. However, in mixed lymphocyte reaction, LiClPAM3 DCs appeared superior in T cell stimulation, and induced Th1 and Th17 differentiation. Moreover, mice pretreated with LiClPAM3 DC displayed a reduced graft survival. Analysis of LiClPAM3 DC culture supernatant revealed high levels of CXCL-1, which was also found in supernatants of co-cultures of LiClPAM3 DC and T cells. Nevertheless, we could not show a role for CXCL-1 in T cell proliferation or activation in vitro. DISCUSSION: LiClPAM3 DCs display in vitro a tolerogenic phenotype with a high IL-10:IL-12 ratio, but appeared to be highly immunogenic, since allograft rejection was accelerated. As yet unidentified LiClPAM3 DC-derived factors, may explain the immunogenic character of LiClPAM3 DCs in vivo.

How can we reduce costs of solid-phase multiplex-bead assays used to determine anti-HLA antibodies?

Solid-phase multiplex-bead assays are widely used in transplantation to detect anti-human leukocyte antigen (HLA) antibodies. These assays enable high resolution detection of low levels of HLA antibodies. However, multiplex-bead assays are costly and yield variable measurements that limit the comparison of results between laboratories. In the context of a Dutch national Consortium study we aimed to determine the inter-assay and inter-machine variability of multiplex-bead assays, and we assessed how to reduce the assay reagents costs. Fifteen sera containing a variety of HLA antibodies were used yielding in total 7092 median fluorescence intensities (MFI) values. The inter-assay and inter-machine mean absolute relative differences (MARD) of the screening assay were 12% and 13%, respectively. The single antigen bead (SAB) inter-assay MARD was comparable, but showed a higher lot-to-lot variability. Reduction of screening assay reagents to 50% or 40% of manufacturers' recommendations resulted in MFI values comparable to 100% of the reagents, with an MARD of 12% or 14%, respectively. The MARD of the 50% and 40% SAB assay reagent reductions were 11% and 22%, respectively. From this study, we conclude that the reagents can be reliably reduced at least to 50% of manufacturers' recommendations with virtually no differences in HLA antibody assignments.

Idiopathic focal segmental glomerulosclerosis: a favourable prognosis in untreated patients?

BACKGROUND: Patients with focal segmental glomerulosclerosis (FSGS) are considered to have a poor prognosis and spontaneous remissions are seldom reported. However, FSGS is not a single disease entity. Our aim was to describe the clinical course in initially untreated patients with recently diagnosed idiopathic FSGS. METHODS: This was a retrospective study of patients with a diagnosis of FSGS by histology, who fulfilled the following criteria: proteinuria >3.5 g/day, normal renal function, duration of proteinuria or hypertension of less than one year, normal-sized kidneys, no underlying renal disease, and a negative family history. Renal biopsies were reviewed without knowledge of the clinical course. RESULTS: Twenty patients (13 male, 7 female) fulfilled the study criteria. Median age was 49.3 (range 21.8 to 73.0) years, serum creatinine 90 +/- 20 micromol/l, proteinuria 10.0 +/- 5.5 g/day and serum albumin 24 +/- 6 g/l. After a median follow-up of 9.4 (2.1-18.6) years, 13 patients (65%) were in remission of proteinuria. Renal function deterioration occurred in seven patients, and prompted treatment in four of them. The ten-year death-censored renal survival was 89%. Renal function deterioration and remission rate could be predicted by selectivity index, serum albumin at three months after renal biopsy and the percentage of glomeruli with segmental sclerosis. CONCLUSION: Focal glomerulosclerosis is not a single disease. Case definition using strict clinical criteria identifies a subgroup of patients with idiopathic FSGS who have a good prognosis. In the majority of these patients immunosuppressive therapy is not warranted.

Human proximal tubule epithelial cells cultured on hollow fibers: living membranes that actively transport organic cations.

The bioartificial kidney (BAK) aims at improving dialysis by developing 'living membranes' for cells-aided removal of uremic metabolites. Here, unique human conditionally immortalized proximal tubule epithelial cell (ciPTEC) monolayers were cultured on biofunctionalized MicroPES (polyethersulfone) hollow fiber membranes (HFM) and functionally tested using microfluidics. Tight monolayer formation was demonstrated by abundant zonula occludens-1 (ZO-1) protein expression along the tight junctions of matured ciPTEC on HFM. A clear barrier function of the monolayer was confirmed by limited diffusion of FITC-inulin. The activity of the organic cation transporter 2 (OCT2) in ciPTEC was evaluated in real-time using a perfusion system by confocal microscopy using 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide (ASP(+)) as a fluorescent substrate. Initial ASP(+) uptake was inhibited by a cationic uremic metabolites mixture and by the histamine H2-receptor antagonist, cimetidine. In conclusion, a 'living membrane' of renal epithelial cells on MicroPES HFM with demonstrated active organic cation transport was successfully established as a first step in BAK engineering.