体外诱导人脐血间充质干细胞向软骨细胞分化的初步研究

Inducing differentiation of human umbilical cord blood mesenchymal stem cells into chondroblast in vitro

目的:探讨人脐带血间充质干细胞存在及体外向成软骨细胞分化的可能性。方法:将无菌条件下采集健康产妇脐带血,用相对密度为1.077g/ml的人淋巴细胞分离液分离脐血单个核细胞,利用间充质干细胞贴壁生长的特性获得MSCs,流式细胞仪检测其表面抗原标志。联合应用TGF-β1和IGF-Ⅰ及含有1×ITS+1的高糖DMEM进行诱导培养,并于诱导前及诱导后第21天留取细胞,甲苯胺蓝染色,免疫细胞化学法检测II型胶原的表达,分析是否诱导出成软骨细胞。结果:脐血MSC强表达CD44、CD105,不表达CD34、CD45。诱导21天甲苯胺蓝染色阳性,特异性表达II型胶原。结论:人足月脐带血中存在MSCs,在TGF-β1等生长因子诱导下可以向具有软骨细胞表型和功能的细胞分化。

Objective To explore whether mesenchymal stem cells could be isolated from human umbilical cord blood and would be able to differentiate into c hondroblast in vitro, Methods Human umbilical cord blood samples were obtained sterilely from healthy parturients. MSCs were isolated by lymphocyte separation medium （density 1.077g/ml）, MSCs were obtained by adhere culture. The surface antigen expression of MSCs was detected by flow cytometry. MSCs were induced with HG-DMEM medium which contained TGF-β1,IGF-Ⅰ and 1 × ITS＋1. Before induction and after 21 days induction ,the cells were stained by Toluidine Blue, the expression of Ⅱ collagens were examined by immunohistochemical staining. Results HUCB MSCs expressed CD44 and CD105 strongly, but CD34 and CD45 were expressed negatively. After 21 days of induction, the cells were positively stained by toluidine blue, and immunohistochemical analysis proved type Ⅱ collagen expression strongly. Conclusion MSCs can obtain from human umbilical cord blood following full-term normal delivery, and can be expanded and induced in vitro to differentiate into chondroblasts by inductive medium.