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Hemolytic uremic syndromes (HUSs) are a heterogeneous group of conditions, only some of which are mediated by complement (complement-mediated HUS). We report the outcome of the 2023 International Society of Nephrology HUS International Forum where a global panel of experts considered the current state of the art, identified areas of uncertainty, and proposed optimal solutions. Areas of uncertainty and areas for future research included the nomenclature of HUS, novel complement testing strategies, identification of biomarkers, genetic predisposition to atypical HUS, optimal dosing and withdrawal strategies for C5 inhibitors, treatment of kidney transplant recipients, disparity of access to treatment, and the next generation of complement inhibitors in complement-mediated HUS. The current rationale for optimal patient management is described., (Copyright © 2024 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Our understanding of the various aspects of pregnancy in women with kidney diseases has significantly improved in the last decades. Nevertheless, little is known about specific kidney diseases. Glomerular diseases are not only a frequent cause of chronic kidney disease in young women, but combine many challenges in pregnancy: immunologic diseases, hypertension, proteinuria, and kidney tissue damage. An international working group undertook the review of available current literature and elicited expert opinions on glomerular diseases in pregnancy with the aim to provide pragmatic information for nephrologists according to the present state-of-the-art knowledge. This work also highlights areas of clinical uncertainty and emphasizes the need for further collaborative studies to improve maternal and fetal health., (Copyright © 2022 International Society of Nephrology. All rights reserved.)

A cyclical corticosteroid-cyclophosphamide regimen is recommended for patients with primary membranous nephropathy at high risk of progression. We hypothesized that sequential therapy with tacrolimus and rituximab is superior to cyclical alternating treatment with corticosteroids and cyclophosphamide in inducing persistent remission in these patients. This was tested in a randomized, open-label controlled trial of 86 patients with primary membranous nephropathy and persistent nephrotic syndrome after six-months observation and assigned 43 each to receive six-month cyclical treatment with corticosteroid and cyclophosphamide or sequential treatment with tacrolimus (full-dose for six months and tapering for another three months) and rituximab (one gram at month six). The primary outcome was complete or partial remission of nephrotic syndrome at 24 months. This composite outcome occurred in 36 patients (83.7%) in the corticosteroid-cyclophosphamide group and in 25 patients (58.1%) in the tacrolimus-rituximab group (relative risk 1.44; 95% confidence interval 1.08 to 1.92). Complete remission at 24 months occurred in 26 patients (60%) in the corticosteroid-cyclophosphamide group and in 11 patients (26%) in the tacrolimus-rituximab group (2.36; 1.34 to 4.16). Anti-PLA2R titers showed a significant decrease in both groups but the proportion of anti-PLA2R-positive patients who achieved immunological response (depletion of anti-PLA2R antibodies) was significantly higher at three and six months in the corticosteroid-cyclophosphamide group (77% and 92%, respectively), as compared to the tacrolimus-rituximab group (45% and 70%, respectively). Relapses occurred in one patient in the corticosteroid-cyclophosphamide group, and three patients in the tacrolimus-rituximab group. Serious adverse events were similar in both groups. Thus, treatment with corticosteroid-cyclophosphamide induced remission in a significantly greater number of patients with primary membranous nephropathy than tacrolimus-rituximab., (Copyright © 2020 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Recurrence of primary membranous nephropathy after transplantation occurs in up to 44% of patients and is driven by PLA2R antibody. Here, we asked whether genetic determinants could improve risk prediction. First, we sequenced PLA2R1 and HLA-D loci in 248 patients with primary membranous nephropathy and identified two independent single nucleotide polymorphisms (SNPs) at risk for primary membranous nephropathy at each locus. These were rs9271188 (intergenic between HLA-DRB1 and HLA-DQA1,) and rs9275086 (intergenic between HLA-DQB1 and HLA-DQA2) at the HLA-D locus along with rs6726925 and rs13018963 at the PLA2R1 locus. Then we investigated whether primary membranous nephropathy at-risk variants were associated with recurrence in a retrospective cohort of 105 donor-recipient pairs and a replication cohort of 40 pairs. Seven SNPs located between HLA-DRB1 and HLA-DQA1 in linkage disequilibrium with rs9271188, and three SNPs in the PLA2R1 region predicted recurrence when presented by the donor, but not when presented by the recipient. The two SNPs in the HLA-D region most strongly associated with recurrence (rs9271705 and rs9271550) were confirmed in the replication cohort. A genetic risk score based on the two best predictors at each locus (rs9271705, rs9271550, rs17830558, and rs3828323) identified a group of patients with high risk of recurrence. Thus, our results suggest that the graft contributes to recurrence of primary membranous nephropathy through the disease susceptibility HLA-D and PLA2R1 SNPs in an autoimmune milieu. Further studies are needed before implementation of genetic testing for these in donor selection., (Copyright © 2020. Published by Elsevier Inc.)

