α-Actinin-4-Mediated FSGS: An Inherited Kidney Disease Caused by an Aggregated and Rapidly Degraded Cytoskeletal Protein

Focal segmental glomerulosclerosis (FSGS) is a common pattern of renal injury, seen as both a primary disorder and as a consequence of underlying insults such as diabetes, HIV infection, and hypertension. Point mutations in theα-actinin-4 gene ACTN4 cause an autosomal dominant form of human FSGS. We characterized the biological effect of these mutations by biochemical assays, cell-based studies, and the development of a new mouse model. We found that a fraction of the mutant protein forms large aggregates with a high sedimentation coefficient. Localization of mutant α-actinin-4 in transfected and injected cells, as well as in situ glomeruli, showed aggregates of the mutant protein. Video microscopy showed the mutant α-actinin-4 to be markedly less dynamic than the wild-type protein. We developed a “knockin” mouse model by replacing Actn4 with a copy of the gene bearing an FSGS-associated point mutation. We used cells from these mice to show increased degradation of mutant α-actinin-4, mediated, at least in part, by the ubiquitin–proteasome pathway. We correlate these findings with studies of α-actinin-4 expression in human samples. “Knockin” mice with a disease-associated Actn4 mutation develop a phenotype similar to that observed in humans. Comparison of the phenotype in wild-type, heterozygous, and homozygous Actn4 “knockin” and “knockout” mice, together with our in vitro data, suggests that the phenotypes in mice and humans involve both gain-of-function and loss-of-function mechanisms

Novel NPHS2 variant in patients with familial steroid-resistant nephrotic syndrome with early onset, slow progression and dominant inheritance pattern

Background Steroid-resistant nephrotic syndrome (SRNS) is a common cause of end-stage renal disease in children but also occurs as an adult-onset condition. In a subset of SRNS patients, pathogenic variants are found in genes coding for podocyte foot process proteins. The aim of this study was to define the role of pathogenic variants in Finnish patients with familial and sporadic SRNS. Methods We analyzed SRNS-related genes NPHS1, NPHS2, NEPH1, ACTN4, TRPC6, INF2, WT1, CD2AP, LAMB2, and PLCE1 for disease-causing variants using direct sequencing of exons and intron/exon boundaries in all members of a family with dominant SRNS with early onset and slow progression to end-stage renal disease. We carried out a whole genome sequencing in two affected and two healthy family members. The function of found podocin variant was studied using co-immunoprecipitation and immunohistochemistry. Podocyte gene sequences were analyzed in a cohort of Finnish non-familial SRNS patients. Results A heterozygous de novo deletion, c. 988_989delCT in NPHS2, was found in all affected family members and in none of their healthy relatives, non-familial patients or controls. No other SRNS-related gene variant, coding or non-coding co-segregated with the disease phenotype in the family. While the truncated podocin remained able to bind nephrin, the expression of nephrin was fragmented and podocin expression reduced. The gene analysis of the non-familial SRNS patients revealed few variants. Conclusion The role of podocin variants in nephrotic syndrome may be more varied than previously thought.Peer reviewe

Podocyte-specific expression of cre recombinase in transgenic mice

Summary: We report a transgenic mouse line that expresses Cre recombinase exclusively in podocytes. Twenty- four transgenic founders were generated in which Cre recombinase was placed under the regulation of a 2.5-kb fragment of the human NPHS2 promoter. Previously, this fragment was shown to drive beta-galactosidase (Β-gal) expression exclusively in podocytes of transgenic mice. For analysis, founder mice were bred with ROSA26 mice, a reporter line that expresses Β-gal in cells that undergo Cre recombination. Eight of 24 founder lines were found to express Β-gal exclusively in the kidney. Histological analysis of the kidneys showed that Β-gal expression was confined to podocytes. Cre recombination occurred during the capillary loop stage in glomerular development. No evidence for Cre recombination was detected in any of 14 other tissues examined. genesis 35:39–42, 2003. © 2002 Wiley-Liss, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/35267/1/10164_ftp.pd

Pathophysiology of focal segmental glomerulosclerosis

Focal segmental glomerulosclerosis (FSGS) is a major cause of idiopathic steroid-resistant nephrotic syndrome (SRNS) and end-stage kidney disease (ESKD). In recent years, animal models and studies of familial forms of nephrotic syndrome helped elucidate some mechanisms of podocyte injury and disease progression in FSGS. This article reviews some of the experimental and clinical data on the pathophysiology of FSGS

Propagation phase-contrast micro-computed tomography allows laboratory-based three-dimensional imaging of articular cartilage down to the cellular level

High-resolution non-invasive three-dimensional (3D) imaging of chondrocytes in articular cartilage remains elusive. The aim of this study was to explore whether laboratory micro-computed tomography (micro-CT) permits imaging cells within articular cartilage

