Construction of human BMP2-IRES-HIF1αmu adenovirus expression vector and its expression in mesenchymal stem cells

Liu,Danping; Hu,Liang; Zhang,Zheng; Li,Quan  Ying; Wang,Guoxian

doi:10.3892/mmr.2012.1237

Construction of human BMP2-IRES-HIF1αmu adenovirus expression vector and its expression in mesenchymal stem cells

Authors:
- Danping Liu
- Liang Hu
- Zheng Zhang
- Quan Ying Li
- Guoxian Wang
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Affiliations: Department of Bone and Joint Surgery, First Affiliated Hospital of Liaoning Medical University, Jinzhou, Liaoning 121000, P.R. China, Department of Orthopaedics, the First People's Hospital of Jingmen, Hubei 448000, P.R. China, Department of Pharmacology, Liaoning Medical University, Jinzhou, Liaoning 121000, P.R. China
Published online on: December 14, 2012 https://doi.org/10.3892/mmr.2012.1237
Pages: 659-663

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Abstract

The present study aimed to construct a novel recombinant adenovirus expression vector Ad-BMP2-IRES-HIF1αmu that expresses human bone morphogenetic protein (BMP2) and mutant hypoxia-inducible factor 1α, and investigated its effects in promoting neogenesis of bone and angiogenesis. The recombinant adenovirus BMP2, HIF1αmu and pIRES2-EGFP expression vectors were constructed and transfected into HEK293A cells. The groups were divided into group A, transfection with Ad-BMP2-IRES-HIF1αmu; group B, transfection with Ad-HIF1αmu-IRES-hrGFP-1; group C, transfection with Ad-BMP2-IRES-hrGFP-1; group D, transfection with Ad-IRES-hrGFP-1; group E, not transfected. Adenovirus liquid was transferred into rabbit mesenchymal stem cells (MSCs) pretreated with dexamethasone at the best multiplicity of infection (MOI). The mRNA and protein expression of BMP2 and HIF1α were detected by RT-PCR and western blot analysis. Adenovirus was successfully packaged. The expression level of HIF1α mRNA in group A and B was markedly higher than that in groups C, D and E, showing a significant difference (P<0.01). There was a significant difference in the expression level of BMP2 mRNA between group A and C (P<0.05) and this was markedly higher than that in groups B, D and E (P<0.01). The protein expression level of HIF1α in group A and B was markedly higher than that in groups C, D and E (P<0.01). The protein expression level of BMP2 in group A and C was markedly higher than that in groups B, D and E (P<0.01). The human BMP2-IRES-HIF1αmu adenovirus expression vector was successfully constructed and the experimental groups formed bone and blood vessels prior to the positive and negative control groups.

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