Cause Of Steroid-resistant Nephrotic Syndrome Research Articles

Phenotypic quantification of Nphs1-deficient mice.

Steroid-resistant nephrotic syndrome (SRNS) is the second most frequent cause of chronic kidney disease in children and young adults. The most severe form of steroid-resistant nephrotic syndrome is congenital nephrotic syndrome Finnish type (CNSF), caused by biallelic loss-of-function variants inNPHS1,encoding nephrin. Since each of the 68 monogenic causes of steroid-resistant nephrotic syndrome represents a rare cause of the disease, tailoring therapeutic interventions to multiple molecular targets remains challenging, suggesting gene replacement therapy (GRT) as a viable alternative. To set the ground for a gene replacement study in vivo, we established rigorous, quantifiable, and reproducible phenotypic assessment of a conditional Nphs1 knockout mouse model. By breeding a floxed Nphs1fl/- mouse (Nphs1tm1Afrn/J) previously studied for pancreatic β-cell survival with a podocin promoter-driven Cre recombinase mouse model (Tg(NPHS2-Cre)295Lbh/J), we generated mice with podocyte-specific nephrin deficiency (Nphs1fl/fl NPHS2-Cre +). We observed a median survival to postnatal day P5 in nephrin-deficient mice, whereas heterozygous control mice and wild type (WT) control group showed 90% and 100% survival, respectively (at P50 days). Light microscopy analysis showed a significantly higher number of renal-tubular microcysts per kidney section in nephrin-deficient mice compared to the control groups (P < 0.0022). Transmission electron microscopy demonstrated reduced foot process (FP) density in nephrin-deficient mice compared to controls (P < 0.0001). Additionally, proteinuria quantitation using urine albumin-to-creatinine ratio (UACR) was significantly higher in nephrin-deficient mice compared to controls. This study represents the first comprehensive description of the kidney phenotype in anephrin-deficient mouse model, laying the foundation for future gene replacement therapy endeavors.

Estimation of the Age of the Kashubian-Specific Pathogenic NPHS2 Variant Responsible for Hereditary Steroid-Resistant Nephrotic Syndrome Points to Its Recent Local Origin

Steroid-resistant nephrotic syndrome (SRNS) is a highly heterogenic kidney disorder resulting from genetic abnormalities or immune system dysfunction affecting the establishment and maintenance of the glomerular filtration barrier. The most common cause of genetic SRNS is biallelic pathogenic variants in NPHS2 gene, especially in individuals with an infantile or childhood onset. The type of the NPHS2 defect implies the course of the disease and the stage of its onset and differs across populations. In a cohort of Polish patients with SRNS, a unique profile of the disease-related NPHS2 variants was identified in patients from northern Poland inhabited by Kashubs, a minority West-Slavic ethnic group known for a local increase of the frequency of several pathogenic variants. Among Kashubian families, the compound heterozygotes c.686G&gt;A/c.1032delT and a single c.1032delT homozygote were the only underlying cause of SRNS. The restricted, Kashubian-only pattern of c.1032delT occurrence, suggesting the founder effect, prompted us to conduct a detailed analysis of its haplotype background to estimate the age of the c.1032delT origin. Eight Kashubian SRNS families were genotyped using the Infinium Global Screening Array-24. The haplotype background analysis was performed using an in-house pipeline designed to solve the phase of the heterozygous genotype data. The age of the c.1032delT mutation was calculated using the gamma method based on the genetic length of ancestral haplotypes shared between two or more individuals carrying this variant. The results of our study indicated a very recent origin of the c.1032delT mutation (~240 years). Genetic screening performed in the general Polish population control corroborates the assumption that the mutation occurred on the specific Kashubian haplotype background. The identification of ancestry-specific Kashubian pathogenic variant can help to develop effective screening and diagnostic strategies as a part of personalized medicine approach in the region.

