title: Construction of human BMP2-IRES-HIF1αmu adenovirus expression vector and its expression in mesenchymal stem cells.
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.
authors.affiliation: Department of Bone and Joint Surgery, First Affiliated Hospital                     of Liaoning Medical University, Jinzhou, Liaoning 121000, P.R. China.
authors.forename: Danping
authors.initials: D
authors.lastname: Liu
authors.forename: Liang
authors.initials: L
authors.lastname: Hu
authors.forename: Zheng
authors.initials: Z
authors.lastname: Zhang
authors.forename: Quan Ying
authors.initials: QY
authors.lastname: Li
authors.forename: Guoxian
authors.initials: G
authors.lastname: Wang
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D055396:Bone Morphogenetic Protein 2 / Q000235:genetics* / Q000378:metabolism; D002478:Cells, Cultured; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics* / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism*; D009154:Mutation; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D014162:Transfection
chemical_list: C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D012333:RNA, Messenger