脑肿瘤干细胞放射敏感性的实验研究

Experimental research of radiosensitivity for brain tumor stem cell

目的评价脑肿瘤干细胞(brain tumorstem cell,BTSC)与胶质瘤细胞系SHG-44细胞放射敏感性的差异。方法SHG-44细胞分别接种于含10%胎牛血清的DMEM/F12培养基和无血清的DMEM/F12培养基(添加bFGF和EGF)中。CD133免疫细胞化学染色鉴定BTSC。0、0.5、1、2、3、4、6、8、10、20GyX线照射SHG-44细胞和BTSC后以流式细胞仪检测两种细胞的细胞周期及细胞凋亡率,绘制SHG-44细胞和BTSC的存活曲线。结果SHG-44细胞中存在BTSC,后者能在无血清的DMEM/F12培养基中存活并悬浮生长,增殖形成克隆球,具有自我更新和增殖能力,表达特异性标志物CD133。两种细胞在X线照射后细胞周期无明显差异,BTSC的细胞凋亡率低于SHG-44细胞(P<0.05),细胞存活率高于SHG-44细胞(P<0.01)。结论BTSC的放射敏感性较SHG-44细胞明显降低,是胶质瘤放疗抵抗的主要原因。

Objective To evaluate difference of radiosensitivity for brain tumor stem cell（BTSC） and human glioma cell line SHG-44 cell.Methods SHG-44 was cultured in DMEM/F12 supplemented with 10% FBS,and in serum-free DMEM/F12 containing bFGF and EGF respectively.BTSC was detected by immunocytochemistrical staining with antibodies against CD133.BTSC and SHG-44 cell were irradiated by X-rays with doses of 0,0.5,1,2,3,4,6,8,10,20 Gy respectively,and flow cytometry assay was used to detecte the apoptosis and analyze the changes of cell cycles.Cell survival curves of BTSC and SHG-44 cell were determined by the measure of cell clone formation.Results SHG-44 cell contained BTSC,which could survive and grow by means of suspension in serum-free medium and proliferate into the clonal sphere.The clonal spheres maintained strong ability of self-renewal and proliferation.The specific marker CD133 could be expressed by BTSC.After BTSC and SHG-44 cell were irradiated by X-rays,flow cytometry assay showed that no different could be found in cell cycles and the ratios of apoptosis in BTSC were lower than that of SHG-44 cell significantly（P〈0.05）.The cell survival curves assay showed that the ratios of cell clone formation in BTSC was higher than that of SHG-44 cell（P〈0.01）.Conclusions Radiosensitivity of BTSC is lower than that of SHG-44 cell,and it is main cause of glioma radioresistence.