In vivo characterization of a podocyte-expressed short podocin isoform

The most common genetic causes of steroid-resistant nephrotic syndrome (SRNS) are mutations in the NPHS2 gene, which encodes the cholesterol-binding, lipid-raft associated protein podocin. Mass spectrometry and cDNA sequencing revealed the existence of a second shorter isoform in the human kidney in addition to the well-studied canonical full-length protein. Distinct subcellular localization of the shorter isoform that lacks part of the conserved PHB domain suggested a physiological role. Here, we analyzed whether this protein can substitute for the canonical full-length protein. The short isoform of podocin is not found in other organisms except humans. We therefore analysed a mouse line expressing the equivalent podocin isoform (podocinΔexon5) by CRISPR/Cas-mediated genome editing. We characterized the phenotype of these mice expressing podocinΔexon5 and used targeted mass spectrometry and qPCR to compare protein and mRNA levels of podocinwildtype and podocinΔexon5. After immunolabeling slit diaphragm components, STED microscopy was applied to visualize alterations of the podocytes’ foot process morphology.Mice homozygous for podocinΔexon5 were born heavily albuminuric and did not survive past the first 24 h after birth. Targeted mass spectrometry revealed massively decreased protein levels of podocinΔexon5, whereas mRNA abundance was not different from the canonical form of podocin. STED microscopy revealed the complete absence of podocin at the podocytes’ slit diaphragm and severe morphological alterations of podocyte foot processes. Mice heterozygous for podocinΔexon5 were phenotypically and morphologically unaffected despite decreased podocin and nephrin protein levels.The murine equivalent to the human short isoform of podocin cannot stabilize the lipid-protein complex at the podocyte slit diaphragm. Reduction of podocin levels at the site of the slit diaphragm complex has a detrimental effect on podocyte function and morphology. It is associated with decreased protein abundance of nephrin, the central component of the filtration-slit forming slit diaphragm protein complex.

Monogenic Causes Identified in 23.68% of Children with Steroid-Resistant Nephrotic Syndrome: A Single-Centre Study

Introduction: Steroid-resistant nephrotic syndrome (SRNS) is the second most common cause of end-stage kidney disease in children, mostly associated with focal segmental glomerulosclerosis (FSGS). Advances in genomic science have enabled the identification of causative variants in 20–30% of SRNS patients. Methods: We used whole exome sequencing to explore the genetic causes of SRNS in children. Totally, 101 patients with SRNS and 13 patients with nephrotic proteinuria and FSGS were retrospectively enrolled in our hospital between 2018 and 2022. For the known monogenic causes analysis, we generated a known SRNS gene list of 71 genes through reviewing the OMIM database and literature. Results: Causative variants were identified in 23.68% of our cohort, and the most frequently mutated genes in our cohort were WT1 (7/27), NPHS1 (3/27), ADCK4 (3/27), and ANLN (2/27). Five patients carried variants in phenocopy genes, including MYH9, MAFB, TTC21B, AGRN, and FAT4. The variant detection rate was the highest in the two subtype groups with congenital nephrotic syndrome and syndromic SRNS. In total, 68.75% of variants we identified were novel and have not been previously reported in the literature. Conclusion: Comprehensive genetic analysis is key to realizing the clinical benefits of a genetic diagnosis. We suggest that all children with SRNS undergo genetic testing, especially those with early-onset and extrarenal phenotypes.

Copy number variation analysis in 138 families with steroid-resistant nephrotic syndrome identifies causal homozygous deletions in PLCE1 and NPHS2 in two families.

Steroid-resistant nephrotic syndrome (SRNS) is the second most common cause of kidney failure in children and adults under the age of 20years. Previously, we were able to detect by exome sequencing (ES) a known monogenic cause of SRNS in 25-30% of affected families. However, ES falls short of detecting copy number variants (CNV). Therefore, we hypothesized that causal CNVs could be detected in a large SRNS cohort. We performed genome-wide single nucleotide polymorphism (SNP)-based CNV analysis on a cohort of 138 SRNS families, in whom we previously did not identify a genetic cause through ES. We evaluated ES and CNV data for variants in 60 known SRNS genes and in 13 genes in which variants are known to cause a phenocopy of SRNS. We applied previously published, predefined criteria for CNV evaluation. We detected a novel CNV in two genes in 2 out of 138 families (1.5%). The 9,673bp homozygous deletion in PLCE1 and the 6,790bp homozygous deletion in NPHS2 were confirmed across the breakpoints by PCR and Sanger sequencing. We confirmed that CNV analysis can identify the genetic cause in SRNS families that remained unsolved after ES. Though the rate of detected CNVs is minor, CNV analysis can be used when there are no other genetic causes identified. Causative CNVs are less common in SRNS than in other monogenic kidney diseases, such as congenital anomalies of the kidneys and urinary tract, where the detection rate was 5.3%. A higher resolution version of the Graphical abstract is available as Supplementary information.

