Differential gene expression in NPHS2-Cre +/+ mouse glomeruli versus wild-type control
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE117331 | GEO | 2019/02/11
REPOSITORIES: GEO
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Project description:Podocytes are the highly specialised cells within the glomeruli of the kidney that maintain the filtration barrier by forming interdigitating foot processes and slit-diaphragms. Disruption to these features result in proteinuria. Studies into podocyte biology and disease has been hampered by a paucity of in vitro models of this non-proliferative cell type. Here we characterise sieved glomeruli from kidney organoids derived from human pluripotent stem cells. Compared to conditionally immortalised podocytes, organoid-derived glomeruli show superior podocyte-specific gene and protein expression, morphology and functional properties. Using CRISPR-derived MAFB reporter iPSC lines, homozygous MAFB mutant organoids recapitulated the anticipated disease related transcriptional changes. Culture of kidney organoids on chicken chorioallantoic membrane resulted in glomerular vascularisation, glomerular filtration barrier assembly, formation of slit diaphragms and fenestrated endothelial cells. This definitively demonstrates that human iPSC kidney organoid-derived glomeruli can serve as an accurate model of human podocytopathies and glomerular disease in vitro.
2018-12-04 | GSE99583 | GEO
Project description:Podocytes are highly specialised cells within the glomeruli of the kidney that maintain the filtration barrier by forming interdigitating foot processes and slit-diaphragms. Disruption to these features result in proteinuria and glomerulosclerosis. Studies into podocyte biology and disease have previously relied on conditionally immortalised cell lines due to the non- proliferative nature of this cell type. Here we describe an advanced model to study both podocyte and glomerular biology using isolated glomeruli from kidney organoids derived from human pluripotent stem cells.
2018-12-04 | GSE111992 | GEO
Project description:Differential gene expression in NPHS2-Cre +/+ mouse glomeruli versus wild-type control
| PRJNA481767 | ENA
Project description:We and others have previously shown that glomerular endothelial cells and podocytes express hypoxia-inducible transcription factors (HIFs). HIFs bind to hypoxia response elements in target genes, such as vascular endothelial growth factor, which is continually produced by podocytes throughout life. To further assess function of HIFs in podocyte biology, podocin-Cre mice were mated with floxed von Hippel-Lindau (VHL) mice to selectively delete VHL, a component of an E3 ligase complex responsible for degradation of HIFs in normoxia. We reasoned that cells lacking VHL would overexpress stable HIFs and upregulate hypoxia-responsive genes. Progeny from these crosses displayed two phenotypes, non-proteinuric with glomerular basement membrane (GBM) defects and proteinuric with GBM defects and end-stage renal failure at ~6 months of age. Gene changes associated with the mild, non-proteinuric phenotype were studied using isolated glomeruli from wildtype and Pod-Cre fVHL mice. At 4 weeks of age, urine was collected and urinary albumin was quantified by Albuwell elisa from Pod-Cre fVHL litters. At 6 weeks of age, glomeruli from 3 wildtype littermate controls and 3 non-proteinuric Pod-Cre fVHL mice were collected using the magnetic bead method. RNA was extracted and hybridized to Affymetrix microarrays.
2010-08-03 | E-GEOD-20235 | biostudies-arrayexpress
Project description:Podocyte specific knockout mice for Parva (Parva-fl/fl*hNPHS2Cre) were generated. Transcriptome profiling (RNA-Seq) and differential gene expression analysis of isolated renal glomeruli from KO and WT mice was performed. Parva KO mice showed transcriptional changes associated with podocyte and glomerular disease.
2022-01-20 | GSE181690 | GEO
Project description:The Wilms tumor-suppressor gene WT1, a key player in renal development, also has a crucial role in maintenance of the glomerulus in the mature kidney. However, molecular pathways orchestrated by WT1 in podocytes, where it is highly expressed, remain unknown. Their defects are thought to modify the cross-talk between podocytes and other glomerular cells and ultimately lead to glomerular sclerosis, as observed in diffuse mesangial sclerosis (DMS) a nephropathy associated with WT1 mutations. To identify podocyte WT1 targets, we generated a novel DMS mouse line, performed gene expression profiling in isolated glomeruli, and identified excellent candidates that may modify podocyte differentiation and growth factor signalling in glomeruli. Scel, encoding sciellin, a protein of the cornified envelope in the skin, and sulf1, encoding a 6-O endosulfatase, are shown to be expressed in wild type podocytes and to be strongly down-regulated in mutants. Co-expression of Wt1, Scel and Sulf1 was also found in a mesonephric cell line, and siRNA-mediated knockdown of WT1 decreased Scel and Sulf1 mRNAs and proteins. By ChIP we show that Scel and Sulf1 are direct WT1 targets. Cyp26a1, encoding an enzyme involved in the degradation of retinoic acid, is shown to be up-regulated in mutant podocytes. Cyp26a1 may play a role in the development of glomerular lesions but does not seem to be regulated by WT1. These results provide novel clues in our understanding of normal glomerular function and early events involved in glomerulosclerosis. Experiment Overall Design: Isolation of glomeruli from mutant (FVB-N4 Wt1+/R394W) and wild-type (FVB-N4 Wt1+/+) was performed after cardiac Dynabead perfusion. GeneChip analysis of glomeruli from 5 Wt1+/R394W mice and 5 Wt1+/+ littermates (N4-FVB) were performed independently. Animals were unweaned 27-day-old males. The Wt1+/R394W mice used were showing little albuminuria (<3 ug/ul on Coomassie blue stained SDS-PAGE gel) and no evidence of mesangial lesions by light microscopy.
