Formation of the Glomerular Microvasculature is Regulated by VEGFR-3
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE198227 | GEO | 2022/12/01
REPOSITORIES: GEO
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Project description:Background: Recent single-cell RNA sequencing (scRNA-seq) analyses have offered much insight into cell-specific gene expression profiles in normal kidneys. However, in diseased kidneys, understanding of changes in specific cells, particularly glomerular cells, remains limited. Methods: To elucidate the glomerular cell–specific gene expression changes in diabetic kidney disease, we performed scRNA-seq analysis of isolated glomerular cells from streptozotocin-induced diabetic endothelial nitric oxide synthase (eNOS)–deficient (eNOS-/-) mice and control eNOS-/- mice. Results: We identified five distinct cell populations, including glomerular endothelial cells, mesangial cells, podocytes, immune cells, and tubular cells. Using scRNA-seq analysis, we confirmed the expression of glomerular cell–specific markers and also identified several new potential markers of glomerular cells. The number of immune cells was significantly higher in diabetic glomeruli compared with control glomeruli, and further cluster analysis showed that these immune cells were predominantly macrophages. Analysis of differential gene expression in endothelial and mesangial cells of diabetic and control mice showed dynamic changes in the pattern of expressed genes, many of which are known to be involved in diabetic kidney disease. Moreover, gene expression analysis showed variable responses of individual cells to diabetic injury. Conclusion: Our findings demonstrate the ability of scRNA-seq analysis in isolated glomerular cells from diabetic and control mice to reveal dynamic changes in gene expression in diabetic kidneys, with variable responses of individual cells. Such changes, which might not be apparent in bulk transcriptomic analysis of glomerular cells, may help identify important pathophysiologic factors contributing to the progression of diabetic kidney disease.
2019-03-08 | GSE127235 | GEO
Project description:There are 3 cell types in a glomerulus: podocytes, mesangial cells and endothelial cells. These cell types play distinct roles in the structure and functions of glomeruli. In order to profile the gene expression of single glomerular cells, we isolated mouse glomeruli by Dynabead/magnetic concentration method and digested them with enzymes to dissociate them into single cells. We loaded the single cell suspension to a Fluidigm C1 Single-Cell Auto Prep System for single cell cDNA preparation. We performed qPCR analyses of marker genes of podocytes (Npsh2, Synaptopodin, WT1), mesangial cells (Gata3, IGFbp5) and endothelial cells (CD31, Tie2) to determine the identity of each cDNA sample. To identify podocyte-specific genes, we mixed 15 mesangial cell cDNA samples and 15 endothelial cell cDNA samples and further divided into 3 aliquots as replicates for sequencing using Illumina HiSeq 2000 system. The resulting data are used to compare with that of podocytes in order to identify podocyte-specific genes.
2017-07-31 | GSE89263 | GEO
Project description:Glomerular diseases are the leading cause of chronic kidney diseases with pathomechanisms largely unclear. It is known that ANGPT2 regulates endothelial cell homeostasis and function via TEK/TIE2 and its deregulation causes endothelial damage. We found that ANGPT2 is upregulated in glomerular diseases and wondered whether it has any effect on glomerular podocytes and mesangial cells given that they have no or low TEK expression. We treated podocytes and mesangial cells in culture with ANGPT2 and found no overt cellular changes. RNA-seq analysis showed that gene expression was altered in both podocytes and mesangial cells and that the regulated genes in the two cell types were fundamentally different. GO and KEGG analyses showed that the two groups of regulated genes were enriched in distinct processes and pathways. These results suggest that ANGPT2 exerts effects on both podocytes and mesangial cells and that increased ANGPT2 may be involved in glomerular injury by affecting podocytes and mesangial cells in addition to endothelial cells.
2022-01-10 | GSE185293 | GEO
Project description:Glomerular diseases are the leading cause of chronic kidney diseases with pathomechanisms largely unclear. It is known that ANGPT2 regulates endothelial cell homeostasis and function via TEK/TIE2 and its deregulation causes endothelial damage. We found that ANGPT2 is upregulated in glomerular diseases and wondered whether it has any effect on glomerular podocytes and mesangial cells given that they have no or low TEK expression. We treated podocytes and mesangial cells in culture with ANGPT2 and found no overt cellular changes. RNA-seq analysis showed that gene expression was altered in both podocytes and mesangial cells and that the regulated genes in the two cell types were fundamentally different. GO and KEGG analyses showed that the two groups of regulated genes were enriched in distinct processes and pathways. These results suggest that ANGPT2 exerts effects on both podocytes and mesangial cells and that increased ANGPT2 may be involved in glomerular injury by affecting podocytes and mesangial cells in addition to endothelial cells.