Genetic analysis and outcomes of Omani children with steroid-resistant nephrotic syndrome.

Nephrotic syndrome (NS) is one of the most common kidney disorders seen by pediatric nephrologists and is defined by the presence of heavy proteinuria (>3.5 g/24 h), hypoalbuminemia (<3.5 g/dL), edema, and hyperlipidemia. Most children with NS are steroid-responsive and have a good prognosis following treatment with prednisolone. However, 10%-20% of them have steroid-resistant nephrotic syndrome (SRNS) and fail to respond to treatment. A significant proportion of these children progress to kidney failure. This retrospective study aimed to determine the underlying genetic causes of SRNS among Omani children below 13 years old, over a 15-year period and included 77 children from 50 different families. We used targeted Sanger sequencing combined with next-generation sequencing approaches to perform molecular diagnostics. We found a high rate of underlying genetic causes of SRNS in 61 (79.2%) children with pathogenic variants in the associated genes. Most of these genetically solved SRNS patients were born to consanguineous parents and variants were in the homozygous state. Pathogenic variants in NPHS2 were the most common cause of SRNS in our study seen in 37 (48.05%) cases. Pathogenic variants in NPHS1 were also seen in 16 cases, especially in infants with congenital nephrotic syndrome (CNS). Other genetic causes identified included pathogenic variants in LAMB2, PLCE1, MYO1E, and NUP93. NPHS2 and NPHS1 genetic variants were the most common inherited causes of SRNS in Omani children. However, patients with variants in several other SRNS causative genes were also identified. We recommend screening for all genes responsible for SRNS in all children who present with this phenotype, which will assist in clinical management decisions and genetic counseling for the affected families.

Kidney Organoids Generated Using an Allelic Series of NPHS2 Point Variants Reveal Distinct Intracellular Podocin Mistrafficking.

NPHS2 variants are the most common cause of steroid-resistant nephrotic syndrome in children >1 month old. Missense NPHS2 variants were reported to cause mistrafficking of the encoded protein, PODOCIN, but this conclusion was on the basis of overexpression in some nonpodocyte cell lines. We generated a series of human induced pluripotent stem cell (iPSC) lines bearing pathogenic missense variants of NPHS2 , encoding the protein changes p.G92C, p.P118L, p.R138Q, p.R168H, and p.R291W, and control lines. iPSC lines were also generated from a patient with steroid-resistant nephrotic syndrome (p.R168H homozygote) and a healthy heterozygous parent. All lines were differentiated into kidney organoids. Immunofluorescence assessed PODOCIN expression and subcellular localization. Podocytes were transcriptionally profiled and PODOCIN-NEPHRIN interaction interrogated. All variant lines revealed reduced levels of PODOCIN protein in the absence of reduced transcription. Although wild-type PODOCIN localized to the membrane, distinct variant proteins displayed unique patterns of subcellular protein trafficking, some unreported. P118L and R138Q were preferentially retained in the endoplasmic reticulum (ER); R168H and R291W accumulated in the Golgi. Podocyte profiling demonstrated minimal disease-associated transcriptional change. All variants displayed podocyte-specific apoptosis, which was not linked to ER stress. NEPHRIN-PODOCIN colocalization elucidated the variant-specific effect on NEPHRIN association and hence NEPHRIN trafficking. Specific variants of endogenous NPHS2 result in distinct subcellular PODOCIN localization within organoid podocytes. Understanding the effect of each variant on protein levels and localization and the effect on NEPHRIN provides additional insight into the pathobiology of NPHS2 variants. This article contains a podcast at https://dts.podtrac.com/redirect.mp3/www.asn-online.org/media/podcast/JASN/2023_01_05_JASN2022060707.mp3.

Whole exome sequencing identifies monogenic forms of nephritis in a previously unsolved cohort of children with steroid-resistant nephrotic syndrome and hematuria.

