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目的本研究旨在从U251胶质瘤细胞系中分离、培养和鉴定脑肿瘤干细胞,观察其的生长特征,为脑肿瘤干细胞的进一步研究打下基础。方法应用简化的无血清培养基和悬浮培养法培养U251细胞,并将获得的脑肿瘤干细胞接种于含血清培养基中观察其分化;交替用含血清培基和无血清培基培养U251细胞,观察其生长特性;用免疫荧光法检测脑肿瘤干细胞中CD133和Nestin的表达。结果 U251细胞接种至无血清培养基后,部分细胞能够存活并增殖为悬浮生长细胞球,并具有很强的自我更新和增殖能力;这种悬浮生长的细胞转移到含血清培基中后可贴壁分化;U251细胞在含血清培基和无血清培基中交替培养时能够快速转换生长方式;脑肿瘤干细胞中CD133和Nestin表达阳性。结论 U251胶质瘤细胞系中含一定比例脑肿瘤干细胞,此种细胞可在体外分离,培养和诱导分化。脑肿瘤干细胞的发现可能将为脑肿瘤研究提供全新的思路。
OBJECTIVE: To isolate, culture and identify brain tumor stem cells from U251 glioma cell lines and to observe the growth characteristics of these cells. This study will lay the foundation for the further study of brain tumor stem cells. Methods U251 cells were cultured in simplified serum-free medium and suspension culture medium. The obtained brain tumor stem cells were inoculated into serum-containing medium to observe the differentiation. U251 cells were cultured alternately with serum-free culture medium and serum-free culture medium. The growth characteristics of CD133 and Nestin in brain tumor stem cells were detected by immunofluorescence. Results After inoculation of U251 cells into serum-free medium, some cells could survive and proliferate into suspension-growing cytoplasm and have strong ability of self-renewal and proliferation. This kind of suspension-growth cells could be transferred to serum-containing culture medium Wall differentiation; U251 cells in serum-containing culture medium and serum-free medium when the rapid conversion of growth patterns; brain tumor stem cells in CD133 and Nestin expression was positive. Conclusion U251 glioma cell line contains a certain percentage of brain tumor stem cells, such cells can be isolated, cultured and induced differentiation. The discovery of brain tumor stem cells may provide a new idea for brain tumor research.