淫羊藿苷对成骨细胞成骨分化的影响及Wnt/-catenin信号系统的关系研究

摘要

目的 探讨淫羊藿苷对成骨细胞分化及其细胞内Wnt/-catenin信号通路的影响.方法 取SD大鼠,采用全骨髓贴壁法分离并体外培养骨髓间充质干细胞(rBMSC),并用贴壁筛选法进行体外培养rBMSC,给予淫羊藿苷3、6、9d后,观察对照组与淫羊藿苷处理组的碱性磷酸酶(ALP)活性,对碱性磷酸酶阳性克隆(CFUALP-F)以及矿化结节形成的影响.采用Western blot方法检测每组-catenin和Runx2组蛋白表达水,采用RT-PCR方法检测淫羊藿苷对Wnt10b、GSK 3、catenin及LEF1 mRNA水平的影响.结果 淫羊藿苷可以提高ALP的活性,碱性磷酸酶克隆数和矿化结节的形成.Western blot结果显示,淫羊藿苷可以提高-catenin和Runx2的蛋白表达水平;RT-PCR结果显示,淫羊藿苷可以提高Wnt10b、GSK-3、-catenin及LEF1表达水平.结论 初步证明淫羊藿苷可以激活Wnt/-catenin信号通路,参与并调控BMSC的成骨分化.%Objective:To investigate the effect of icariin on osteogenic differentiation of osteoblasts and Wnt/catenin signaling pathway in the cells.Methods:SD rats were selected and rat bone mnarrow derived mesenchymal stem cells(rBMSC) were isolated and cultured by whole bone marrow adherent method in vitro,with adherent screening method.On the 3rd,6th and 9th day being treated with icariin,alkaline phosphatase(ALP) activity was observed in the control group and icariin treatment group,the alkaline phosphatase positive clone (CFUALP-F)and the influence of the formation of mineralized nodules were observed.Western blot method was used to detect the expression of-catenin and Runx2 protein in each group.The effect of Wntl0b,GSK-3,-catenin and LEF1 mRNA levels were detected by RT-PCR method.Results:The activity of ALP,the number of alkaline phosphatase and the formation of mineralized nodules could be enhanced by the activity of the alkaline phosphatase.Western blot results showed that the protein expression levels of-catenin and Runx2 could be improved by the expression of the protein.RT-PCR results showed that the expression levels of Wnt10b,GSK-3,-catenin and LEF1 were increased.Conclusion:Icariin can activate the Wnt/-catenin signaling pathway and participate in and regulate the BMSC osteogenic differentiation.