摘要
目的:探讨差速贴壁法从早期人胚股骨分离、纯化骨髓间充质干细胞(MSCs)的可行性,并观察培养细胞是否向神经元样细胞诱导分化。方法:取2～3月龄新鲜健康流产人胚股骨,切碎,接种玻璃培养瓶内培养8～12h,待较多的成纤维细胞贴壁,立刻将未贴壁细胞转种至塑料培养瓶培养,并多次传代;用甲氧酚(BHA)和二甲亚砜(DMSO)诱导培养的细胞,免疫细胞化学方法检测诱导后细胞神经元烯醇化酶(NSE)、神经巢蛋白(Nestin)、胶质纤维酸性蛋白(GFAP)表达情况。结果:原代培养3d可见塑料培养瓶内有长梭形细胞生长,至第10天前后形成集落样克隆,传至第4代后呈单层漩涡状排列的长梭形的成纤维样细胞,已无明显细胞克隆形成,经多次传代,细胞保持良好的增殖能力和稳定的性状;免疫细胞化学方法显示未诱导细胞NSE、Nestin、GFAP阴性,诱导后细胞NSE、Nestin呈阳性,GFAP阴性,间接证实培养所得细胞是MSCs。结论:差速贴壁培养法从早期人胚股骨中分离纯化MSCs,去除可能的成纤维细胞污染,是简便有效的,可供有关研究参考选择。
Objective To investigate the feasibility of expansion and purification of bone marrow mesenchymal stem cells (MSCs) derived from human fetal thighbone in vitro and their differentiation into neuron cells. Methods Cells were isolated by differential adhesion from human fetal thighbone and expanded in culture medium. MSCs were induced to differentiate into neuron-like cells with dimethylsulfoxide (DMSO) and butylated hydroxyanisole (BHA). Specific markers were detected by immunohistochemistry. Results MSCs were purified and expanded to be undifferentiated cells in vitro with the culturing method. Immunohistochemistry staining showed that the induced MSCs expressed neuronspecific enolase (NSE) and nestin, but did not express glial fibrillary acidic protein (GFAP). Conclusion This culturing method of differential attachment is a reliable method of primary culturing the isolated MSCs from human fetal thighbone successfully, which can be used for further basic research and clinical applications.
出处
《东南大学学报（医学版）》
CAS
2005年第4期218-221,共4页
Journal of Southeast University(Medical Science Edition)
基金
国家自然科学基金资助项目(30271352)
江苏省自然科学基金资助项目(BK2001170)。
