Experimental Study On Rat Bone Marrow Mesenchymal Stem Cells-derived Neural Stem Cell Differentiation Into Vascular Endothelial-like Cells

Objective: To explore whether rat bone marrow mesenchymal stem cells-derived neural stem cells can differentiate into vascular endothelial-like cells or not in vitro.Methods: In this study,the whole bone marrow from femur,tibia and humerus of rats were isolated and cultured in DMEM/F12.To observe the morphological changes of bone marrow mesenchymal stem cells during culture and describe the growth curve of P3(passage 3)cells and explore the optimal inoculum density of BMSCs for culture.Flow cytometry was used to detect the expression of CD34,CD44,CD45,CD90.The P3 BMSCs were cultured in neural stem cell culture medium(20ng/mL bFGF,20ng/m L EGF,B27).During the culture,detecting the expression of CD133 and Nestin by immunocytochemistry.During the VEC culture,observing the morphological changes of the cells and detecting the expression of CD31 、 vWF by immunocytochemistry.And then,the neural stem cells were also cultured in Matrigel medium to observe weather form the capillary-like structure.Results: 1.At the beginning of culture,BMSCs were clustered and spindle-shape.The result of flow cytometry showed that the expression of CD44(91.4%)and CD90(93.7%)were positive,CD34 and CD45 were negative.2.The growth curve of P3 cells was S-type,and the optimal inoculum density was from 5×104/mL to1×105/m L.3.The cells were lumpish during the first 1 to 2 days,immunocytochemistry showed that the expression of CD133 and Nestin were positive.4.The cells were flat,triangular or polygonal after the neurospheres were cultured for7 days.The cells appearance seems to like paving stone shaped.The capillary-like structure can be formed in matrigel medium.Conclusion: 1.In vitro,the rat bone marrow mesenchymal stem cells can be induced to differentiate into neural stem cells in serum free suspension culture.2.Neural stem cells can be induced to differentiate into vascular endothelial-like cells in vitro,and the capillary like structure can be formed in matrigel gel.