Effect Of Human Umbilical Cord Mesenchymal Stem Cells On Podocytic Apoptosis Induced By High Glucose

Background:Diabetic nephropathy (DN) is one of the most common complications of capillaries in patients with diabetes and has become the leading cause of end-stage renal disease (ESRD) with more and more patients with diabetes. Some studies have suggested a close relationship between the occurrence and development of DN proteinuria and podocytopathies, including podocyte apoptosis, detachment from the glomerular basement membrane, cellular hypertrophy and foot process fusion. More importantly, several experimental studies have shown that protecting podocyte can prevent the development or ameliorate renal injury in diabetes. At present, there is no satisfactory method or target for the treatment of DN. Currently available treatments involving strict glycemic and blood pressure control and renal replacement therapy. Mesenchymal stem cells (MSCs) have strong proliferation ability and multipotential differentiation, which have been widely used in curing lots of diseases, as well as DN. Our previously experiments have shown that MSCs injected via the tail vein into rats with DN alleviated kidney injury and reduced proteinurea, but the mechanism was not very clear. We used high glucose (HG) to induce podocytic apoptosis, and explored the effects of human umbilical cord mesenchymal stem cells(HUC-MSCs) on podocytic apoptosis.Objective:To explore the effects of HUC-MSCs on podocytic apoptosis induced by high glucose and the underlying mechanisms.Methods:HUC-MSCs were harvested from umbilical cords from full-term newborns, and then were characterized through adherent culture with fibroblast-like morphology, differentiation into osteocytes and adipocytes and flow-cytometric analysis of cell surface antigens. Mouse podocytes were mainly divided into four groups according to treatment:normal glucose (NG), mannitol control (NG+Ma), High glucose (HG) and HUC-MSCs co-culture group(HUC-MSCs). After 72 hours, cytometry and hoechst staining were used to detect the apoptosis rates.Western blot was used to mesure the ratio of active PARP to total PARP and the level of Bcl-2. ELISA was used to detect the secretion of hepatocyte growth factor (HGF). Neutralizing antibody (NtAb) of HGF was used to block its function and the recombinant human hepatocyte growth factor (rhHGF) was added to induce its function. The methods mentioned above were used to detect the apoptosis of podocytes.Results:1. HUC-MSCs were adherent cultured with a typical fibroblast-like morphology. They were able to differentiate into adipogenic and osteogenic cells. Flow cytometric analysis showed that MSCs were positive for CD90 (98.5%), CD 105 (64.3%), CD73 (96.3%), and negative for CD34 (0.3%), CD45 (0.5%).2. High glucose induced podocytic apoptosis in a time-dependent manner. After 72 hours, high glucose increased the podocytic apoptosis rate(P<0.05), the expression of PARP(P<0.05), decreased the expression of Bcl-2(P<0.05). HUC-MSCs co-culture decreased the podocytic apoptosis rate(P<0.05) and the expression of PARP(P<0.05), increased the expression of Bcl-2(P<0.05).3. The rhHGF decreased the podocytic apoptosis rate(P<0.05) and the expression of PARP(P<0.05), increased the expression of Bcl-2(P<0.05), similarly to HUC-MSCs. Blockade of HGF increased the podocytic apoptosis rate(P<0.05) and the expression of PARP(P<0.05), decreased the expression of Bcl-2(P<0.05).Conclusion:1. It is suggested that HUC-MSCs co-culture can reduce podocytic apoptosis induced by HG.2. It is suggested that HUC-MSCs ameliorate podocytic apoptosis probably through secreting soluble HGF.