胶质母细胞瘤肿瘤干细胞体外分离培养鉴定及生物学特性的研究

目的:探讨人脑胶质母细胞瘤(GBM)肿瘤干细胞的培养方法,观察其体外增殖、分化等生物学特性。方法:应用改良神经干细胞培养基,11例人脑GBM标本中悬浮培养GBM的肿瘤细胞。采用有限稀释法筛选分离具有连续克隆能力的单细胞。通过免疫组化及电镜观察的方法,确定其干细胞特征;通过核型分析探讨该细胞的肿瘤性质;将确定的脑肿瘤干细胞(BTSCs)接种于不含生长因子的干细胞培养基,观察其在体外条件下分化的特点。结果:本组样本中10例(1例污染)获得具有连续克隆和增殖能力及干细胞特征的肿瘤细胞。结论:人脑GBM中存在具有自我更新和增殖潜能的BTSCs,在体外可将其分离、培养和纯化。

Isolation, Culture and Identification and Biological Character Research of Brain Tumor Stem Cells in Glioblastoma Multiforme in Vitro

Aim:To explore the method of culturing human brain tumor stem cell in glioblastoma in vitro with reformed neural stem cell culturing medium,meanwhile to study biological characters on proliferation and differentiation and so on. Methods:Tumor cells were isolated in 11 cases of fresh-resected glioblastoma multiforme by limited dilution analysis and monoclonal formation assay. Immunohistochemistry dectection and electron microscope observation were conducted to determine if those isolated tumor cells were stem cell. And the karyotype analysis was conducted to understand the nature of the chromosome in those isolated cells. The differatiative features of" brain tumor stem cell"was observed,in vitro,by cultured in growth factor-free culture medium. Results:In 10 samples of glioblastoma( 1 was polluted) a small amount of tumor cells were obtained,which were of continuous proliferating and renewing,with positive immunohistochemistry-staining on Nestin and CD133. Electron microscope observation indicated those cell of immature cellular feature. Those cells showed the tumor karyotype,disclosing the cells were derived from tumor. Conclusion:There were a small amount of tumor cells in human brain glioblastoma which had capacity of continuous proliferating,renewing and multi-potent differentiation. Those tumor cells could be isolated,cultured and purified inv itro.