In Vitro Differentiation Of Human Bone Marrow Mesenchymal Stem Cells Into Vessel Endothelial Cells And The Bilogical Characteristics Of Vessel Endothelial Cells

Objective:To provide certain research foundation for the future application of tissue engineering,by studying the endothelial cells on the source of saphenous vein and bone marrow mesenchymal stem cells isolated and cultured which in vitro induced by chemical agents, and comparing the biological characteristics and cell function analysis of the two sources of endothelial cells, analyzing the feasibility of as seed cells for future application.Materials:10ml bone marrow was extracted from the sternum of surgical patients in sterile conditions; the saphenous venous segment was obtained from the undergoing CABG surgery.Methods:hECs were separated from the venous wall and cultivated by collagenase digestion method in vitro; hMSCs were separated and cultivated by density gradient centrifugation method in vitro and induced to endothelial cell differentiation hMSC-ECs by VEGF,bFGF and ECGF, observed the biology characteristics before and after of the cells, the induction rate of differentiated cells were determined by flow cytometry (FCM); The cellular phenotypes (CD31,CD34,Ⅷfactor,a-SMA, Vimentin) and collagen I amount by western blot were performed.function (NO,PGI-2,ET-1 and t-PA) were determined by immunohistochemistry, and analyzed by statistical comparison.Results:The endothelial cells separated and cultivated on the source of the human great saphenous vein in early days had a slower growth proliferation, Immunohistochemical staining showed positive on antigen CD34, CD31 andⅦfactor; and the separated and cultivated endothelial cells on the source of human bone marrow stromal cells grew fast, the growth became uniformly spindle-shaped and expanded exponentially. Immunohistochemical staining revealed that BMSCs were positive on a-SMA and Vimentin. After induction hMSC-ECs also expressed CD31, CD34 andⅧfactor related antigen. Western blot results showed that the three kinds of cells all had the function to secrete I type collagen; cell function showed that:PGI2 and ET-1 were much higher in hECs group than in other two groups (p<0.01), the content of t-PA in three groups had no significant difference (p>0.05); the NO content determination indicated, it had no differences between hECs group and hMSC-ECs group, yet all significant higher than in hMSCs group (p<0.05), after freeze-stored recovery the characteristics and functions.of the cells had no obvious change.Conclusion:Human autogenous saphenous vein sources of hECs and hMSCs bone marrow-derived are both successfully separated, cultivated and amplified in vitro.And that can be successful in vitro to endothelial method differentiation. The growth ability and cell function of hMSCs are stronger, then obtained relatively simple and has basically met the function of endothelial cells. hECs and hMSCs all can become tissue engineering seeds cells. Although the overall cell function of hECs is slightly above hMSCs. But hMSCs has special biological characteristics and extremely promised becoming the best choice seeded cells.