Alport syndrome (AS), atypical hemolytic-uremic syndrome (aHUS), and fibronectin-glomerulopathy (FG) are rare forms of glomerular diseases that manifest in a combination of proteinuria, hematuria, and hypertension, referred to as nephritic syndrome. Due to phenotypic overlays, steroid-resistant nephrotic syndrome (SRNS) and nephritic syndrome have been difficult to discern diagnostically. SRNS is more common than nephritic syndrome and is the second leading cause of childhood-onset CKD. Fourteen monogenic causes of AS, aHUS, and FG and 60 monogenic causes of SRNS have been identified. As whole exome sequencing (WES) allows for unequivocal molecular genetic diagnostics, we hypothesize to be able to identify causative mutations in genes known to cause nephritic syndrome in patient cohorts with a clinical diagnosis of SRNS. We identified patients with hematuria and steroid-resistant proteinuria in an international patient cohort that we had submitted to WES and who were unsolved for known monogenic causes of SRNS. These 70 patients from 65 individual families were subsequently analyzed for causative mutations in 14 AS, aHUS, or FG causing genes. WES data were compared to a control cohort of 76 patients from 75 families that were diagnosed with nephronophthisis-related ciliopathies (NPHP-RC) and to a control cohort of 83 individuals from 75 families with SRNS, but without hematuria. We detected likely pathogenic genetic variants in 3 of 65 families (4.6%) in 2 of the 14 genes analyzed. We confirmed that in cohorts of childhood-onset SRNS, patients with nephritic syndrome can be discerned by WES. The findings highlight the importance of clinical genetic testing for therapeutic and preventative measures in patients with proteinuria. A higher resolution version of the Graphical abstract is available as Supplementary information.

Role of fibroblast specific protein 1 expression in the progression of adriamycin-induced glomerulosclerosis

Focal segmental glomerulosclerosis (FSGS) is a commonly occurring cause of steroid-resistant nephrotic syndrome and frequently progresses to renal failure. Podocyte epithelial-mesenchymal transition (EMT) is thought to induce podocyte detachment in glomerular diseases, and severe degeneration and shedding of glomerular podocytes plays a major role in the progression of FSGS. We showed that fibroblast specific protein 1 (FSP1), an EMT marker, is strongly expressed in podocytes of FSGS patients, but the significance of podocyte expression of FSP1 to the pathophysiology of FSGS remained unclear. Here, we investigated FSP1 expression in podocytes from mice with adriamycin (ADR)-induced nephropathy, a murine model of FSGS. The number of FSP1-positive (FSP1+) podocytes was increased in ADR-treated mice and positively correlated with the degree of proteinuria and glomerulosclerosis in ADR-treated mice. ADR-induced FSGS and the attendant proteinuria were significantly ameliorated in FSP1 knockout mice as compared to wild type mice. These findings indicate that podocyte expression of FSP1 plays a crucial role in the pathogenesis of FSGS, which makes FSP1 a potential target for treatment of FSGS.

