人脐静脉来源间充质干细胞向神经样细胞分化的体外分离扩增实验

Differentiation of mesenchymal stem cells from human umbilical cord vein into nerve-like cells:An experiment on in vitro separation and amplification

目的:探讨人脐静脉来源的间充质干细胞的体外分离、纯化、扩增及向神经元样细胞的定向分化,为间充质干细胞治疗神经系统疾病提供实验依据。方法:实验于2004-06/2005-03在华中科技大学同济医学院附属协和医院血研所完成。无菌条件下取正常人脐静脉,10g/L胶原酶Ⅱ消化脐静脉内皮细胞,以IM DM作为培养基进行培养和纯化细胞,检测其生物学特点;用全反式维甲酸诱导,倒置相差显微镜观察诱导前后细胞的形态;免疫组织化学、免疫荧光化学和W estern blot方法检测分化前后细胞上的神经丝蛋白和胶质纤维酸性蛋白的表达。结果:脐静脉来源的细胞呈纤维样贴壁生长,具有骨髓间充质干细胞的生物学特征;诱导分化后,大部分间充质干细胞呈现典型神经样细胞形态,出现神经丝蛋白和胶质纤维酸性蛋白表达。结论:人脐静脉可以分离培养出间充质干细胞,并可以在体外定向分化为神经样细胞,为神经系统疾病的治疗提供细胞来源。

AIM： To investigate the isolation, purification, expansion and differentiation of mesenchymal stem cells （MSCs） derived from human umbilical cord vein into nerve-like cells in vitro, so as to provide experimental evidence for MSCs in treating disease of nervous system. METHODS： The experiment was finished in the.Institute of Hematology, Union Hospital affiliated to Tongji Medical College, Huazhong University of Science and Technology between June 2004 and March 2005. By 10 g/L collagenase II digestion, endothelial cells were isolated from human um- bilical cord vein and cultured in IMDM medium. Their biological charac- teristics were detected. Induced with all-trans-retinoic-acid （ATRA）, the morphologoca; changes of MSCs were observed under inverted phase contrast microscope. The expressions of neurofilament and glial fibrillary acidic protein were detected with immunohistochemistry, immunofluorescence and Western blot before and after differentiation. RESULTS： The cells from the cord vein displayed a fibroblast-like morphology adhering to the culture plate and shared the similar biological characteristics with MSCs from bone marrow. After the inducement, many MSCs turned into typical morphology of nerve-like cells and neurofilament and GFAP expressions appeared. CONCLUSION： MSCs can be derived from human umbilical cord vein and be induced into nerve-like cells in vitro. They could potentially be an excellent source of MSCs for the treatment of nervous system diseases.