ABSTRACT
BACKGROUND:Epidermal growth factor is an auxiliary growth factor,but its effect on the growth of bone marrow mesenchymal stem cells (BMSCs) is uncertain.OBJECTIVE:To establish a mature method for isolation,extraction and identification of rat BMSCs,to investigate the effects of epidermal growth factor (EGF) on the proliferation and migration ability of BMSCs and to explore its potential mechanisms at the same time.METHODS:Rat BMSCs were isolated and purified using the improved whole bone marrow adherence method.After the cells were subcultured to the third generation,we detected the expression of cells surface antigens CD29,CD45 and CD90 by flow cytometry.BMSCs were further identified by osteogenic and adipogenic differentiation.Meanwhile,the effect of EGF on the proliferation of passage 3 BMSCs was measured by cell counting kit-8 and clonogenic assay.And the migration of P3 cells was verified by Transwell chamber.In addition,we detected the expression of proteins related to PI3K/Akt and nuclear factor-κB signaling pathways by western blot assay.RESULTS AND CONCLUSION:The primary BMSCs were polygonal and spindle-shaped,and then gradually appeared to be spindle-shaped.The results of flow cytometry demonstrated that the passage 3 cells were positive for CD29 and CD90,but negative for CD45.Furthermore,we successfully induced the osteogenic and adipogenic differentiation of BMSCs in vitro.Additionally,our data demonstrated that EGF promoted the proliferation and migration of passage 3 BMSCs.The relative expression levels of p-Akt and Bcl-2 of PI3K/Akt signaling pathway was up-regulated and the expression of Bax was down-regulated.At the same time,the relative expression level of phosphorylated p65 of nuclear factor-κB signaling pathway was up-regulated and the expression of phosphorylated inhibitor κB was down-regulated.Moreover,the downstream protein of matrix metalloproteinase-9 was up-regulated.Those proteins were related to the migaration of BMSCs.In summary,our results suggest that EGF could promote the proliferation and migration of BMSCs.