Kidney Cell and Organ Culture
A goal of our Center is to generate cell cultures from nephrons and isolate embryonic kidneys for organ cultures. The performance of cell- and organ-based assays will allow investigators to identify and characterize biochemical and molecular mechanisms controlling kidney cell and organ function.
The VCKD Cell Culture Facility will aid researchers in performing cell-based assays by developing nephron-segment specific cell populations from wild type and genetically altered animal models. Three different preparations of kidney cells can be performed:
- Primary cell culture. These cells have the advantage that their phenotype most closely resembles that of the nephron segments from which they were originally derived. However these cells cannot be propagated for long periods of time and can only be passaged a few times. In the case of cells prepared from conditional knockout mice, the cells can either be generated from mice that have already been crossed with the appropriate Cre mice or can be generated from floxed mouse and then infected with adenoCre in vitro.
- Immortalized cells. These cells are made by transfecting the primary cell populations with temperature sensitive conditional SV40 large T antigen, which results in transformation of the cell population. These cells can be easily propagated.
- Conditionally immortalized cells. These cells are obtained by crossing the transgenic mice of interest with mice carrying a conditionally immortalized temperature sensitive SV40 large T gene (called “Immortomice”), following which cells are isolated and grown. The advantage of these cells is that they can be easily propagated and their phenotype most closely resembles that of the nephron segments from which they were originally derived.
The VCKD Organ Culture Facility will aid researchers in performing kidney organ based assays by isolating E12.5 mouse embryonic kidneys from wild type and transgenic mice. Embryonic kidneys can be cultured on transwells for 3 days and the number of branches quantified. This assay is a great model for defining signaling pathways and mechanisms that regulate branching morphogenesis of the ureteric bud.
Expertise in isolation of mesangial and glomerular endothelial cells
Expertise in isolation of medullary interstitial cells
Expertise in isolation of podocytes and proximal tubule cells
Novel methods for kidney cell immortalization
Expertise in isolation of collecting duct cells and embryonic kidney culture