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

Abstract EN

Objective.

To construct the eukaryotic expression vector hypoxia-inducible factor 1α-pcDNA3.1 and to investigate its transfective efficiency into mesenchymal stem cells (MSCs) in vitro and the expression of HIF-1α gene in MSCs.

Methods.

mRNA of Wistar Rats' myocardial cells was extracted, and cDNA was synthesized with Reverse Transcription Kit, HIF-1α was amplified by polymerase chain reaction (PCR), and constructed into pcDNA3.1.

Transfected HIF-1α-pcDNA3.1 into MSCs by liposome mediated method.

The expression of HIF-1α in the cells was detected by Western Blot Analysis and ELISA.

Results.

Eukaryotic expression vector HIF-1α-pcDNA3.1 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-1α gene transfected MSCs, the expression of HIF-1α mRNA and HIF-1α protein were both increased obviously.

Conclusion.

HIF-1α was cloned successfully.

HIF-1α-pcDNA3.1 can be transfected into MSCs by liposome-mediated method effectively and which resulting stable expression of HIF-1α in transfected MSCs.