摘要
目的:探讨纯度较高的小鼠脑微血管内皮细胞的分离与原代培养。方法:取8～14周龄BALB/c或C57/BL6小鼠,无菌分离脑组织,采用改进的两种方法即2次酶消化和1次密度梯度离心法及1次酶消化和1次密度梯度离心法分离小鼠脑微血管段,接种于涂布有Ⅳ型胶原的培养皿上进行原代培养,相差显微镜下观察细胞形态,并以免疫荧光法鉴定Ⅷ因子相关抗原。结果:改进的两种方法均可见内皮细胞在接种后12～16 h从微血管段周围爬出,5～7 d时细胞呈"漩涡状"生长,并可见细胞汇合,90%以上培养的细胞Ⅷ因子相关抗原表达阳性。结论:改进后的两种方法均能进行较高纯度的小鼠腑微血管内皮细胞的分离和原代培养。
Objective： To explore the method of primary culture of the mouse cerebral microvascular endothelial cells （MCMEC）. Methods： The mouse brains were isolated from 8-14 weeks BALB/c or C57/BL6 mice. Two modified methods, two-step enzyme digestion with one-step gradient centrifugation and one-step enzyme digestion with one-step gradient centrifugation, were used to isolate relatively pure cerebral microvessel fragments. The microvessel fragments were seeded and cultured on dishes coated with type Ⅳ collagen. The MCMEC were identified according to the morphological observation under the phase contrast microscopy and the detection of factor H-associated antigen by immumofluorescence method. Results ： By the two modified methods, cells in migrating from microvessel fragments were observed 12-16 hours after seeding, and became spindle-shaped and confluent after 5-7 days. More than 90% of the cells expressed factor H-associated antigen. Conclusion： The primary culture and its purification in MCMEC has been suc- cessfully established in using these two modified methods.
出处
《解剖学杂志》
CAS
CSCD
北大核心
2007年第5期530-533,630,共5页
Chinese Journal of Anatomy
基金
国家自然科学基金(30271210)
关键词
脑微血管内皮细胞
细胞
培养的
小鼠
cerebral microvascular endothelial cells
cell, cultured
mouse