Membranous nephropathy (MN) is the most common cause of primary nephrotic syndrome among adults. The identification of phospholipase A2 receptor (PLA2R) as target antigen in most patients changed the management of MN dramatically, and provided a rationale for B-cell depleting agents such as rituximab. The efficacy of rituximab in inducing remission has been investigated in several studies, including 3 randomized controlled trials, in which complete and partial remission of proteinuria was achieved in approximately two-thirds of treated patients. Due to its favorable safety profile, rituximab is now considered a first-line treatment option for MN, especially in patients at moderate and high risk of deterioration in kidney function. However, questions remain about how to best use rituximab, including the optimal dosing regimen, a potential need for maintenance therapy, and assessment of long-term safety and efficacy outcomes. In this review, we provide an overview of the current literature and discuss both strengths and limitations of "the new standard.", (© 2021 Published by Elsevier, Inc., on behalf of the International Society of Nephrology.)

Primary forms of minimal change disease and focal segmental glomerulosclerosis are rare podocytopathies and clinically characterized by nephrotic syndrome. Glucocorticoids are the cornerstone of the initial immunosuppressive treatment in these two entities. Especially among adults with minimal change disease or focal segmental glomerulosclerosis, relapses, steroid dependence or resistance are common and necessitate re-initiation of steroids and other immunosuppressants. Effective steroid-sparing therapies and introduction of less toxic immunosuppressive agents are urgently needed to reduce undesirable side effects, in particular for patients whose disease course is complex. Rituximab, a B cell depleting monoclonal antibody, is increasingly used off-label in these circumstances, despite a low level of evidence for adult patients. Hence, critical questions concerning drug-safety, long-term efficacy and the optimal regimen for rituximab-treatment remain unanswered. Evidence in the form of large, multicenter studies and randomized controlled trials are urgently needed to overcome these limitations., Competing Interests: Declaration of Competing Interest VA reports grants from ADDMEDICA, outside the submitted work; AB reports consulting fees from Chemocentryx, from Merck/MSD and from Astra Zeneca, outside the submitted work; MB reports lecture fees from Boehringer Ingelheim, from Bristol-Myers Squibb, from Novartis, travel support from Hexal, lecture fees and travel support from Vifor Fresenius and lecture fee from Servier, outside the submitted work; CKC reports grants from GlaxoSmithKline, grants and consultancy fees from Retrophin, outside the submitted work; SF reports lecture fees from VIFOR Pharma, outside the submitted work; CK reports being funded by National Institute for Health Research (NIHR) as academic clinical fellow (ACF), during the conduct of the study. All the other authors declared no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Zinc fingers and homeoboxes (ZHX) proteins are heterodimeric transcriptional factors largely expressed at the cell membrane in podocytes in vivo. We found ZHX2-based heterodimers in podocytes, with ZHX2-ZHX1 predominantly at the cell membrane of the podocyte cell body, and ZHX2-ZHX3 at the slit diaphragm. In addition to changes in overall ZHX2 expression, there was increased podocyte nuclear ZHX3 and ZHX2 in patients with focal segmental glomerulosclerosis, and increased podocyte nuclear ZHX1 in patients with minimal change disease. Zhx2 deficient mice had increased podocyte ZHX1 and ZHX3 expression. Zhx2 deficient mice and podocyte specific Zhx2 overexpressing transgenic rats develop worse experimental focal segmental glomerulosclerosis than controls, with increased nuclear ZHX3 and ZHX2, respectively. By contrast, podocyte specific Zhx2 overexpressing transgenic rats develop lesser proteinuria during experimental minimal change disease due to peripheral sequestration of ZHX1 by ZHX2. Using co-immunoprecipitation, the interaction of ZHX2 with aminopeptidase A in the podocyte body cell membrane, and EPHRIN B1 in the slit diaphragm were noted to be central to upstream events in animal models of minimal change disease and focal segmental glomerulosclerosis, respectively. Mice deficient in Enpep, the gene for aminopeptidase A, and Efnb1, the gene for ephrin B1 developed worse albuminuria in glomerular disease models. Targeting aminopeptidase A in Zhx2 deficient mice with monoclonal antibodies induced albuminuria and upregulation of the minimal change disease mediator angiopoietin-like 4 through nuclear entry of ZHX1. Thus, podocyte ZHX2 imbalance is a critical factor in human glomerular disease, with minimal change disease disparities mediated mostly through ZHX1, and focal segmental glomerulosclerosis deviations through ZHX3 and ZHX2., (Copyright © 2019 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Immunoglobulin A (IgA) nephropathy (IgAN), the most common glomerulonephritis worldwide, is characterized by IgA depositions in the kidney. Deficiency of CD37, a leukocyte-specific tetraspanin, leads to spontaneous development of renal pathology resembling IgAN. However, the underlying molecular mechanism has not been resolved. Here we found that CD37 expression on B cells of patients with IgAN was significantly decreased compared to B cells of healthy donors. Circulating interleukin (IL)-6 levels, but not tumor necrosis factor-α or IL-10, were elevated in Cd37 -/- mice compared to wild-type mice after lipopolysaccharide treatment. Cd37 -/- mice displayed increased glomerular neutrophil influx, immune complex deposition, and worse renal function. To evaluate the role of IL-6 in the pathogenesis of accelerated renal pathology in Cd37 -/- mice, we generated Cd37xIl6 double-knockout mice. These double-knockout and Il6 -/- mice displayed no glomerular IgA deposition and were protected from exacerbated renal failure following lipopolysaccharide treatment. Moreover, kidneys of Cd37 -/- mice showed more mesangial proliferation, endothelial cell activation, podocyte activation, and segmental podocyte foot process effacement compared to the double-knockout mice, emphasizing that IL-6 mediates renal pathology in Cd37 -/- mice. Thus, our study indicates that CD37 may protect against IgA nephropathy by inhibition of the IL-6 pathway., (Copyright © 2018 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Background: Despite the increasing use of pre- and post-hydration protocols and low osmolar instead of high osmolar iodine containing contrast media, the incidence of contrast induced nephropathy (CIN) is still significant. There is evidence that contrast media cause ischemia reperfusion injury of the renal medulla. Remote ischemic preconditioning (RIPC) is a non-invasive, safe, and low cost method to reduce ischemia reperfusion injury. The aim of this study is to investigate whether RIPC, as an adjunct to standard preventive measures, reduces contrast induced acute kidney injury in patients at risk of CIN., Methods: The RIPCIN study is a multicenter, single blinded, randomized controlled trial in which 76 patients at risk of CIN received standard hydration combined with RIPC or hydration with sham preconditioning. RIPC was applied by four cycles of 5 min ischemia and 5 min reperfusion of the forearm. The primary outcome measure was the change in serum creatinine from baseline to 48 to 72 hours after contrast administration., Results: With regard to the primary endpoint, no significant effect of RIPC was found. CIN occurred in four patients (2 sham and 2 RIPC). A pre-defined subgroup analysis of patients with a Mehran risk score ≥11, showed a significantly reduced change in serum creatinine from baseline to 48 to 72 hours in patients allocated to the RIPC group (Δ creatinine -3.3 ± 9.8 μmol/L) compared with the sham group (Δ creatinine +17.8 ± 20.1 μmol/L)., Conclusion: RIPC, as an adjunct to standard preventive measures, does not improve serum creatinine levels after contrast administration in patients at risk of CIN according to the Dutch guideline. However, the present data indicate that RIPC might have beneficial effects in patients at a high or very high risk of CIN (Mehran score ≥ 11). The RIPCIN study is registered at: http://www.controlled-trials.com/ISRCTN76496973., (Copyright © 2015 European Society for Vascular Surgery. Published by Elsevier Ltd. All rights reserved.)

