Abstract
SummaryDuring nephrogenesis, stem/progenitor cells differentiate and give rise to early nephron structures that segment to proximal and distal nephron cell types. Previously, we prospectively isolated progenitors from human fetal kidney (hFK) utilizing a combination of surface markers. However, upon culture nephron progenitors differentiated and could not be robustly maintained in vitro. Here, by culturing hFK in a modified medium used for in vitro growth of mouse nephron progenitors, and by dissection of NCAM+/CD133− progenitor cells according to EpCAM expression (NCAM+/CD133−/EpCAM−, NCAM+/CD133−/EpCAMdim, NCAM+/CD133−/EpCAMbright), we show at single-cell resolution a preservation of uninduced and induced cap mesenchyme as well as a transitioning mesenchymal-epithelial state. Concomitantly, differentiating and differentiated epithelial lineages are also maintained. In vitro expansion of discrete stages of early human nephrogenesis in nephron stem cell cultures may be used for drug screening on a full repertoire of developing kidney cells and for prospective isolation of mesenchymal or epithelial renal lineages for regenerative medicine.
Highlights
26 million Americans, one in every nine, harbor kidney disease (Trivedi, 2010)
100 mm. (Right) Immunofluorescence staining for SIX2 and EpCAM in human fetal kidney (hFK) cultures shows that the described heterogeneous ‘‘niches’’ appear only in cells grown in mNPEM, but not in serum-free medium (SFM) or serum-containing medium (SCM)
50 mm and 100 mm. (C) A heatmap representing gene expression levels that were obtained from microfluidic multiplex qPCR of ‘‘bulk’’ RNA from freshly dissociated hFK cells and for hFK cells grown in mNPEM, SFM, or SCM for 7 days
Summary
26 million Americans, one in every nine, harbor kidney disease (Trivedi, 2010). Treatment options for patients with renal failure are limited. The alternatives available to patients who succumb to terminal renal disease are either supportive treatment in the form of dialysis or whole organ replacement by kidney transplantation. The shortage of donor organs and the long wait time on the recipient list hamper renal transplantation (Daar, 2006). The number of patients with terminal renal disease has increased, and the treatment costs for these patients exceed the cumulative costs of treating cancer patients (Trivedi, 2010). Due to the growing number of patients with kidney disease and the limited treatment options, alternative treatments are clearly in need
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