摘要
目的探讨体外培养和鉴定胚胎大鼠神经干细胞(neural stem cells,NSCs)及诱导其分化为类毛细胞。方法从SD系胚鼠大脑分离NSCs,在无血清培养基中培养,用含10%胎牛血清的DMEM/F12培养基诱导其分化为神经元和星形胶质细胞:将NSCs球放到含鼠尾胶原包被的盖玻片6孔培养板中培养,然后将乳鼠基底膜体外培养后汲取其上清液与NSCs一起培养,14～21天后通过免疫荧光和免疫组化法检测毛细胞标志物肌球蛋白(myosin)Ⅶa和钙视网膜蛋白(calretinin)。结果培养的NSCs胞体透亮,折光性好,分化后的细胞免疫组化示神经元特异性烯醇化酶、胶原纤维酸性蛋白、myosinⅦa和calretinin阳性。结论在无血清条件下能培养出活性很好的NSCs,并且能诱导其分化为神经元、星形胶质细胞和类毛细胞。
OBJECTIVE To study the culturing and identifying neural stem cells (NSCs) from rat embryos in vitro and inducing them to differentiate into hair-like cells.METHODS NSCs were isolated from brain of SD rat embryos and cultured in serum- free conditional medium.Then NSCs were induced to differentiate into neurons and astrocytes in DMEM/ F12 medium containing 10 % fetal bovine serum. The NSCs were added to rat tail collagen coated coverglasses in 6-well culture plates in DMEM/F12 medium containing 10 % fetal bovine serum.The basal lamina was isolated from P0 to P3 rats and cultured in the same medium.The supernatant was added to the NSCs suspension mentioned above and co-cultured for 14 to 21 days.Then expression of hair cell markers (myosin Vlla and calretinin) was detected by immunocytochemical analyses.RESULTS The cell bodies of NSCs were transparent with good refraction.After differentiation,the cells demonstrated neuron specific enolase (NSE),glial fibrillary acidic protein(GFAP),myosin Vlla and calretinin immunoreactively.CONCLUSION NSCs were cultured well in serum-free conditional medium and they could be induced to differentiate into neurons, astrocytes and hair-like cells.
出处
《中国耳鼻咽喉头颈外科》
北大核心
2008年第7期404-407,共4页
Chinese Archives of Otolaryngology-Head and Neck Surgery
基金
国家自然科学基金(30672308)
关键词
大鼠
胚胎
干细胞
毛细胞
Rats
Embryo
Stem Cells
Hair Cells