In rodents, parietal epithelial cells (PECs) migrating onto the glomerular tuft participate in the formation of focal segmental glomerulosclerosis (FSGS) lesions. We investigated whether immunohistologic detection of PEC markers in the initial biopsies of human patients with first manifestation of idiopathic nephrotic syndrome with no immune complexes can improve the sensitivity to detect sclerotic lesions compared with standard methods. Ninety-five renal biopsies were stained for claudin-1 (PEC marker), CD44 (activated PECs), and LKIV69 (PEC matrix); 38 had been diagnosed as early primary FSGS and 57 as minimal change disease. PEC markers were detected on the tuft in 87% of the biopsies of patients diagnosed as primary FSGS. PEC markers were detected in FSGS lesions from the earliest stages of disease. In minimal change disease, no PEC activation was observed by immunohistology. However, in 25% of biopsies originally diagnosed as minimal change disease the presence of small lesions indicative of a sclerosing process were detected, which were undetectable on standard periodic acid-Schiff staining, even though only a single histologic section for each PEC marker was evaluated. Staining for LKIV69 detected lesions with the highest sensitivity. Two novel PEC markers A-kinase anchor protein 12 and annexin A3 exhibited similar sensitivity. In summary, detection of PECs on the glomerular tuft by immunostaining improves the differentiation between minimal change disease and primary FSGS and may serve to guide clinical decision making., (Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

The Nijmegen Biomedical Study is a population-based cross-sectional study conducted in the eastern part of the Netherlands. As part of the overall study, we provide reference values of estimated glomerular filtration rate (GFR) for this Caucasian population without expressed risk. Age-stratified, randomly selected inhabitants received a postal questionnaire on lifestyle and medical history. In a large subset of the responders, serum creatinine was measured. The GFR was then measured using the abbreviated Modification of Diet in Renal Disease (MDRD) formula. To limit possible bias, serum creatinine was calibrated against measurements performed in the original MDRD laboratory. The study cohort included 2823 male and 3274 female Caucasian persons aged 18-90 years. A reference population of apparently healthy subjects was selected by excluding persons with known hypertension, diabetes, cardiovascular- or renal diseases. This healthy study cohort included 1660 male subjects and 2072 female subjects, of which 869 of both genders were 65 years or older. The median GFR was 85 ml/min/1.73 m(2) in 30-to 34-year-old men and 83 ml/min/1.73 m(2) in similar aged women. In these healthy persons, GFR declined approximately 0.4 ml/min/year. Our study provides age- and gender-specific reference values of GFR in a population of Caucasian persons without identifiable risk.

