淫羊藿苷对小鼠脂肪干细胞成骨分化的促进作用

Effect of icariin on osteogenic differentiation of adipose-derived mesenchymal stem cells

目的 探讨淫羊藿苷(icariin,ICA)对小鼠脂肪干细胞(adipose-derived mesenchymal stem cells,ADSCs)成骨分化的促进作用。方法 分离培养2周龄C57BL/6J雄鼠的ADSCs,显微镜下观察培养细胞形态。采用成骨和成脂诱导分化培养液分别对细胞进行成骨和成脂诱导,茜素红和油红O染色检测其多向分化潜能。以含浓度为10^-7 mol/L淫羊藿苷的培养液培养作为实验组,单纯培养液培养作为对照组,RT-PCR检测细胞成骨分化相关基因Runt相关转录因子2(runt-related transcription factor 2,Runx2)、骨桥蛋白(osteopontin,OPN)、碱性磷酸酶(alkaline phosphatase,ALP)的表达,免疫荧光法分析细胞骨钙素(osteocalcin,OCN)的表达。结果 显微镜下观察培养细胞呈纤维细胞样梭形,呈簇分布。成骨分化茜素红染色及成脂分化油红O染色结果都呈强阳性。ICA实验组成骨相关基因Runx2、OPN、ALP的表达水平均显著高于培养基对照组,差异具有统计学意义( P <0.05),并且ICA实验组OCN蛋白的表达强于培养液对照组。结论 浓度为10^-7 mol/L的淫羊藿苷具有促进脂肪干细胞成骨分化的作用。

Objective To explore the effect of icariin(ICA) on osteogenic differentiation of adipose-derived mesenchymal stem cells(ADSCs). Methods ADSCs of C57BL/6J male mice aged 2 weeks were isolated and cultured, and the cell morphology was observed under light microscope. Multi-potency property of ADSCs was identified after osteoblasts and adipocytes induction, and tested by Alizarin red staining and oil red O staining, respectively. Culture medium containing 10^-7 mol/L icariin was used as ICA group and the culture medium alone was chosen as control group. The expression levels of runt-related transcription factor 2(Runx2), osteopontin(OPN) and alkaline phosphatase(ALP) were detected by RT-PCR, and the expression of osteocalcin(OCN) protein was analyzed by immunofluorescence staining. Results ADSCs appeared as spindle-shaped cells in small colonies. The differentiation of ADSCs into osteoblast and adipocyte-like cells was confirmed by positive Alizarin red staining and oil red O staining. The expression levels of Runx2, OPN and ALP in ICA group were significantly higher than those in control group( P <0.05). The protein expression of OCN in ICA group was higher than that in control group. Conclusion Icariin at a concentration of 10^-7 mol/L can promote the osteogenic differentiation of adipose-derived mesenchymal stem cells.