MO1027EFFICACY OF CALCINEURIN INHIBITORS IN CHILDREN WITH MONOGENIC STEROID-RESISTANT NEPHROTIC SYNDROME

Abstract Background and Aims Monogenic causes of steroid-resistant nephrotic syndrome (SRNS) have been reported for up to one-third of children depending on age of the disease onset. Immunosuppressive treatment of genetic SRNS with calcineurin inhibitors (CNIs) is still controversial. The aim of the study was to investigate the efficacy of CNIs with focus on inducing remission and long-term kidney function in children with monogenic SRNS. Method Retrospective analysis of efficacy CNIs in five children (2M/3F) with monogenic SRNS was performed. Kidney biopsy prior CNIs revealed FSGS (n=4) and MCD (n=1). The initial cyclosporine (CsA) dose was 5 mg/kg/24h to keep a target level of 80-150 ng/ml and tacrolimus (TAC) - 0.1 mg/kg/24h to achieve a target level of 5-10 ng/ml. CsA took all 5 patients with subsequent switching to TAC in 2 children due to cosmetic side effects. The median follow-up period was 165.0 (IQR: 59.0; 185.5) months. Next generation sequencing (NGS) was used for identification of pathogenic variants in all patients. Results The median age at onset of monogenic SRNS was 33.0 (IQR: 16.5; 63.0) months. 2/5 (40%) patients presented with acute SRNS, 1/5 (20%) child with infantile NS, 1/5 (20%) - with isolated nephrotic range proteinuria with hypoalbuminemia and 1/5 (20%) - with NS and extrarenal features of Nail-Patella syndrome. NGS identified previously described pathogenic variants in all 5 children, including NPHS2 homozygous c.28dup (p.Glu87Ter) (n=1), NPHS2 compound heterozygous c.868G&amp;gt;A (p.Val290Met) in combination with c.686G&amp;gt;A (p.Arg229Gln) (n=1), LMX1B heterozygous c.788T&amp;gt;G (p.Val263Gly) (n=1), LMX1B heterozygous c.737G&amp;gt;A (p.Arg246Gln) (n=1), and COL4A3 heterozygous c.2962G&amp;gt;A (p.Gly988Arg) variant (n=1). The median time from diagnosis to initiation of CNIs treatment was 72.0 (IQR: 33.0; 93.0) months. CNIs induced complete remission in 2/5 (40%) patients, presented with acute SRNS, including one girl with MCD due to NPHS2 compound heterozygous variants with mutation-dependent pathogenicity of one (p.R229Q) of them and one boy with FSGS due to COL4A3 heterozygous variant (n=1). Partial remission was induced by CNIs in 2/5 (40%) girls with FSGS due to LMX1B heterozygous variants with isolated SRNS (n=1) and Nail-Patella syndrome (n=1). The median duration of CNIs treatment to obtain complete or partial remission was 13.5 (IQR: 6.8; 15.8) months. Acute CNIs-associated nephrotoxicity had 2 patients with LMX1B variants. At the last follow up full and partial responders to CNIs treatment aged of 16.5 (IQR: 11.8; 17.5) years had CKD-1 (n=3) and CKD-2 (n=1). 1/5 (20%) boy with NPHS2-associated infantile NS was CNI resistant and developed CKD-5 at the age of 6.5 years with subsequent living related kidney transplantation. Conclusion We found that 4/5 (80%) children with monogenic SRNS demonstrated partial or full response to CNIs treatment with stable long-term kidney function. We assume that CNIs might improve podocyte function by stabilization of their cytoskeleton disrupted in patients with monogenic SRNS.

Crumbs2 Is an Essential Slit Diaphragm Protein of the Renal Filtration Barrier.

Crumbs2 is expressed at embryonic stages as well as in the retina, brain, and glomerular podocytes. Recent studies identified CRB2 mutations as a novel cause of steroid-resistant nephrotic syndrome (SRNS). To study the function of Crb2 at the renal filtration barrier, mice lacking Crb2 exclusively in podocytes were generated. Gene expression and histologic studies as well as transmission and scanning electron microscopy were used to analyze these Crb2podKO knockout mice and their littermate controls. Furthermore, high-resolution expansion microscopy was used to investigate Crb2 distribution in murine glomeruli. For pull-down experiments, live cell imaging, and transcriptome analyses, cell lines were applied that inducibly express fluorescent protein-tagged CRB2 wild type and mutants. Crb2podKO mice developed proteinuria directly after birth that preceded a prominent development of disordered and effaced foot processes, upregulation of renal injury and inflammatory markers, and glomerulosclerosis. Pull-down assays revealed an interaction of CRB2 with Nephrin, mediated by their extracellular domains. Expansion microscopy showed that in mice glomeruli, Crb2 and Nephrin are organized in adjacent clusters. SRNS-associated CRB2 protein variants and a mutant that lacks a putative conserved O-glycosylation site were not transported to the cell surface. Instead, mutants accumulated in the ER, showed altered glycosylation pattern, and triggered an ER stress response. Crb2 is an essential component of the podocyte's slit diaphragm, interacting with Nephrin. Loss of slit diaphragm targeting and increasing ER stress are pivotal factors for onset and progression of CRB2-related SRNS.