It has now been firmly established that, not only ischemia/reperfusion, but also cold itself causes damage during kidney transplantation. Iron chelators or anti-oxidants applied during the cold plus rewarming phase are able to prevent this damage. At present, it is unknown if these measures act only during the cold, or whether application during the rewarming phase also prevents damage. We aimed to study this after cold normoxic and hypoxic conditions. LLC-PK1 cells were incubated at 4 degrees C in Krebs-Henseleit buffer for 6 or 24h, followed by 18 or 6h rewarming, respectively. Cold preservation was performed under both normoxic (95% air/5% CO2) and hypoxic (95% N2/5% CO2) conditions. The iron chelator 2,2'-DPD (100 microM), anti-oxidants BHT (20 microM) or sibilinin (200 microM), and xanthine oxidase inhibitor allopurinol (100 microM) were added during either cold preservation plus rewarming, or rewarming alone. Cell damage was assessed by LDH release (n=3-9). Addition of 2,2'-DPD and BHT during cold hypoxia plus rewarming did, but during rewarming alone did not prevent cell damage. When added during rewarming after 6h cold normoxic incubation, BHT and 2,2'-DPD inhibited rewarming injury compared to control (p<0.05). Allopurinol did not prevent cell damage in any experimental set-up. Our data show that application of iron chelators or anti-oxidants during the rewarming phase protects cells after normoxic but not hypoxic incubation. Allopurinol had no effect. Since kidneys are hypoxic during transplantation, measures aimed at preventing cold-induced and rewarming injury should be taken during the cold.

Global glomerulosclerosis can be divided in the vascular (obsolescent) type and the glomerulopathic (solidified) type. In biopsies from children with recurrent nephrotic syndrome owing to minimal change nephropathy (MCN), we noticed small, globally sclerosed glomeruli that appeared to be distinct from global glomerulosclerosis. These small sclerosed glomeruli are best described as involuted glomeruli. We have characterized these involuted glomeruli in detail. We studied biopsies of 18 children (11 male, 7 female) with frequently relapsing MCN and evaluated possible explanatory variables. The involuted glomeruli can be differentiated from the other types of global glomerulosclerosis. Most notable is the presence of vital podocytes and parietal epithelial cells, which have retained their staining characteristics, in between the matrix, and the absence of periglomerular and tubulo-interstitial fibrosis. We observed involuted glomeruli in 12 out of 18 biopsies; the median percentage of involuted glomeruli was 6% (range 0-33%). The percentage of involuted glomeruli correlated with age at renal biopsy and the interval between onset of disease and time of renal biopsy, but not with gender, age at onset of disease, or prednisone dose. Multivariate analysis revealed that the interval between onset of disease and time of renal biopsy was the only independent predictor. In conclusion, glomerular involution is a special form of global glomerulosclerosis. The absence of periglomerular and tubulo-interstitial fibrosis suggests a different pathogenesis. Glomerular involution is a slow process. The clinical data suggest that involution is related to the duration of the disease process.

Recent studies have reported a significant increase of proteinuria in kidney transplant recipients who were switched from a calcineurin inhibitor (CI) to sirolimus. This has (partly) been ascribed to the hemodynamic renal effects of CI withdrawal. We have evaluated the evolution of proteinuria in renal transplant recipients who underwent conversion from azathioprine to sirolimus. In a randomized, prospective, multicenter study called RESCUE (Recurrent cutanEous Squamous cell Carcinoma Under RapamunE) the efficacy and safety is investigated of conversion to sirolimus in stable renal transplant recipients with a cutaneus squamous cell carcinoma (SCC). In our center 25 patients have been included in this study of which 13 patients were randomized to continue their current immunosuppressive treatment and 12 to conversion to sirolimus. After a mean follow-up of 360 days mean proteinuria increased from 0.37+/-0.34 to 1.81+/-1.73 g/24 h after conversion to sirolimus (P<0.005). In the control group there was no change in proteinuria. A significant increase of proteinuria was observed in all seven patients with proteinuria before conversion, whereas proteinuria remained absent in all patients without previous proteinuria. Two of the patients with proteinuria were converted from cyclosporine and five were converted from azathioprine to sirolimus. Sirolimus was discontinued in five patients with proteinuria, and in all of them proteinuria declined to baseline values. Our study demonstrates that conversion from azathioprine to sirolimus after kidney transplantation may cause a reversible increase of proteinuria. Sirolimus-induced proteinuria therefore cannot be ascribed to the hemodynamic renal effects of withdrawal of CI.

