高氧促进胶质瘤干细胞分化致化疗增敏的体外研究  

Hyperoxia-induced enhanced sensitivity to chemotherapy of glioma stem cells in vitro

作　　者：万文武[1] 汪攀[1] 兰川[1] 吴南[1] Wan Wenwu Wang Pan Lan Chuan Wu Nan(Department of Neurosurgery, Key Laboratory of Neurotrauma Prevention and Treatment, Southwest Hospital, Third Military Medical University, Chongqing 400038, Chin)

机构地区：[1]第三军医大学附属第一医院神经外科,全军神经外科研究所,全军神经创伤防治重点实验室,重庆400038

出　　处：《中华神经外科杂志》2017年第3期293-298,共6页Chinese Journal of Neurosurgery

基　　金：国家自然科学基金（81272784,81672493）

摘　　要：目的通过体外研究初步探讨高氧诱导胶质瘤干细胞分化致化疗增敏的效应。方法干细胞培养基诱导胶质瘤干细胞（U87、GL261），免疫荧光染色鉴定肿瘤干细胞标记蛋白的表达。胶质瘤干细胞在不同氧浓度条件（高氧95％O2、常氧21％O2）处理后采用免疫印迹法、实时荧光定量PCR（qRT—PCR）检测干性标记蛋白、分化蛋白、耐药蛋白的表达情况。胶质瘤干细胞在不同氧浓度条件下预处理后添加替莫唑胺干预，行半数抑制浓度（IC50）及凋亡检测。结果荧光检测表明干细胞培养基诱导的胶质瘤细胞高表达干细胞标记蛋白CD133、Nestin，同时高表达耐药标记蛋白MGMT。qRT—PCR及Western blot结果提示，高氧干预胶质瘤干细胞后，其干细胞标记蛋白CD133、Nestin表达水平均低于常氧处理组（P〈0．05），分化标记蛋白GFAP表达水平增高（P〈0．05），而耐药标记蛋白MGMT则显著降低（P〈0．05）。胶质瘤干细胞不同氧浓度预处理后添加替莫唑胺干预，凋亡结果提示高氧组凋亡率显著高于常氧组（P〈0．05）；而IC50检测提示，高氧组IC50值显著低于常氧组（P〈0．05）。结论高氧通过抑制胶质瘤干细胞干性表达并促进其分化致化疗增敏，本研究为高压氧联合化疗治疗胶质瘤并改善胶质瘤患者的预后提供新的理论依据。Objective To investigate the enhancement effect of hyperoxia on chemotherapy sensitivity of glioma stem cells in vitro. Methods The U87 and GL261 glioma stem cells were cultivated by stem cell culture medium, and the expression levels of labeled proteins of tumor stern cells were identified with immunofluoreseence staining. The U87 and GL261 glioma stem cells were both treated under high （ 95% ） oxygen and normal （21%） oxygen. The Western Blot and qRT-PCR were applied to detect the expression levels of stem cell labeled proteins, differential proteins and resistant proteins after the treatment. After the pretreatment of U87 and GL261 glioma stem cells under different oxygen concentration, the chemotherapy drug temozolomide （TMZ） was added in the culture medium of each group and ICs0 and apoptosis detection was subsequently conducted. Results The fluorescence detection results showed that U87 and GL261 glioma stem cells cultured in stem cell medium highly expressed the labeled proteins of tumor stem ceils including CD133 as well as Nestin and the drug resistance marker protein MGMT. The expression levels of CD133, Nestin and MGMT were lower after the treatment of hyperoxia in glioma stem cells detected with qRT-PCR and Western-blot （ P 〈 0.05 ）, while the differential labeled protein GFAP were much higher （ P 〈 0.05 ）. The apoptosis tests showed that the apoptosis value in TMZ-treated high oxygen group was significantly higher than that in the normal oxygen group （P 〈 0.05 ）. Similarly, the ICs0 value in the TMZ-treated high oxygen group was significantly lower than that in the normal oxygen group （P 〈 0. 05 ）. Conclusions High oxygen environment could promote the chemotherapy sensitivity of gliomas by inhibiting the property expression of the glioma stem cells. This study has provided a new theoretical foundation for the glioma treatment using high pressure oxygen combined with chemotherapy which might improve the outcomes.

关 键 词：神经胶质瘤 肿瘤干细胞 高氧 化疗增敏 

分 类 号：R739.41[医药卫生—肿瘤]