淫羊藿苷促进入骨髓间充质干细胞成软骨分化的研究  

The experimental study on icariin promoting chondrogenic differentiation of human bone marrow mesenchymal stem cells

作　　者：肖军[1] 马德彰[1] 杨钟华[1] 张山锋[1] 刘军[1] 

机构地区：[1]华中科技大学同济医学院附属普爱医院骨科,武汉430034

出　　处：《中华实验外科杂志》2013年第7期1357-1359,共3页Chinese Journal of Experimental Surgery

摘　　要：目的探讨应用淫羊藿苷促进人骨髓间充质干细胞（hMSCs）成软骨分化的可行性及效果。方法体外分离培养hMSCs，取第3代细胞实验。将hMSCs分别用无血清高糖培养基（H．DMEM）和含1×10-8、1×10-7、1×10-6、1×10-5mol／L淫羊藿苷的软骨诱导液（CM）诱导，显微镜下观察细胞形态变化，噻唑蓝（MTT）法检测各组细胞的增殖。以5×10 9/L的细胞密度离心构建微团，诱导培养3周。逆转录一聚合酶链反应（RT—PCR）检测各组蛋白多糖和Ⅱ型胶原mRNA的表达。免疫组织化学、甲苯胺蓝染色检测Ⅱ型胶原和蛋白多糖表达。结果5组微团分别由无血清H．DMEM、含1×10-8、1×10-7、1×10-6、1×10-5mol／L淫羊藿苷的CM诱导3周后，除H—DMEM组无Ⅱ型胶原和蛋白多糖表达外，其他各组Ⅱ型胶原蛋白和蛋白多糖的表达呈浓度依赖性增加，ImageProPlus6．0软件测得各组Ⅱ型胶原平均吸光度（A）值分别为0．0214±0．0035、0．1182±0．0105、0．1886±0．0184、0．2903±0．0261、0．3476±0．0287，各组差异均有统计学意义（P〈0．05）。结论淫羊藿苷能促进hMSCs成软骨分化，软骨分化程度呈浓度依赖性增加。Objective To explore the feasibility and effect of icariin on promoting chondrogenic differentiation of human bone marrow mesenchymal stem cells （hMSCs）. Methods hMSCs were isolated and cultured in vitro, and the cells from Passage 3 were used in this study, hMSCs were randomly grouped into five according to the kinds of induction medium-serum-free DMEM-H, chondrogenic medium supple- mented with 1×10-8、1×10-7、1×10-6、1×10-5mol/Licariin. The proliferation of hMSCs was inves- tigated by methyl thiazol tetrazolium （MTT） assay. The cells were resuspended and pellets were construc- ted by centrifugation at a density of 5 ×109/L, and then continued to be kept for three weeks. Reverse transcriptase-polymerase chain reaction （RT-PCR） was performed to detect the expression of mRNA of type II collagen and aggrecan. Immunohistochemistry and toluidine blue staining were performed to detect the expression of type II collagen and proteoglycan. Results After three weeks＇ induction, Group A did not express mRNA of type II collagen and aggrecan, the mRNA transcripts of type ]] collagen and proteogly- can in other four groups kept increasing in a concentration-dependent manner. The mean absorbanee （A） values of collagen type ]] were, respectively, （0. 0214 ± 0.0035）, （0. 1182 ± 0.0105）, （0. 1886 ± 0. 0184）, （0. 2903 ±0. 0261 ）, （0. 3476 ±0. 0287） with significant differences （P 〈0. 05）. Conclusion Icariin can promote chondrogenic differentiation of hMSCs. Treated with a certain concentration of icariin, the expression of type 11 collagen and proteoglycan increases in a concentration-dependent manner.

关 键 词：成人骨髓间充质干细胞 淫羊藿苷 微团培养 软骨分化 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]