Experimental Study of the Effects of Marrow Mesenchymal Stem Cells Transfected with Hypoxia‐Inducible Factor‐1α Gene

OBJECTIVE: To construct the eukaryotic expression vector hypoxia-inducible factor 1alpha-pcDNA(3.1) and to investigate its transfective efficiency into mesenchymal stem cells (MSCs) in vitro and the expression of HIF-1alpha gene in MSCs. METHODS: mRNA of Wistar Rats' myocardial cells was extracted, and cDNA was synthesized with Reverse Transcription Kit, HIF-1alpha was amplified by polymerase chain reaction (PCR), and constructed into pcDNA(3.1). Transfected HIF-1alpha-pcDNA(3.1) into MSCs by liposome mediated method. The expression of HIF-1alpha in the cells was detected by Western Blot Analysis and ELISA. RESULTS: Eukaryotic expression vector HIF-1alpha-pcDNA(3.1) was constructed successfully. Analyzed by flow cytometer, The MSCs' surfaces mark were CD44+, SH3(CD73)+, CD34-, CD45- and the CD44+ cells and SH3(CD73)+ cells were 94.7% and 97.3%, respectively, showing the high purity of the cultured MSCs. After inducing, the cultured MSCs can differentiate into osteoblasts and adipocytes successfully. In HIF-1alpha gene transfected MSCs, the expression of HIF-1alpha mRNA and HIF-1alpha protein were both increased obviously. CONCLUSION: HIF-1alpha was cloned successfully. HIF-1alpha-pcDNA(3.1) can be transfected into MSCs by liposome-mediated method effectively and which resulting stable expression of HIF-1alpha in transfected MSCs.