Collapsing focal segmental glomerulosclerosis (cFSGS) is characterized by hyperplasia of glomerular epithelial cells. In a mouse model of FSGS and in a patient with recurrent idiopathic FSGS, we identified the proliferating cells as parietal epithelial cells (PECs). In the present study, we have evaluated the origin of the proliferating cells in cFSGS associated with human immunodeficiency virus (HIV) and pamidronate. We performed a detailed study of glomerular lesions in biopsies of two patients with HIV-associated cFSGS and a nephrectomy specimen of a patient with pamidronate-associated cFSGS. Glomeruli were studied by serial sectioning using light and electron microscopy and immunohistochemistry to determine the epithelial cell phenotype. We used Synaptopodin, vascular endothelial growth factor, and CD10 as podocyte markers, CK8 and PAX2 as PEC markers and Ki-67 as marker of cell proliferation. The newly deposited extracellular matrix was characterized using antiheparan sulfate single-chain antibodies. The proliferating cells were negative for the podocyte markers, but stained positive for the PEC markers and the cell proliferation marker Ki-67. The proliferating PAX-2 and CK8 positive cells that covered the capillary tuft were always in continuity with PAX-2/CK8 positive cells lining Bowman's capsule. The matrix deposited by these proliferating cells stained identically to Bowman's capsule. Our study demonstrates that PECs proliferate in HIV and pamidronate-associated cFSGS. Our data do not support the concept of the proliferating, dedifferentiated podocyte.

Sirolimus is a new and potent immunosuppressive agent. Recently, increased proteinuria has been recognized as an important complication. However, the mechanism thereof has remained unclear. We describe a patient who received sirolimus as standard therapy after living donor kidney transplantation. Within 10 days the patient developed a severe proteinuria that disappeared completely after substituting tacrolimus for sirolimus. Renal biopsy disclosed normal glomeruli even without effacement of the podocytic foot processes. Using FITC labeled anti-albumin antibodies we noted complete absence of albumin in the proximal tubules, whereas an abundant albumin staining was observed in a control patient with a comparable level of proteinuria due to a recurrence of focal segmental glomerulosclerosis after transplantation. Our data suggest that sirolimus can induce severe proteinuria, and that reduced tubular protein reabsorption contributes to the protein loss.

Background: Focal segmental glomerulosclerosis (FSGS) is one of the most common patterns of glomerular injury encountered in human renal biopsies. Epithelial hyperplasia, which can be prominent in FSGS, has been attributed to dedifferentiation and proliferation of podocytes. Based on observations in a mouse model of FSGS, we pointed to the role of parietal epithelial cells (PECs). In the present study we investigated the relative role of PECs and podocytes in human idiopathic FSGS., Methods: We performed a detailed study of lesions from a patient with recurrent idiopathic FSGS by serial sectioning, marker analysis and three-dimensional reconstruction of glomeruli. We have studied the expression of markers for podocytes, PECs, mesangial cells, endothelium, and myofibroblasts. We also looked at proliferation and composition of the deposited extracellular matrix (ECM)., Results: We found that proliferating epithelial cells in FSGS lesions are negative for podocyte and macrophage markers, but stain for PEC markers. The composition of the matrix deposited by these cells is identical to Bowman's capsule., Conclusion: Our study demonstrates that PECs are crucially involved in the pathogenesis of FSGS lesions.

Background: Nephrotic syndrome in childhood is mainly due to minimal change nephropathy. In general, it is characterized by selective proteinuria, by steroid responsiveness, and histologically by podocytic foot process effacement. Familial presentation is rare and mainly restricted to one generation., Methods: We describe the occurrence of a familial nephropathy in a mother and two daughters. An initial diagnosis of minimal change nephropathy was made, but subsequently unique features became apparent. During follow-up, detailed studies of renal function and urinary protein excretion were performed. Available frozen renal biopsy material was revised and processed for immunofluorescence to detect abnormalities in the expression of heparan sulfate proteoglycans. The latter results were compared with renal biopsies of a control group composed of five adult patients with minimal change nephropathy., Results: The mother and two daughters were proteinuric since their early childhood. The mother revealed a persistent nephrotic syndrome for more than 20 years despite treatment with various immunosuppressive drugs. Likewise, treatment with prednisone was ineffective in the daughters. All three patients retained normal renal function during follow-up. Detailed measurements revealed that the proteinuria was incredibly selective (selectivity index approximately 0.01), and there was no evidence of tubulointerstitial damage, as reflected by a normal excretion of the low-molecular weight proteins beta(2)-microglobulin and alpha1-microglobulin. Renal biopsy performed in the mother and one daughter was thought to be compatible with minimal change nephropathy. However, histologically, two remarkable findings were made. By electron microscopy, there was no evidence of foot process retraction; specifically, the foot process width and slit pore diameter were normal. Furthermore, in contrast to the control patients, the expression of heparan sulfate polysaccharide side chains, as reflected by the staining with monoclonal antibody JM403, was normal., Conclusions: We propose that this family represents a new familial nephropathy. The molecular basis of the permeability defect remains to be identified.