No evidence for genotype/phenotype correlation in NPHS1 and NPHS2 mutations

Primary steroid-resistant nephrotic syndrome (SRNS) is characterized by childhood onset of proteinuria and progression to end-stage renal disease. In 26% of cases it is caused by recessive mutations in NPHS2 (podocin). Congenital nephrotic syndrome (CNS) is caused by mutations in NPHS1 (nephrin) or NPHS2 . In three families mutations in NPHS1 and NPHS2 had been reported to occur together, and these tri-allelic mutations were implicated in genotype/phenotype correlations. To further test the hypothesis of tri-allelism, we examined a group of 62 unrelated patients for NPHS1 mutations, who were previously shown to have NPHS2 mutations; 15 of 62 patients had CNS. In addition, 12 CNS patients without NPHS2 mutation were examined for NPHS1 mutations. Mutational analysis yielded three different groups. (1) In 48 patients with two recessive NPHS2 mutations (11 with CNS), no NPHS1 mutation was detected, except for 1 patient, who had one NPHS1 mutation only. This patient was indistinguishable clinically and did not have CNS. (2) In 14 patients with one NPHS2 mutation only (4 with CNS), we detected two additional recessive NPHS1 mutations in the 4 patients with CNS. They all carried the R229Q variant of NPHS2 . The CNS phenotype may be sufficiently explained by the presence of two NPHS1 mutations. (3) In 12 patients without NPHS2 mutation (all with CNS), we detected two recessive NPHS1 mutations in 11 patients, explaining their CNS phenotype. We report ten novel mutations in the nephrin gene. Our data do not suggest any genotype/phenotype correlation in the 5 patients with mutations in both the NPHS1 and the NPHS2 genes.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/47823/1/467_2004_Article_1629.pd

African ancestry allelic variation at the MYH9 gene contributes to increased susceptibility to non-diabetic end-stage kidney disease in Hispanic Americans

Recent studies identified MYH9 as a major susceptibility gene for common forms of non-diabetic end-stage kidney disease (ESKD). A set of African ancestry DNA sequence variants comprising the E-1 haplotype, was significantly associated with ESKD. In order to determine whether African ancestry variants are also associated with disease susceptibility in admixed populations with differing genomic backgrounds, we genotyped a total of 1425 African and Hispanic American subjects comprising dialysis patients with diabetic and non-diabetic ESKD and controls, using 42 single nucleotide polymorphisms (SNPs) within the MYH9 gene and 40 genome-wide and 38 chromosome 22 ancestry informative markers. Following ancestry correction, logistic regression demonstrated that three of the E-1 SNPs are also associated with non-diabetic ESKD in the new sample sets of both African and Hispanic Americans, with a stronger association in Hispanic Americans. We also identified MYH9 SNPs that are even more powerfully associated with the disease phenotype than the E-1 SNPs. These newly associated SNPs, could be divided into those comprising a haplotype termed S-1 whose association was significant under a recessive or additive inheritance mode (rs5750248, OR 4.21, P < 0.01, Hispanic Americans, recessive), and those comprising a haplotype termed F-1 whose association was significant under a dominant or additive inheritance mode (rs11912763, OR 4.59, P < 0.01, Hispanic Americans, dominant). These findings strengthen the contention that a sequence variant of MYH9, common in populations with varying degrees of African ancestry admixture, and in strong linkage disequilibrium with the associated SNPs and haplotypes reported herein, strongly predisposes to non-diabetic ESKD

Expression of TRPC6 in Renal Cortex and Hippocampus of Mouse during Postnatal Development

TRPC6, a member of the TRPC family, attracts much attention from the public because of its relationship with the disease. In both the brain and kidney, TRPC6 serves a variety of functions. The aim of the present study was to observe the expression and effects of TRPC6 in renal cortex and hippocampus during early postnatal development of the mouse. In the present study, immunohistochemistry and Western blotting were used to detect the expression of TRPC6 in the mouse kidney and hippocampus of postnatal day 1, 3, 5, 7, 14, 21, 28 and 49 (P1, P3, P5, P7, P14, P21, P28 and P49). Results showed that the expression of TRPC6 was increased in the mouse hippocampus, and there was a significant increase between P7 and P14 during the postnatal development. Meanwhile, the expression of TRPC6 was also detected in glomerulus and tubules, and a decreased expression was found during postnatal maturation of mouse renal cortex. From these in vivo experiments, we concluded that the expression of TRPC6 was active in the developing mouse kidney cortex, and followed a loss of expression with the development of kidney. Meanwhile, an increased expression was found in the hippocampus with the development. Together, these data suggested that the developmental changes in TRPC6 expression might be required for proper postnatal kidney cortex development, and played a critical role in the hippocampus during development, which formed the basis for understanding the nephrogenesis and neurogenesis in mice and provided a practically useful knowledge to the clinical and related research

Genetics of focal segmental glomerulosclerosis

The recent advances in understanding the pathophysiology of focal segmental glomerulosclerosis (FSGS) and molecular function of glomerular filtration barrier come directly from genetic linkage and positional cloning studies. The exact role and function of the newly discovered genes and proteins are being investigated by in vitro and in vivo mechanistic studies. Those genes and proteins interactions seem to change susceptibility to kidney disease progression. Better understanding of their exact role in the development of FSGS may influence future therapies and outcomes in this complex disease

Glucose Monitoring During Pregnancy

Self-monitoring of blood glucose in women with mild gestational diabetes has recently been proven to be useful in reducing the rates of fetal overgrowth and gestational weight gain. However, uncertainty remains with respect to the optimal frequency and timing of self-monitoring. A continuous glucose monitoring system may have utility in pregnant women with insulin-treated diabetes, especially for those women with blood sugars that are difficult to control or who experience nocturnal hypoglycemia; however, continuous glucose monitoring systems need additional study as part of larger, randomized trials