2009-10-09 | E-GEOD-18358 | biostudies-arrayexpress
Project description:The specific contribution of the two TNF-receptors Tnfr1 and Tnfr2 to TNF-induced inflammation in the glomerulus is unknown. In mice, TNF exposure induces glomerular expression of inflammatory mediators like adhesion molecules and chemokines in vivo, and glomerular accumulation of leukocytes. To examine Tnfr-specific inflammatory responses in intrinsic glomerular cells but not infiltrating leukocytes we performed microarray gene expression profiling on intact glomeruli isolated from wild-type and Tnfr-deficient mice following TNF exposure in vitro. Glomeruli were isolated from male C57BL/6J wild-type and Tnfr-deficient mice applying a magnetic bead-based isolation technique. Isolated intact glomeruli were stimulated with TNF for 12 hours in vitro for subsequent RNA extraction and hybridization on Affymetrix microarrays.
2013-09-11 | E-GEOD-43928 | biostudies-arrayexpress
Project description:Mouse glomeruli and brain capillary fragments were prepared. Podocytes were separated from isolated glomeruli from 8-day-old Podocin-Cre, Z/EG double transgenic mice as follows: Isolated glomeruli were incubated with trypsin solution containing 0.2 % trypsin-EDTA (Sigma-Aldrich), 100ug/ml Heparin and 100U/ml DNase I in PBS for 25 min at 37 degree, with mixing by pipetting every 5 min. The trypsin was inactivated with soybeans trypsin inhibitor (Sigma-Aldrich) and the cell suspension sieved through a 30 um pore size filter (BD bioscience, Franklin Lakes, NJ). Cells were collected by centrifugation at 200 x g for 5 min at 4 degree and resuspended in 1ml PBS supplemented with 0.1 % BSA. To separate GFP-expressing (GFP+) and GFP-negative (GFP-) cells, glomerular cell were sorted using a FACSVantage SE (BD, San Jose, CA, USA) operating at a sheath pressure of 22 psi. Autofluorescent cells were excluded by analyzing the emission of orange light (585 nm). To allow for standardization of results, the samples such as glomerular RNA, brain capillary RNA, rest of kidnay RNA, podocyte RNA and foxc2 glomreular RNA were hybridized in competition with common reference samples.
2006-10-31 | E-MEXP-492 | biostudies-arrayexpress
Project description:We and others have previously shown that glomerular endothelial cells and podocytes express hypoxia-inducible transcription factors (HIFs). HIFs bind to hypoxia response elements in target genes, such as vascular endothelial growth factor, which is continually produced by podocytes throughout life. To further assess function of HIFs in podocyte biology, podocin-Cre mice were mated with floxed von Hippel-Lindau (VHL) mice to selectively delete VHL, a component of an E3 ligase complex responsible for degradation of HIFs in normoxia. We reasoned that cells lacking VHL would overexpress stable HIFs and upregulate hypoxia-responsive genes. Progeny from these crosses displayed two phenotypes, non-proteinuric with glomerular basement membrane (GBM) defects and proteinuric with GBM defects and end-stage renal failure at ~6 months of age. Gene changes associated with the mild, non-proteinuric phenotype were studied using isolated glomeruli from wildtype and Pod-Cre fVHL mice.
2010-08-03 | GSE20235 | GEO
Project description:To investigate the impact of various NPHS2 homozygous point mutations on podocyte biology in induced pluripotent stem cell (iPSC)-derived human kidney organoids, an iPSC allelic series was generated from a control fibroblast line and two iPSC lines were derived from patient blood samples (one homozygous R168H and one unaffected relative) iPSCs were differentiated into kidney organoids, organoids glomeruli were harvested and we performed gene expression profiling using data obtained from RNA-seq of 3 sets of 4 organoids for each genotype at D7+14 of our differentiation protocol (Takasato et al, Nature 2016)
2022-09-07 | GSE205647 | GEO