Background: Aminopeptidase A is an enzyme that is present on podocytes and is involved in the degradation of angiotensin II. In previous studies in mice, we administered single monoclonal antibodies directed against aminopeptidase A. We observed that only monoclonal antibodies that inhibited aminopeptidase A enzyme activity caused albuminuria., Methods: In this study, the effects of the combined injections of two monoclonal anti-aminopeptidase A antibodies (mAbs) were studied, using a combination of anti-aminopeptidase A mAbs that were directed against two different domains involved in the aminopeptidase A enzyme activity (ASD-3 or ASD-37) and an anti-aminopeptidase A mAb not related to the enzyme active site (ASD-41)., Results: An injection of the combinations ASD-3/37 (total 4 mg, 1:1 ratio) and ASD-37/41 (total 4 mg, 1:1 ratio) in doses that do not cause albuminuria when given alone (4 mg) induced massive albuminuria at day 1 after injection. The combination ASD-3/41 had no effect. This albuminuria was not dependent on systemic immune mediators of inflammation and could not merely be related to a blockade of aminopeptidase A enzyme activity. However, a correlation was observed between the induction of albuminuria and the aggregation of the mAbs injected and aminopeptidase A on the podocytes. An injection of the combinations ASD-3/37 or ASD-37/41 did not cause an increase in systemic blood pressure. The treatment with a combination of enalapril and losartan lowered blood pressure (53 +/- 10 vs. 90 +/- 3 mm Hg in untreated mice) and reduced the acute albuminuria by 55% (11,145 +/- 864 vs. 24,517 +/- 2448 micrograms albumin/18 hr in untreated mice). However, similar effects were observed using triple therapy. Therefore, the reduction of albuminuria by the combined treatment of enalapril/losartan seems to be the consequence of the reduction in the systemic blood pressure. These findings argue against a specific role for angiotensin II in this model., Conclusions: The combined injection of two mAbs directed against different domains of aminopeptidase A induces a massive albuminuria in mice, which is not merely dependent on angiotensin II. We hypothesize that the direct binding of mAbs to at least two pathogenic domains on aminopeptidase A triggers the podocyte to release mediators that are involved in the observed albuminuria.

In patients with proteinuric renal diseases the rate of progression of renal insufficiency is determined by the level of blood pressure and proteinuria. It has been demonstrated that strict blood pressure control with angiotensin converting enzyme (ACE)-inhibitors or beta-blockers, aimed at reaching values below 130/80 mm Hg, attenuates the deterioration of renal function. In general, the beneficial effects of these drugs are reflected in a parallel lowering of proteinuria. Calcium channel blockers are effective antihypertensive drugs, however, their safety in patients with proteinuric renal diseases and renal insufficiency may be questioned because of reported untoward effects on urinary protein excretion. The present review discusses the potential benefits and risks of calcium channel blockers (CCBs) in the treatment of patients with renal diseases. To this end we have evaluated the effects of these drugs in animal models of progressive renal injury. In these animal models adverse effects of CCBs have been reported which are attributed to an impairment of autoregulation. In patients with proteinuria, the dihydropyridine CCBs do not lower proteinuria despite a reduction of blood pressure. Studies on the effects on the course of renal function are limited, however, the available data do suggest that this class of CCBs may be less advantageous than other antihypertensive drugs, thus arguing against the use of these agents as first-line drugs in patients with proteinuric renal diseases. Information on the effects of the non-dihydropyridine CCBs is limited to a small number of studies in patients with diabetic renal disease. Although the data suggest that these classes of CCBs might be more beneficial, more studies are needed, particularly in patients with non-diabetic renal diseases, before founded conclusions can be reached.

Renal ischemia results in adenosine triphosphate (ATP) depletion, particularly in cells of the proximal tubule (PT), which rely heavily on oxidative phosphorylation for energy supply. Lack of ATP leads to a disturbance in intracellular homeostasis of Na+, K+ and Cl-. Also, cytosolic Ca2+ levels in renal PTs may increase during hypoxia [1], presumably by a combination of impaired extrusion and enhanced influx [2]. However, Ca2+ influx was previously measured using radiolabeled Ca2+ and at varying partial oxygen tension [2]. We have now used to Mn2(+)-induced quenching of fura-2 fluorescence to study Ca2+ influx in individual rat PTs during normoxic and hypoxic superfusion. Normoxic Ca2+ influx was indeed reflected by the Mn2+ quenching of fura-2 fluorescence and this influx could be inhibited by the calcium entry blocker methoxyverapamil (D600; inhibition 50 +/- 2% and 35 +/- 3% for 10 and 100 mumol, respectively). La3+ completely blocked normoxic Ca2+ influx. Hypoxic superfusion or rat PTs did not induce an increase in Ca2+ influx, but reduced this influx to 79 +/- 3% of the normoxic control. We hypothesize that reducing Ca2+ influx during hypoxia provides the cell with a means to prevent cellular Ca2+ overload during ATP-depletion, where Ca2+ extrusion is limited.

