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.
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.
Project description:We identified the gene Far2, encoding the fatty acyl-coA reductase 2, to be associated with mesangial matrix expansion (MME) in the mouse (PMID: 24009241). In order to verify this association we obtained the C57BL/6N-Far2tm2a(KOMP)Wtsi/2J (JR#018805) strain from The Jackson Laboratory's KOMP2 program and compared this strain to it's control strain (C57BL/6N) for several renal characteristics. At 6 months of age the knockout mice have a significantly better kidney function (measured as glomerular filtration rate) but the MME is at a comparable level. However, as MME increases in the control strain at 12 months of age, MME does not increase in the knockout until 18 months of age. In order to explore changes at the gene expression level, we compared RNA sequence reads from 6-month old kidneys. Our analysis showed a decrease of RNA expression for several tubular damage markers (NGAL, KIM-1) and an increase in several genes in the fatty acid metabolism pathway.
Project description:To investigate dynamic changes in glomerular cells, including podocyte, mesangial cells and glomerular endothelial cells, in the development of diabetic nephropathy We then performed gene expression profiling analysis using data obtained from RNA-seq of glomerular cells of control(m/m), diabetic (db-/- 6-week-old) and diabetic nephropathy (db-/- 10-week-old with albuminuria) mice
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.
Project description:Single cell RNAseq analysis of kidneys from P0 mouse demonstrated that expression of Vegfc by podocytes and mesangial cells is dispensable for glomerular development.
Project description:Available single-cell RNA-seq analyses have revealed that individual cells of the same type differ substantially in gene expression. We wonder whether glomerular mesangial cells, the cell type that support the unique structure of glomeruli, also exhibit big difference in gene expression among individual cells; and what biological information could be obtained from the single-mesangial cell RNA-seq data. Therefore, we isolated mouse glomeruli by Dynabead/magnetic concentration method, and digested them with enzymes to dissociate the cells. We loaded the single cell suspension to a Fluidigm C1 Single-Cell Auto Prep System for single cell cDNA preparation. The cDNA samples were analyzed with mesangial cell marker, Gata3,and the Gata3 positive samples were amplified and underwent sequecing using Illumina Highseq 2000 system.
Project description:Extrarenal viral infections commonly trigger glomerulonephritis mostly in association with immune complex disease. The immunoglobulin component of immune complexes can activate glomerular cell Fc receptors but whether complexed viral nucleic acids contribute to glomerular inflammation remains unknown. Glomerular mesangial cells express TLR3 but lack TLR7-9, hence, it is unclear whether mesangial cells can recognize and respond to viral ssRNA or DNA. Here we studied the immune responses activated by 3P-RNA (5'Triphosphate RNA) and Non-CpG DNA (Double stranded DNA) in primary mesangial cells (PMC). We used microarrays to detail the global programme of gene expression that induced by 3P-RNA and Non-CpG DNA. Keywords: diffrent ligands