小鼠胚胎干细胞定向诱导分化为软骨样细胞的初步研究  

Preliminary study of chondrogenic differentiation of mouse embryonic stem cells

作　　者：王晓云[1] 丛笑倩[1] 刘晓音[1] 吴春芳[1] 吴娟娟[1] 殷德民[1] 刘伟[1] 曹谊林[1] 

机构地区：[1]上海市第二医科大学第九人民医院整复外科上海市组织工程研究重点实验室,上海200011

出　　处：《上海第二医科大学学报》2004年第4期250-253,274,共5页Acta Universitatis Medicinalis Secondae Shanghai

基　　金：国家重点基础研究发展规划资助项目("973"项目)(G1999054309);上海市"重中之重"重点学科基金资助.

摘　　要：目的 全反式维生素A酸(RA)联合转化生长因子-β1(TGF-β1)及骨形态发生蛋白-2(BMP2)等多种因子定向诱导小鼠胚胎干细胞(ESC)分化,探索这几种细胞因子对ESC的成软骨定向诱导分化作用。方法 小鼠ESC在小鼠胚胎成纤维细胞(MEF)饲养层上培养扩增,在1×109mol/L全反式RA条件下制备拟胚体(EB),贴壁后联合应用TGF-β1及BMP2诱导分化,经形态学、免疫细胞化学和RT-PCR检测细胞性质。结果 ESC在悬浮培养下,经RA初步诱导后给予TGF-β1及BMP2等因子继续诱导,贴壁后EB周边可爬出多角形分化细胞,诱导分化细胞和EB表达Ⅱ型胶原和软骨发生早期特异性转录因子Scleraxis。结论 在RA预诱导EB分化后,TGF-β1和BMP2等因子可定向诱导小鼠ESC表达软骨特异性相关基因和蛋白,为胚胎干细胞成为软骨组织工程种子细胞提供了可能。Objective To study the differentiation of mouse embryonic stem cells ( ESC ) induced by retinoic acid (RA) , transforming growth factor-β1 (TGF-β1 ) and bone morphogeneic protein-2 (BMP2) , and explore chondro-genic induction effect of these factors. Methods After being in vitro cultured and expanded, mouse ES cells were made into embryoid bodies (EBs) with the medium added 1×10-9mol/L all trans-RA. The EBs were induced by TGF-β1 , BMP2 and other factors. The induced cells derived form EBs were analyzed by morphology, immunocyto-chemistry and reverse transcription-polymerase chain reaction (RT-PCR). Results EBs were formed by ES cells cultured in suspension with RA for five days. When attached to the dish, EBs would give rise to a large number of polygonal cells given TGF-β1 , BMP2 and other induction factors. Induced EBs and differentiation cells expressed type Ⅱ collagen and transcription factor of Scleraxis, which usually expresses during early stage of embryo chondrogenesis. Conclusion After 1×10-9mol/L RA pre-stimulating the differentiation of EB, TGF-β1, and BMP2 can induce mouse ES cells to express specific gene and protein of chondrocytes, which provided the possibility for ES cells to become seed cells for cartilage tissue engineering.

关 键 词：小鼠 胚胎干细胞 定向诱导 细胞分化 维生素A酸 转化生长因子β1 骨形态发生蛋白-2 软骨组织工程 

分 类 号：Q813.5[生物学—生物工程] R318.17[医药卫生—生物医学工程]