New kid on the block: NOS1AP is a newly recognized genetic cause of steroid-resistant nephrotic syndrome in infants

Steroid-resistant nephrotic syndrome (SRNS) is a rare but important cause of chronic kidney disease and kidney failure in children. Mutations in podocyte genes explain more than two-thirds of early-onset SRNS presenting in the first year of life.1 In a recent publication in Science Advances, Majmundar et al. identified recessive mutations in nitric oxide synthase 1 adaptor protein (NOS1AP) as another genetic cause of early-onset SRNS.2

CLINICAL COURSE OF SCHIMKE IMMUNO-OSSEOUS DYSPLASIA AFTER KIDNEY TRANSPLANTATION

Schimke immuno-osseous dysplasia (SIOD) is one of the rare causes of steroid-resistant nephrotic syndrome. Extra-renal symptoms of this autosomal recessive disease include spondyloepiphyseal dysplasia, disproportional growth failure and defective cellular immunity. Kidney transplantation (KT) is indicated in these patients when their kidney problems progress to end-stage renal disease (ESRD). Due to the rarity of this condition, clinical course of SIOD after KT is not well known yet. Here we present our experience of SIOD after KT. Since 2014 three children with focal segmental glomerulosclerosis (FSGS) confirmed by biopsy were diagnosed as SIOD through genetic analysis. Onset age of proteinuria was 7, 5, 3 years, respectively. All of them had disproportional growth failure and typical facial feature. They reached ESRD in about two years after diagnosis of proteinuria. At their age of 9, 7, 9 years, these patients had living donor KT from their parents. After KT they suffered from multiple infections. Case 1 had PCP pneumonia requiring ventilator care 2 months after KT, then in 3 months disseminated adenovirus infection occurred along with CMV, EBV, and JC virus infection. The patient succumbed to the infection in 2 months despite aggressive management. Case 2 had CMV infection and PCP pneumonia after 3 months of KT. She suffered from skin infection and otitis. Case 3 suffered from respiratory failure due to CMV infection after 5 months of KT. Recurrent hospitalization was required due to various infections. Allograft has survived for 3.5 years in case 3 despite low-dose tacrolimus single medication (< 5ng/mL), but case 1 lost his graft function along with disseminated adenovirus infection. Case 2 had graft failure after 5 years of KT from acute rejection, probably due to insufficient dosage of tacrolimus and poor compliance. As Extra-renal presentation case 1 had renal vein thrombosis extended to IVC and pulmonary hypertension before KT, lowering his oxygen saturation to 80-90% in room air. Case 2 had severe developmental dysplasia of the hip requiring orthopedic surgery at the age of 5. She had sitting height and leg length ratio of 0.96, vertebral flattening and lordosis on spine x-ray suggesting spondyloepiphyseal dysplasia. After KT she suffered constant headache and Moyamoya syndrome was dignosed. She had an encephaloduroarteriosynangiosis when she was 10yrs 4month old. Case 3 suffered from spontaneous bacterial peritonitis and recurrent rhabdomyolysis along with neutropenia and thrombocytopenia before KT. In three years of KT his neutropenia and thrombocytopenia aggravated and bone marrow study suggested aplastic anemia. Hematopoietic stem cell transplantation is under consideration. In summary, SIOD is a rare systemic disease presenting as FSGS. While KT is indicated for their ESRD, their extra-renal symptoms progress regardless of KT. With its defective cellular immunity multiple infections complicate the clinical course after KT in SIOD.

Genetic Study in Korean Pediatric Patients with Steroid-Resistant Nephrotic Syndrome or Focal Segmental Glomerulosclerosis

Steroid-resistant nephrotic syndrome (SRNS) is one of the major causes of end-stage renal disease (ESRD) in childhood and is mostly associated with focal segmental glomerulosclerosis (FSGS). More than 50 monogenic causes of SRNS or FSGS have been identified. Recently, the mutation detection rate in pediatric patients with SRNS has been reported to be approximately 30%. In this study, genotype-phenotype correlations in a cohort of 291 Korean pediatric patients with SRNS/FSGS were analyzed. The overall mutation detection rate was 43.6% (127 of 291 patients). WT1 was the most common causative gene (23.6%), followed by COQ6 (8.7%), NPHS1 (8.7%), NUP107 (7.1%), and COQ8B (6.3%). Mutations in COQ6, NUP107, and COQ8B were more frequently detected, and mutations in NPHS2 were less commonly detected in this cohort than in study cohorts from Western countries. The mutation detection rate was higher in patients with congenital onset, those who presented with proteinuria or chronic kidney disease/ESRD, and those who did not receive steroid treatment. Genetic diagnosis in patients with SRNS provides not only definitive diagnosis but also valuable information for decisions on treatment policy and prediction of prognosis. Therefore, further genotype-phenotype correlation studies are required.