To elucidate potential mechanisms of ischemic renal injury, investigators often use drugs that interfere with specific pathological pathways and study their protective efficacy in in vitro models of ischemia, such as isolated renal proximal tubules subjected to hypoxia. However, the protective effects of certain drugs may depend on non-specific membrane-stabilizing properties. We have studied the effects of several drugs on membrane integrity using osmotic lysis of erythrocytes as a model system. Freshly isolated rabbit erythrocytes were subjected to a hypotonic shock, and the protective effects of various calcium channel blockers, phospholipase inhibitors, free fatty acids, the NO-synthase inhibitor L-NAME, the amino acid glycine and its receptor-analogue strychnine, and two chloride channel blockers were examined. Most agents protected erythrocytes against hypotonic hemolysis when added to the medium in the same concentration range as used in suspensions of hypoxic proximal tubules. Only the protective agents that proposedly act via a blockade of chloride influx (glycine, strychnine and the chloride channel blockers), did not attenuate hypotonic hemolysis. The erythrocyte hemolysis assay may provide an easy and rapid method to screen for non-specific membrane-stabilizing effects of potentially cytoprotective agents.

Recent in vitro studies have demonstrated that the amino acid glycine affords profound protection against hypoxic injury in isolated rat proximal tubules and the isolated perfused rat kidney. In the present study we have examined the in vivo effect of glycine in a rat model of ischemic acute renal failure. Acute renal failure was induced in rats by right uninephrectomy and clamping of the left renal artery for 45 or 60 minutes. Glycine infusion was initiated 60 minutes before renal artery clamping at a rate to increase serum glycine levels above 2.0 mmol/L. Control rats received a 5% dextrose infusion. In the control rats, both 45 minutes and 60 minutes of clamping resulted in reversible acute renal failure and a serum creatinine level after 24 hours averaging 2.1 +/- 0.3 mg/dl (mean +/- SEM) and 3.3 +/- 0.4 mg/dl, respectively. Glycine treatment did not attenuate the decrease of renal function in the 45-minute and 60-minute models, with serum creatinine averaging 2.1 +/- 0.4 mg/dl and 3.4 +/- 0.5 mg/dl, respectively. Histologic examination also did not reveal any differences between control and glycine-treated rats. Therefore, we conclude that infusion of glycine does not afford in vivo protection against acute ischemic renal injury in the rat. The in vitro protective effects of glycine therefore cannot be extrapolated to the in vivo situation.

The effects of phosphate (PO4) removal from Krebs Henseleit buffer on freshly isolated rat proximal tubules (rPT) were assessed by measuring Ca2+ uptake (nmol/mg protein), cellular adenosine triphosphate (ATP) (nmol/mg), tissue K+ content (nmol/mg) and lactate dehydrogenase (LDH) as an index of cell integrity. Ca2+ uptake increased by 50% in rPT incubated in zero PO4 medium as compared to control (2.6 +/- 0.1 vs. 3.9 +/- 0.19, P less than 0.001) and LDH release increased 2.5-fold from 14.2 +/- 0.6 to 31.6 +/- 1.6%, P less than 0.001. Neither verapamil (200 microM) nor mepacrine (50 microM) reduced Ca2+ uptake or decreased LDH release suggesting that the increased Ca2+ uptake was not occurring through potential operated channels and that phospholipase-induced cell injury was not the cause of increased LDH release. Either glycine (2 mM) or extracellular fluid acidosis (pH 7.06), however, significantly diminished rPT injury and Ca2+ uptake. Specifically, as compared to the increased LDH released in untreated. PO4-depleted rPT, LDH release was diminished significantly by glycine treatment (31.0 +/- 0.9 vs. 15.5 +/- 1.6%, P less than 0.001) or acidosis (30.3 +/- 0.04 vs. 19.2 +/- 0.9%, P less than 0.01). Ca2+ uptake did not increase in glycine treated tubules (2.6 +/- 0.1 vs. 2.8 +/- 0.2 nmol/mg, NS) or in the presence of acidosis (2.6 +/- 0.1 vs. 2.97 +/- 0.17 nmol/mg, NS). ATP concentrations were markedly reduced by PO4 depletion (2.8 +/- 0.2 vs. 4.8 +/- 0.3 nmol/mg, P less than 0.001) and remained at low levels during either acidosis or glycine-induced protection.(ABSTRACT TRUNCATED AT 250 WORDS)

Genetic predisposition to essential hypertension has been proposed as a risk factor for the development of diabetic nephropathy in type 1 (insulin-dependent) diabetes mellitus. An increased sodium-lithium countertransport activity (NaLiCT) has been suggested as a genetic marker for essential hypertension. We therefore evaluated NaLiCT in diabetic patients with (N = 39) or without (N = 23) diabetic nephropathy (DNP), patients with non-diabetic renal diseases (N = 42) and in healthy controls (N = 24). The NaLiCT was elevated in both diabetic patient groups compared to healthy controls (median 244; range 134 to 390 mumol.liter cells-1.hr-1), but was not different in patients with DNP (median 314; range 162 to 676), without DNP (median 325; range 189 to 627) and patients with non-diabetic renal disease (median 300; range 142 to 655). The genetic predisposition to DNP is illustrated by the fact that diabetic sibs of probands with DNP showed a higher occurrence of DNP than diabetic sibs of patients without DNP. We analyzed whether familial DNP clustered with an increased NaLiCT. The NaLiCT in sibs concordant for the presence of DNP (N = 10; median 307; range 217 to 428 mumol.liter cells-1.hr-1) was not significantly different from that in sibs concordant for absence of DNP (N = 15; median 279; range 189 to 442). We conclude that erythrocyte sodium-lithium countertransport activity cannot be used as a marker to identify patients at risk for the development of diabetic nephropathy.

