不同微环境对人骨髓间充质干细胞体外增殖和分化的影响  

Effects of different microenvironments on cell proliferation and differentiation of human bone marrow mesenchymal stem cells in vitro

作　　者：袁君杰[1] 谢幼专[1] 卢霄 卢建熙 

机构地区：[1]上海市骨科内植物重点实验室,上海交通大学医学院附属第九人民医院骨科,200011 [2]上海贝奥路生物材料有限公司,201114

出　　处：《国际骨科学杂志》2013年第6期435-438,共4页International Journal of Orthopaedics

基　　金：国家自然科学基金(81071453);上海交通大学医工交叉基金(YG2011MS24);上海高校创新团队建设基金(二期);上海市骨科内植物重点实验室建设基金(08DZ2230330);上海教委重点学科建设基金(J50206)

摘　　要：目的评价不同微环境对人骨髓间充质干细胞(hBMSC)体外增殖和分化的影响。方法采用全骨髓贴壁分离法培养hBMSC,传至第3代后分别在4种微环境下培养:10%FBS组、10%CS组、1 g/L植物源重组人血清白蛋白(OsrHSA)组、单纯DMEM组。采用CCK-8比色法测定细胞一周增殖率。以不加成骨诱导液为对照组,实验组添加成骨诱导液后第1周和第2周行碱性磷酸酶染色,第3周和第4周行茜素红染色,评估不同时段各组细胞成骨分化程度。结果采用单纯DMEM组培养hBMSC,细胞增殖率基本不变,后期下降;10%FBS组、10%CS组和1 g/L OsrHSA组均能显著提高hBMSC增殖率(P<0.05),以10%FBS组效果最好。碱性磷酸酶染色和茜素红染色显示,单纯DMEM组和对照组均未发现钙沉积阳性细胞;10%FBS组、10%CS组和1 g/L OsrHSA组在各阶段均出现阳性染色,随着诱导时间延长颜色明显加深;10%CS组和1 g/L OsrHSA组各时间段阳性染色程度明显较10%FBS组弱,10%CS组其次,1 g/L OsrHSA组最差。结论细胞外基质微环境对hBMSC生长分化至关重要,含血清的微环境能更好地促进hBMSC增殖及维持hBMSC分化特性。Objective To evaluate the influence of different microenvironments on cell proliferation and differentiation of human bone marrow meaencbymal stem cells （hBMSCs） in vitro. Methods The hBMSCs were cultured using whole bone marrow adherence separation method. The hBMSCs of primary culture and subculture＇ s first and second generation were cultured in DMEM containing 10% fetal bovine serum （FBS）. While, P3 was cultured in DMEM containing the following microenvironment ingredients respectively： 10 % FBS group, 10% calf serum （CS） group, 1 g/L oryza sativa recombinant human serum albumin （OsrHSA） group and nothing. The proliferation rate of one week of hBMSCs was evaluated by CCK-8 colorimetric assay. The one without osteogenesis induced liquid was considered as a control group, the experimental group was evaluated by alkaline phosphatase staining at 1 and 2 weeks, alizarin red staining at the 3 and 4 weeks after osteogenesis induced liquid added. During these time, the osteogenic differentiation＇s degree were estimated. Results The data from CCK-8 colorimetric assay showed that the proliferation rate of cell was unchanged in the early time, and decreased in the last among the pure DMEM group. Compared with the pure DMEM group, the group of using 10% FBS, 10%CS and 1 g/L OsrHSA could also significantly increase the differentiation of hBMSCs （P〈0. 05）, and 10%FBS group works best. The pure DMEM group and the control group had not been discovered calcium deposits positive cells through alkaline phosphatase staining and alizarin red staining. Positive staining occurred at any stage in the 10% FBS group, 10% CS group and 1 g/L OsrHSA group. The longer you induced, the deeper coulour you get. Compared with 10%FBS group, the degree of positive staining of 10%CS group and 1g/L OsrHSA group was significantly weaker in any stage. 10% CS group was the second weaker one, and the worst one is 1 g / L OsrHSA group. Conclusions The extracellular matrix microenviroment is essential for the proliferation a

关 键 词：微环境 骨髓间充质干细胞 增殖 分化 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]