Expression of osteogenic genes in rat bone marrow mesenchymal stem cells infected by lentivirus carrying hypoxia-inducible factor-1 alpha

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Abstract

BACKGROUND:

Human hypoxia-inducible factor-1 alpha can regulate the expression of osteogenic and angiogenic genes, and promote osteogenic activity.

OBJECTIVE:

To observe the expression of osteogenic genes in rat bone marrow mesenchymal stem cells carrying human hypoxia-inducible factor-1 alpha slow virus infection.

METHODS:

Hypoxia-inducible factor-1 alpha was obtained from Hela cells using RT-PCR. Lentivirus expression vector plasmid carrying hypoxia-inducible factor-1 alpha (Lenti-HIF-1α-eGFP) was constructed. 293Ta cells with LentiPac HIV mixed packaging plasmid was packaged, and then lentivirus was obtained. Rat bone marrow mesenchymal stem cells were isolated and cultured using direct whole bone marrow adherent method. Bone marrow mesenchymal stem cells were identified using flow cytometry. Bone marrow mesenchymal stem cells were infected with slow virus for 1, 4, 7 and 14 days. Bone morphogenetic protein-2, osteocalcin, osteopontin and alkaline phosphatase expression levels were detected in bone marrow mesenchymal stem cells using real-time fluorescent quantitative PCR.

RESULTS AND CONCLUSION:

Bone marrow mesenchymal stem cells were effectively infected with Lenti-HIF-1α-eGFP. Real-time fluorescent quantitative PCR results revealed that bone morphogenetic protein-2, osteocalcin, osteopontin and alkaline phosphatase began to obviously overexpress from 4 days after infection with Lenti-HIF-1α-eGFP until 14 days. Results suggested that hypoxia-inducible factor-1 alpha could elevate the osteogenic activity of bone marrow mesenchymal stem cells.

Subject headings:

bone marrow; mesenchymal stem cells; hypoxia-inducible factor 1, alpha subunit; lentivirus; transfection

Funding:

the Natural Science Foundation of Guangdong Province, No. S2012010008129; the Special Funds for the Development of Strategic Emerging Industries of Shenzhen City, No. 20120614154551201; the International Cooperation Project of Commission on Science and Technology Innovation of Shenzhen City, No. GJHZ20130412153906740; the Technology Development Project of Commission on Science and Technology Innovation of Shenzhen city, No. CXZZ20120614160234842

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