Nephrin Is Expressed on the Surface of Insulin Vesicles and Facilitates Glucose-Stimulated Insulin Release

OBJECTIVE-Nephrin, an immunoglobulin-like protein essential for the function of the glomerular podocyte and regulated in diabetic nephropathy, is also expressed in pancreatic P-cells, where its function remains unknown. The aim of this study was to investigate whether diabetes modulates nephrin expression in human pancreatic islets and to explore the role of nephrin in P-cell function. RESEARCH DESIGN AND METHODS-Nephrin expression in human pancreas and in MIN6 insulinoma cells was studied by Western blot, PCR, confocal microscopy, subcellular fractionation, and immunogold labeling. Islets from diabetic (n = 5) and nondiabetic (n = 7) patients were compared. Stable transfection and siRNA knockdown in MIN-6 cells/human islets were used to study nephrin function in vitro and in vivo after transplantation in diabetic immunodeficient mice. Live imaging of green fluorescent protein (GFP)-nephrin-transfected cells was used to study nephrin endocytosis. RESULTS-Nephrin was found at the plasma membrane and on insulin vesicles. Nephrin expression was decreased in islets from diabetic patients when compared with nondiabetic control subjects. Nephrin transfection in MIN-6 cells/pseudoislets resulted in higher glucose-stimulated insulin release in Vitro and in vivo after transplantation into immunodeficient diabetic mice. Nephrin gene silencing abolished stimulated insulin release. Confocal imaging of GFP-nephrin-transfected cells revealed nephrin endocytosis upon glucose stimulation. Actin stabilization prevented nephrin trafficking as well as nephrin-positive effect on insulin release. CONCLUSIONS-Our data suggest that nephrin is an active component of insulin vesicle machinery that may affect vesicle-actin interaction and mobilization to the plasma membrane Development of drugs targeting nephrin may represent a novel approach to treat diabetes. Diabetes 59:190-199, 2010