体外诱导人骨髓间充质干细胞分化为软骨细胞的研究  

Chondrocytic differentiation of mesenchymal stem cells in vitro

作　　者：黄建荣[1] 李建华[2] 李卫平[1] 沈慧勇[1] 王志勇[1] 宋卫东[1] 刘尚礼[1] 韩运[1] 

机构地区：[1]中山大学第二附属医院骨科,广州510120 [2]广州医学院生理学教研室,广州510182

出　　处：《中国临床解剖学杂志》2006年第6期662-665,共4页Chinese Journal of Clinical Anatomy

基　　金：广东省自然科学基金资助项目(5300677)

摘　　要：目的:探讨胰岛素样生长因子-I(IGF-I)和软骨细胞培养液体外诱导骨髓间充质干细胞(MSCs)向软骨细胞分化的可能性。方法:采用Percoll分离液分离培养人胚MSCs,体外扩增,用流式细胞仪测定MSCs的CD44、CD71、CD34、CD45表达;在第4代MSCs中加入的条件培养基进行诱导,根据条件培养基的不同分为4组:①IGF-I组:培养基中加入100ng/mLIGF-I;②软骨细胞培养液组:加入软骨细胞培养液;③联合诱导组:培养液中加入100ng/mLIGF-I和软骨细胞培养液;④对照组:培养基中不加入IGF-I和软骨细胞培养液。采用倒置显微镜观察、II型胶原免疫组化、细胞内蛋白多糖(PG)含量测定等方法判断诱导细胞的形态变化和表达软骨基质的能力。结果:体外培养的MSCs呈现成纤维细胞样形态,流式细胞仪检测结果显示,MSCs阳性表达CD44,阴性表达CD34,CD45;IGF-I和软骨细胞培养液联合诱导15d后,MSCs开始呈现软骨细胞的形态特点,II型胶原免疫组化呈阳性,而未诱导的MSCs组呈阴性;联合诱导组细胞内PG含量为8.92μg/ml,显著高于IGF-I组、软骨细胞培养液组和对照组,但低于正常软骨细胞(P<0.01)。结论:IGF-I和软骨细胞培养液能诱导MSCs向软骨细胞分化。Objective： To investigate the differentiation of human fetal mesenchymal stem cells （MSCs） exposed to insulin-like growth factors-Ⅰ （IGF-Ⅰ） and chondrocyte-derived medium into chondrocytes in vitro. Methods： Human fetal MSCs were isolated and cultured in DMEM. Cell surface antigen was analyzed by flow cytometry. And then MSCs were cultured with 100 ng/mL IGF-Ⅰ medium （IGF-Ⅰ group）, chondrocyte-derived medium （chondrocyte-derived medium group）, 100 ng/mL IGF-Ⅰ plus chondrocyte-derived medium （combined treated group）, or without IGF-Ⅰ and chondrocyte-derived medium （control group） respectively. After 15 days, cultures were assessed by collagen types Ⅱ immunohistochemistry and proteoglycan production. Results： The isolated and cultured human fetal MSCs displayed a fibroblast-like morphology. MSCs were uniformly positive for CD44, and did not express CD34 and CD45. MSCs in monolayer cultures treated with IGF-Ⅰ and chondrocyte-derived medium exhibited a chondrocytic phenotype and expressed collagen types Ⅱ. Proteoglycan contents in combined treated MSCs group were significantly higher than IGF-Ⅰ group, chondrocyte-derived medium group and control group. However, proteoglycan production of MSCs treated with IGF-Ⅰ and chondrocyte-derived medium remained lower than cultured chondrocytes. Conclusions： Human fetal MSCs can be induced and differentiated into chondrocyte-like cells when cultured in IGF-Ⅰ and chondrocyte-derived medium.

关 键 词：骨髓间充质干细胞 胰岛素样生长因子 软骨细胞 

分 类 号：Q343[生物学—遗传学]