Transcription factor MafB in podocytes protects against the development of focal segmental glomerulosclerosis

Focal segmental glomerulosclerosis (FSGS) is a common cause of steroid-resistant nephrotic syndrome. Spontaneous remission of FSGS is rare and steroid-resistant FSGS frequently progresses to renal failure. Many inheritable forms of FSGS have been described, caused by mutations in proteins that are important for podocyte function. Here, we show that a basic leucine zipper transcription factor, MafB, protects against FSGS. MAFB expression was found to be decreased in the podocytes of patients with FSGS. Moreover, conditional podocyte-specific MafB-knockout mice developed FSGS with massive proteinuria accompanied by depletion of the slit diaphragm-related proteins (Nphs1 and Magi2), and the podocyte-specific transcription factor Tcf21. These findings indicate that MafB plays a crucial role in the pathogenesis of FSGS. Consistent with this, adriamycin-induced FSGS and attendant proteinuria were ameliorated by MafB overexpression in the podocytes of MafB podocyte-specific transgenic mice. Thus, MafB could be a new therapeutic target for FSGS.

Genetic tests in children with steroid-resistant nephrotic syndrome

Steroid-resistant nephrotic syndrome (SRNS) is a common cause of chronic kidney disease in children, and a considerable number of patients progress to end-stage renal disease. SRNS is a highly heterogeneous disorder, both clinically and genetically, and more than 50 monogenic causes of SRNS, including isolated and syndromic forms, have been identified. Recent large-cohort studies indicate that at least 30% of childhood-onset SRNS cases are genetic. The benefits of definitive molecular diagnosis by genetic testing include the avoidance of unnecessary and potentially harmful diagnostic procedures (e.g., kidney biopsy) and treatment (e.g., steroid and immunosuppressants), detection of rare and potentially treatable mutations (e.g., coenzyme Q10 biosynthesis pathway defect), prediction of prognosis (e.g., posttransplant recurrence), and providing precise genetic counseling. Furthermore, the identification of novel disease-causing genes could provide new insights into the pathogenic mechanisms of SRNS. Therefore, whenever accessible and affordable, genetic testing is recommended for all pediatric patients with SRNS, and should certainly be performed in patients with a higher probability of genetic predisposition based on genotype-phenotype correlation data. The genetic testing approach should be determined for each patient, and clinicians should, therefore, be aware of the advantages and disadvantages of methods currently available, which include Sanger sequencing, gene panel testing, and whole-exome or whole-genome sequencing. Importantly, the need for precise and thorough phenotyping by clinicians, even in the era of genomics, cannot be overemphasized. This review provides an update on recent advances in genetic studies, a suggested approach for the genetic testing of pediatric patients with SRNS.

Centromere protein I (CENPI) is a candidate gene for X-linked steroid sensitive nephrotic syndrome.

Individuals with proteinuria in association with hypoalbuminemia, edema, and hyperlipidemia are considered as having nephrotic syndrome (NS). NS is the most common kidney disease seen in the paediatric age group. NS is usually classified into steroid resistant nephrotic syndrome (SRNS) and steroid sensitive nephrotic syndrome (SSNS). More than 58 genes have been identified as a monogenic cause of SRNS, however, the genetic architecture of childhood SSNS remains poorly understood. Here in this study, we performed sequencing of 66 NS candidate genes followed by whole genome SNP genotyping and whole exome sequencing in SSNS families with multiple affected individuals. NS candidate genes sequencing did not identify any pathogenic variant in the known genes. Homozygosity mapping based on an autosomal recessive model failed to detect any shared loss of heterozygosity region in the genome. An unbiased and hypothesis-free exome data analysis identified a missense variant (c.383G>A; p.Arg128Gln) in the CENPI gene. Sanger sequencing of both parents, unaffected and affected individuals confirmed an X-linked inheritance pattern of the variant (c.383G>A) with SSNS phenotype. The variant (c.383G>A) is very rare and is potentially damaging. Collectively, these observations suggest that a specific pathogenic link between SSNS development and alteration in CENPI exists. However, human mutations in CENPI causing SSNS have not been reported hitherto. Identification of genetic defects underlying SSNS will help in understanding the precise aetiology of SSNS and improved management of children with NS.