Results from animal experiments have suggested that treatment with recombinant human erythropoietin (rHuEPO) causes changes in renal hemodynamics which are detrimental to renal function. Therefore, the effects of correction of the anemia by rHuEPO on glomerular filtration rate (GFR; inulin clearance) and effective renal plasma flow (ERPF; PAH clearance) were studied in eight pre-dialysis patients. The studies were done before (Hct 0.24 +/- 0.05 liter/liter) and at 89 +/- 19 days after the start of rHuEPO therapy (Hct 0.39 +/- 0.03 liter/liter). To further evaluate the effects of ACE inhibition, 25 mg of captopril was given orally after baseline values had been obtained. Baseline GFR, renal blood flow (RBF) and filtration fraction (FF) did not change during rHuEPO therapy. At low hematocrit (Hct) captopril induced a significant increase in ERPF and RBF, and a decrease in MAP. After correction of the hematocrit the blood pressure lowering effect of captopril remained unchanged. However, captopril no longer induced changes in ERPF and RBF. We conclude that the increase in hematocrit had no adverse effects on GFR. The results suggest that changes in hematocrit may influence the effects of ACE inhibition on efferent vascular resistance. Therefore, the hematocrit should be taken into account when evaluating studies on the effects of ACE inhibition in the progression of chronic renal failure.

To investigate whether the administration of cimetidine can improve the reliability of creatinine as a marker of GFR, we compared the creatinine clearance (CCr) to the clearance of the true filtration markers 51Cr-EDTA (CEDTA) and inulin (CIn), after oral ingestion of cimetidine in 10 healthy men and 29 patients with varying degrees of renal dysfunction. After administration of cimetidine for three to six days, serum creatinine level rose in all participants, while CEDTA and CIn remained stable in a subgroup of 14 subjects in whom they were measured before as well as after the administration of cimetidine. The mean (+/- SD) ratios of CCr to CEDTA (N = 39) and of CCr to CIn (N = 19) after ingestion of cimetidine were 1.02 +/- 0.13 and 1.01 +/- 0.13, respectively, and did not differ significantly from unity. This high degree of accuracy of the cimetidine-aided CCr was present over the entire range of renal function in the study population. Our results also indicated an improved precision of the cimetidine-aided measurement of CCr, resulting in a variability that did not differ significantly from that of the measurement of CEDTA or CIn. We conclude that after oral administration of cimetidine, the creatinine clearance can be used as a reliable measure of GFR within a broad range of renal function.

In patients with a nephrotic syndrome administration of prednisolone causes an increase of proteinuria. To elucidate the mechanism of this effect we have studied the acute proteinuric effect of prednisolone, 125 to 150 mg intravenously, in nine patients (7 M, 2F) with a nephrotic syndrome. Mean age (+/- SD) of the patients was 53 +/- 6 years, mean endogenous creatinine clearance 104 +/- 30 ml/min, and mean proteinuria 7.7 +/- 3.0 g/24 hr. After administration of prednisolone, urinary total protein excretion rose in all patients from a mean (+/- SEM) of 4.89 +/- 0.59 mg/min before to 9.09 +/- 0.99 mg/min at five hours after administration (P less than 0.01). Glomerular filtration rate (inulin clearance), effective renal plasma flow (PAH clearance), and filtration fraction did not change significantly. The increases of urinary excretion of albumin (median %: +92%), IgG (median %: +88%), and transferrin (median %: +76%) were comparable and correlated significantly. Urinary excretion of beta 2-microglobulin did not change significantly however. We conclude that intravenous administration of prednisolone to patients with a nephrotic syndrome causes an increase in urinary protein excretion rate which cannot be explained by changes in renal hemodynamics or tubular protein reabsorption, and which therefore must be the result of a change in glomerular permselectivity characteristics.

In a prospective randomised trial, 72 recipients of cadaver renal allografts received cyclosporin for 3 months followed by azathioprine and prednisone (cyclosporin group), and 71 received azathioprine and prednisone from the day of transplantation (conventional group). Graft survival was better in the cyclosporin group at 3 months and 1 year (93% and 80%) than in the conventional group (83% and 70%). This was not a significant difference. The incidence of acute rejection episodes in the first 3 months was significantly lower in the cyclosporin group (35% versus 77%, p less than 0.00001), as was the number of grafts lost because of immunological failure (1 versus 10, p less than 0.02). After conversion, renal function improved. Only 5 patients had acute rejection after conversion. These episodes were easily reversible in all cases and did not lead to graft loss. The numbers of grafts lost after conversion were similar in the two groups. Conversion of cyclosporin to azathioprine 3 months after renal transplantation is a safe procedure that obviates the long-term toxic effects of cyclosporin.