淫羊藿次苷Ⅰ与其原型药物淫羊藿苷对rBMSCs成骨性分化的影响比较

Comparative study on the osteogenic differentiation effects of icariin and icariside Ⅰ in rat bone marrow stromal cells

目的观察淫羊藿次苷Ⅰ(IcarisideⅠ)与其原型药物淫羊藿苷(Icariin,ICA)对大鼠骨髓间充质干细胞(rat bone marrow stromal cells,r BMSCs)成骨分化过程中碱性磷酸酶(ALP)、骨钙素(BGP)蛋白表达量及ALP、BGP、Osterix和Runx-2 mRNA表达的影响。方法贴壁筛选法体外培养r BMSCs,以1×10-5mol/L的ICA和IcarisideⅠ对r BMSCs的成骨性分化进行药物干预。ELISA法检测ICA组、IcarisideⅠ组和空白对照组及转化液组之间ALP活性、BGP分泌量;RT Real-Time PCR法检测ALP、BGP、Osterix和Runx-2 mRNA基因表达。结果与空白对照组相比,IcarisideⅠ与其原型药物ICA均可显著增强r BMSCs的ALP活性,促进BGP的分泌,提高ALP、BGP、Osterix和Runx-2的mRNA水平(P<0.01)。IcarisideⅠ与ICA组比较,其ALP、BGP蛋白表达量及基因表达明显高于ICA组(P<0.01),Osterix和Runx-2的基因表达两组无显著性差异。结论 IcarisideⅠ也能促进r BMSCs成骨性分化,并且其活性高于原型药物ICA,也是淫羊藿发挥抗骨质疏松活性的主要成分。(2)提示补肾中药淫羊藿其补肾壮骨作用与促进ALP、BGP蛋白分泌量,上调ALP、BGP、Osterix和Runx-2的mRNA水平有关。

Objective To evaluate the effects of icariin and its major metabolite IcarisideⅠon the protein expression of ALP and BGP and the mRNA expression of ALP,BGP,Osterix and Runx-2 during osteogenic differentiation in rat bone marrowstromal cells（ r BMSCs）. Methods By using the adherencemethod,BMSCs were cultured. Icariin and IcarisideⅠwere supplemented into the culture at 1 × 10^-5mol/L,respectively. The experiment groups were blank control,positive control,Icariin and IcarisideⅠgroups.ELISA was performed to examine the protein expression of ALP and BGP on the 6th,9th,and 12 th day. The expressions of ALP,BGP,Osterix and Runx-2 genes were detected using RT Real-Time PCR. Results Compared with that of the blank control group,Icariin and IcarisideⅠsignificantly improved ALP activity and ALP and BGP amount（ P〈0. 01）,and enhanced the gene expressions of ALP,BGP,Osterix and Runx-2（ P〈0. 01）. Compared with the Icariin group,Icariside I had significantly stronger effect on the gene expressions of ALP and BGP protein（ P〈0. 01）,but there were no significant differences between the groups for Osterix and Runx-2 gene expressions. Conclusion 1） Icariside could promote the osteogenic differentiation of r BMSCs greater than Icariin,and is the main active anti-osteoporosis ingredient of the herb epimedii; 2） The function of the herb epimedii in strengthening bone of is associated with the increased production of ALP and BGP,and the upregulation of the mRNA expression of ALP,BGP,Osterix and Runx-2.