Expression of vascular endothelial growth factor gene in bone marrow mesenchymal stem cells / 吉林大学学报(医学版)

ABSTRACT

Objective To study the vascular endothelial growth factor(VEGF) expression and secretion of mesenchymal stem cells(MSCs) of rabbit transfected with pcDNA3-VEGF165 expression plasmid in order to construct a kind of tissue engineering artificial skull with more blood supply.Methods By gene reconstruction method,the VEGF165 gene was cloned into eukaryotic expression plasmid pcDNA3 and recombined eukaryotic expression plasmid pcDNA3-VEGF165 was constructed;By lipofectamine transfection method,pcDNA3-VEGF165 expression plasmid was transfected into MSCs of rabbit;By RT-PCR and Western blotting methods,the VEGF mRNA expression and VEGF protein secretion in the MSCs were detected.Results Recombined eukaryotic expression plasmid pcDNA3-VEGF165 was confirmed to be true by double enzyme digestion,two strips came to appear in 5400 bp and 600 bp of gelose electrophoresis and their sizes accorded with pcDNA3 plasmid and VEGF165 accordingly;Primarily cultured and subcultured MSCs of rabbit were successfully performed and the MSCs storehouse of rabbit was established,primary MSCs presented lymphoid form at first and then the morphology of them became circulara,polygonal or irregular forms,they were more and more like fibroblastic cells after subcultured cultivation.The expressions of VEGF mRNA and VEGF protein in the MSCs were found after transiently transfection by RT-PCR and Western blotting methods.Conclusion Recombined eukaryotic expression plasmid pcDNA3VEGF165 can be transfected into MSCs of rabbit effectively by lipofectamine and the VEGF expression can be detected in the MSCs after transfection.