补肾活血汤调控Runx2及Osterix促进BMSCs成骨分化的作用研究

Effect of Bushen Huoxue Decoction on Regulating Runx2 and Osterix to Promote Osteogenic Differentiation of BMSCs

目的:探讨补肾活血汤提高骨髓间充质干细胞(bone mesenchymal cells,BMSCs)Runx2及Osterix基因表达,促进成骨分化的作用。方法:从双侧股骨和胫骨分离大鼠BMSCs并使用腺病毒载体转染BMSCs细胞,构建过表达Runx2细胞株,Q-PCR法对转染细胞株进行鉴定,CCK8法评价补肾活血汤对BMSCs活性的影响,ALP半定量活性检测法评价补肾活血汤对BMSCs成骨分化的影响,RT-PCR、Western Blot法检测补肾活血汤对成骨相关标志物Runx2和Osterix表达量的影响。结果:成功构建过表达Runx2的BMSCs细胞,补肾活血汤组在25μg/m-200μg/ml之内对细胞活性没有影响,与正常培养组比较补肾活血汤100μg/ml组ALP活性显著提高,与正常培养组比较补肾活血汤50μg/ml、100μg/ml组Runx2和Osterix的mRNA和蛋白相对表达量显著增加。结论:补肾活血汤能提高BMSCs中Runx2和Osterix的表达,促进成骨分化。

Objective： to investigate the effects of the Bushen Huoxue Decoction（ BHD） on the expression of Runx2 and Osterix and osteogenesis differentiation in bone marrow mesenchymal stem cells（ BMSCs）. Method： BMSCs were separated from bilateral femur and tibia of rats and lentiviral vector was transfected into the BMSCs with or without overexpressed Runx2. The transfected cells were identified by Q-PCR,ALP activity was recruited to evaluate differentiation effect of BHD; Cytotoxicity was evaluated by cell counting kit 8. Moreover,RT-PCR,Western Blot were recruited to detected the expression of Runx2 and Osterix. Results： lentitiral vector were transfected successfully into BMSCs with overexpressed Runx2; No Cytotoxicity was presented between the concentration 25 ~ 100 μg/ml; Compared with control group,ALP activities were improved after treatment with BHD; Compared with control group,with the treatment with BHD,the gene and protein expression level of Runx2 and Osterix were both up-regulated. Conclusions： BHD improved the expression of Runx2 and Osterix in BMSCs and promoted the osteogenesis differentiation of BMSCs.