2022-01-10 | GSE185292 | GEO
Project description:Renal blood filtration occurs in a functional unit called the glomerulus. The resident cell types comprise the filtration barrier, namely podocytes, mesangial cells and glomerular endothelial cells. Here we introduce a glomerular cell isolation protocol, which enables the separation of these three cell types in sufficient amounts and purity to allow detailed protein-biochemical investigations. We demonstrate that the expression of fluorescent transgenes in glomerular cells can result in proteome artifacts. We show that different mouse strains have different glomerular cell type proteomes. Further, we demonstrate the power of the technique to identify new glomerular cell type-enriched proteins and demonstrate the potential of this globally applicable technique in the dissection of cell-specific disease responses and intra-glomerular cell-type crosstalk.
2021-11-12 | PXD024481 | Pride
Project description:Protoemics of endothelia, podocytes, mesangial cells from mouse
2022-06-09 | PXD016238 | Pride
Project description:Purpose: Next-generation sequencing (NGS) was used to define the transcriptome of native mouse podocytes and non-podocytes glomerular cells as part of a project aiming to define the molecular fingerprint of mouse podocytes. Method: Glomeruli from 29 Gt(ROSA)26Sortm4(ACTB-tdTomato,-EGFP)Luo/J x hNPHS2Cre mice at the age of 10 weeks were purified and a single cell solution was prepared to seperate GFP-expressing (podocytes) and GFP-negative (non-podocytes glomerular cells) cells by FACS sorting. RNA was extracted and prepared for further analysis using directional, polyA+ library preparation. An Illumina HiSeq2500 was used for a paired-end sequencing of 100 cycles . Salmon and Sleuth were used for downstream analysis. Results: A total of 100 Million reads each from podocytes and non-podocytes glomerular cells could be used for further analysis.
2019-01-01 | E-MTAB-5457 | biostudies-arrayexpress
Project description:Podocytes, highly differentiated glomerular epithelial cells, are essential for the maintenance of glomerular filtration barrier. Podocyte dysfunction in podocytes is a major determinant of proteinuric kidney disease. By RNA sequencing analysis in ADR-treated podocytes with or without MYDGF overexpression, we observed the significant changes of genes important in regulating cell cycle in podocytes with ADR treatment.
2021-10-06 | GSE185226 | GEO
Project description:Heparan sulfate (HS) is an important component of the kidney anionic filtration barrier, the glomerular basement membrane (GBM). HS chains attached to proteoglycan protein cores are modified by sulfotransferases in a highly ordered series of biosynthetic steps resulting in immense structural diversity due to negatively charged sulfate modifications. 3-O-sulfation is the least abundant modification generated by a family of seven isoforms but creates the most highly sulfated HS domains. We analyzed the kidney phenotypes in the Hs3st3a1, Hs3st3b1 and Hs3st6 -knockout (KO) mice, the isoforms enriched in kidney podocytes. Individual KO mice show no overt kidney phenotype, although Hs3st3b1 kidneys were smaller than wildtype (WT). Furthermore, Hs3st3a1-/-; Hs3st3b1-/- double knockout (DKO) kidneys were smaller but also had a reduction in glomerular size relative to wildtype (WT). Mass spectrometry analysis of kidney HS showed reduced 3-O-sulfation in Hs3st3a1-/- and Hs3st3b1-/-, but not in Hs3st6-/- kidneys. Glomerular HS showed reduced HS staining and reduced ligand-and-carbohydrate engagement (LACE) assay, a tool that detects changes in binding of growth factor receptor-ligand complexes to HS. Interestingly, DKO mice have increased levels of blood urea nitrogen, although no differences were detected in urinary levels of albumin, creatinine and nephrin. Finally, transmission electron microscopy showed irregular and thickened GBM and podocyte foot process effacement in the DKO compared to WT. Together, our data suggest that loss of 3-O-HS domains disrupts the kidney glomerular architecture without affecting the glomerular filtration barrier and overall kidney function.
2024-07-05 | GSE261059 | GEO
Project description:The glomerular filtration barrier prevents large serum proteins from being lost into the urine. It is not known, however, why the filter does not routinely clog with large proteins that enter the glomerular basement membrane (GBM). Here we provide evidence that an active transport mechanism exists to remove immunoglobulins that accumulate at the filtration barrier. We found that FcRn, an IgG and albumin transport receptor, is expressed in podocytes and functions to internalize IgG from the GBM. Mice lacking FcRn accumulated IgG in the GBM as they aged and tracer studies showed delayed clearance of IgG from the kidneys of FcRn deficient mice. Supporting a role for this pathway in disease, saturating the clearance mechanism potentiated the pathogenicity of nephrotoxic sera. These studies support the idea that podocytes play an active role in removing proteins from the GBM and suggest that genetic or acquired impairment of the clearance machinery is likely to be a common mechanism promoting glomerular diseases. Keywords: Unique cell type expression analysis
2008-01-01 | GSE10017 | GEO