体外培养骨髓源性神经干细胞死亡情况的观察  

Death of bone marrow stromal cells-derived neural stem cells cultured in vitro

作　　者：沈雪[1] 徐如祥[1] 马宏伟[2] 姜晓丹[1] 刘智良[2] 邹雨汐[1] 

机构地区：[1]南方医科大学珠江医院神经外科,广东神经外科研究所,广州510282 [2]空军总医院神经外科,北京100036

出　　处：《中华神经医学杂志》2008年第2期118-121,130,共5页Chinese Journal of Neuromedicine

摘　　要：目的动态观察体外培养不同时间段骨髓源性神经干细胞（BMSCs-d-NSCs）的死亡情况，了解其体外培养条件下衰退和死亡的机制与规律。方法密度梯度离心法分离培养大鼠骨髓基质细胞，以神经干细胞专用培养基促使其向神经干细胞分化．用AO染色、Rhodamine123染色、TUNEL法及AnnexinV-EGFP，PI双染在荧光显微镜下直接观察．并以流式细胞仪检测不同培养阶段细胞凋亡和坏死情况。结果多种检测方法证实，BMSCs-d-NSCs在体外培养条件下1周、2周、3周、4周、8周均会出现不同程度的以凋亡为主的衰退和死亡现象，培养4周以上细胞凋亡和坏死情况较3周内明显增多。结论体外培养BMSCs-d-NSCs的衰退和死亡有凋亡和坏死两种机制的共同参与．并以凋亡为主。Objective To observe the death of bone marrow stromal cells-derived neural stem cells （BMSCs-d-NSCs） cultured in vitro, and study its mechanisms and regularity. Methods BMSCs were isolated using density gradient centrifugation, cultured, and induced to differentiate into NSCs using the special medium. The degeneration and death of cells were researched by the methods of AO staining, Rhodamine 123 staining, TUNEL, Annexin V-EGFP/PI double staining. Apoptotic and necrotic cells were observed by fluorescence microscope or phase-contrast microscope. The ratio of apoptotic and necrotic cells were detected by flow cytometry. Results Apoptosis and necrosis were confirmed in the cells cultured in vitro at 1, 2, 3, 4, 8 weeks, respectively, and the ratio of apoptotic cells were higher than the ratio of necrotic cells. The death rate of cells cultured for 4 weeks was higher than that for 3 weeks. Conclusion The mechanisms of BMSCs-d-NSCs cultured in vitro undergoing degeneration and death are apoptosis and necrosis, especially the former.

关 键 词：神经干细胞 细胞凋亡 坏死 细胞培养技术 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学]