骨形态发生蛋白2和7共转染骨髓间充质干细胞的成骨分化

背景：骨形态发生蛋白最主要的作用是诱导骨形成，但因提取困难、代谢速度快、难以准确控制其使用浓度、价格昂贵等限制了其在体外和体内相关研究中的应用。目的：构建含特异细胞生长因子基因骨形态发生蛋白2，7的腺病毒载体，观察骨形态发生蛋白2和骨形态发生蛋白7基因共转染对兔骨髓间充质干细胞成骨分化的促进作用。方法：将全骨髓贴壁法分离得到兔骨髓间充质干细胞传至第3代，分为5组，空白组和常规诱导组分别以常规培养基和成骨诱导分化培养基培养；骨形态发生蛋白2， 7腺病毒转染组：分别单独以骨形态发生蛋白2、骨形态发生蛋白7腺病毒进行转染；联合转染组：以2种骨形态发生蛋白腺病毒联合转染。结果与结论：转染第7天，联合转染组成骨相关基因Runx-2、Osx、Ⅰ型胶原和碱性磷酸酶mRNA表达均较其他各组增高(P < 0.05)；骨形态发生蛋白2，7腺病毒转染组上述指标较空白组和常规诱导组增高(P < 0.05)。转染第14天，联合转染组4个指标均较其他各组明显增高(P < 0.05)；同时，4个指标表达均高于第7天(P < 0.05)。病毒转染后第7天，联合转染组Ⅰ型胶原和骨钙素蛋白表达均较其他组明显增高(P < 0.05)。提示对骨髓间充质干细胞进行骨形态发生蛋白2腺病毒和骨形态发生蛋白7腺病毒共转染后，较单基因转染和成骨诱导液诱导更能促进成骨相关基因和蛋白的表达。

Bone morphogenetic protein 2 co-transfected with bone morphogenetic protein 7 forosteogenic differentiation of bone marrow mesenchymal stem cells

BACKGROUND:The most important role of bone morphogenetic protein is the induction of bone formation some factors, including difficulty in sample extract, rapid metabolic rate, difficulty in accurate concentration control and high cost, limit the application for in vitro and in vivo studies.OBJECTIVE:To establish the adenoviral vectors containing bone morphogenetic protein 2 and bone morphogenetic protein 7 in order to observe the promotion effect of co-transfection of bone morphogenetic protein 2 and bone morphogenetic protein METHODS:The rabbit bone marrow mesenchymal stem cells separated by the whole bone marrow adherence method were passaged to the third generation, and divided into five groups: the control group and the conventional induction group were cultured with conventional culture medium and osteo-induction culture medium; bone morphogenetic protein 2 group and bone morphogenetic protein 7 group were transfected with bone morphogenetic protein 2 adenovirus and bone morphogenetic protein 7 adenovirus; the combination group was co-transfected with bone morphogenetic protein 2 adenovirus and bone morphogenetic protein 7 adenovirus. RESULTS AND CONCLUSION:At 7 days after transfection, the expression levels of Runx-2, Osx, alkaline phosphatase and Collagen Ⅰ mRNA in the combination group were higher than those in the other four groups (P < 0.05); the indicators above in the bone morphogenetic protein 2 group and bone morphogenetic protein 7 group were higher than those in control group and the conventional induction group (P < 0.05). At 14 days after transfection, the expression of Runx-2, Osx,alkaline phosphatase and collagen Ⅰ mRNA in the combination group were higher than those in the other four groups (P < 0.05); and the indicators at 14 days after transfection were higher than those at 7 days after trasnfection (P < 0.05). At 7 days after transfection, the expression levels of collagen Ⅰ and osteocalcin protein in the combination group were higher than those in the other four groups (P < 0.05). After co-transfected by both bone morphogenetic protein 2 and bone morphogenetic protein 7 adenoviral vectors, bone marrow mesenchymal stem cells could express high levels of osteogenesis related gene in both mRNA level and protein level than induced by conventional culture medium or osteo-induction culture medium.