骨保护蛋白基因转染成骨细胞后的表达及对其生物学行为的影响

目的 将人骨保护蛋白(OPG)基因转入体外培养的人成骨细胞,研究OPG基因在转染成骨细胞中的表达,并分析OPG基因转染对成骨细胞生物学行为的影响.方法 首先行人成骨细胞体外培养,然后用质粒peDNA3.1-hOPG转染成骨细胞,应用逆转录-聚合酶链反应(RT-PCR)和Western blot检测转染细胞OPG mRNA和蛋白质的表达.观察OPG基因转染后成骨细胞的增殖情况,对成骨细胞表达的碱性磷酸酶(ALP)和骨钙素(BGP)的含量进行检测,观察转基因细胞的生物学特性.结果 RT-PCR和Western blot检测结果表明在OPG基因转染后,成骨细胞表达的OPGmRNA和蛋白质含量均较未转染组增加.在OPG基因转染后,可明显促进成骨细胞的增殖,成骨细胞表达的ALP和BGP的含量均显著上升,而且在量效图中可以观察到随着OPG基因转染剂量增加,其增加程度也增加.结论 成骨细胞可作为转基因的受体细胞,成功表达目的 基因.转染OPG基因的成骨细胞可稳定、高效的表达OPG.OPG基因转染成骨细胞生物学特性稳定,而且可明显促进转染成骨细胞的成骨特性的表达.

The expression of osteoprotegerin and the biological characteristics of the osteoblast transfected by human osteoprotegerin gene

Objective To investigate the osteoprotegerin (OPG) gene expression in transfected osteoblasts in vitro and the effects of OPG gene transfer on the biological characteristics of osteoblast. Methods The osteoblast was transfected by pcDNA3. l-hOPG,the OPG mRNA were detected by RTPCR and the OPG protein expression were determined by Western blot after 24h transfection. Osteoblast proliferation,and ALP activity,as well as BGP expression were determined in both groups. Results The OPG mRNA and protein expression were higher in the pcDNA3. 1-hOPG transfected osteoblasts than the non-transfected osteoblasts. The osteoblast proliferation, BGP and ALP contents of the transfected cells increased significantly than the control (P<0. 05). Conclusion The osteoblast transfected by OPG gene could express OPG stably and effectively. The osteoblast transfected by OPG gene keeps a stable biological characteristics,and the OPG gene transfection promotes the bone formation.