Clinicopathologic Features and the Association with Short-Term Outcome of Primary Membranous Nephropathy in Children: A Single-Center Study from Pakistan.

Membranous nephropathy (MN) is an uncommon cause of steroid-resistant nephrotic syndrome in children. Our study aimed to determine the clinicopathologic features of primary MN in children and their association with short-term outcome. This observational study was conducted from January 2009 to June 2017 at the Pediatric Nephrology Department. A total of 50 children were diagnosed with primary MN. Their clinical, laboratory, and histopathological findings on renal biopsy were recorded. The minimum follow-up was for six months. Clinicopathologic features were correlated with the outcome at the last follow-up. Data analysis was done using IBM SPSS Statistics for Windows software version 20.0. The mean age at onset was 10.92 ± 3.08 years (range: 4-17 years). The male-to-female ratio was 3:1. The serum albumin of ≤2.5 g/dL was seen in 40 patients (80%), hypertension was present in 38 (76%), and heavy proteinuria was seen in 32 children (70%). The mean estimated glomerular filtration rate (eGFR) at presentation was 178.71 ± 0.78 mL/min/1.73 m2. At the initial visit, nine children (18.4%) were in chronic kidney disease stage 2 and one (2%) in stage 4. Phospholipase A2 receptor antibody was present in five (15%) of 32 children tested. At the last follow-up (28 interquartile range: 25.5 months), 11 children (26%) were in complete remission and 25 (66%) had achieved partial remission. The mean eGFR had reduced to 145.84 ± 78.05 mL/min/1.73 m2. Patients with normal initial eGFR were more likely to go into remission (P = 0.001). The short-term outcome of childhood primary MN is relatively good in our setup. A multicenter collaborative study is required to determine prognostic factors and to standardize treatment in this uncommon nephropathy.

A rare cause of steroid-resistant nephrotic syndrome in a child: Answers.

A rare cause of steroid-resistant nephrotic syndrome in a child: Answers.

TBC1D8B Mutations Implicate RAB11-Dependent Vesicular Trafficking in the Pathogenesis of Nephrotic Syndrome

Mutations in about 50 genes have been identified as monogenic causes of nephrotic syndrome, a frequent cause of CKD. These genes delineated the pathogenetic pathways and rendered significant insight into podocyte biology. We used whole-exome sequencing to identify novel monogenic causes of steroid-resistant nephrotic syndrome (SRNS). We analyzed the functional significance of an SRNS-associated gene in vitro and in podocyte-like Drosophila nephrocytes. We identified hemizygous missense mutations in the gene TBC1D8B in five families with nephrotic syndrome. Coimmunoprecipitation assays indicated interactions between TBC1D8B and active forms of RAB11. Silencing TBC1D8B in HEK293T cells increased basal autophagy and exocytosis, two cellular functions that are independently regulated by RAB11. This suggests that TBC1D8B plays a regulatory role by inhibiting endogenous RAB11. Coimmunoprecipitation assays showed TBC1D8B also interacts with the slit diaphragm protein nephrin, and colocalizes with it in immortalized cell lines. Overexpressed murine Tbc1d8b with patient-derived mutations had lower affinity for endogenous RAB11 and nephrin compared with wild-type Tbc1d8b protein. Knockdown of Tbc1d8b in Drosophila impaired function of the podocyte-like nephrocytes, and caused mistrafficking of Sns, the Drosophila ortholog of nephrin. Expression of Rab11 RNAi in nephrocytes entailed defective delivery of slit diaphragm protein to the membrane, whereas RAB11 overexpression revealed a partial phenotypic overlap to Tbc1d8b loss of function. Novel mutations in TBC1D8B are monogenic causes of SRNS. This gene inhibits RAB11. Our findings suggest that RAB11-dependent vesicular nephrin trafficking plays a role in the pathogenesis of nephrotic syndrome.