摘要
目的研究α-硫辛酸（LA）对大鼠胶质瘤细胞（C6细胞）缺氧复氧诱导损伤的保护作用，并初步探讨其作用机制。方法体外常规培养大鼠C6细胞，取对数生长期细胞用于实验。实验分正常组、模型组、10μmol／LLA组、25μmol／LLA组、50μmol／LLA组。后4组细胞建立缺氧复氧模型，缺氧前12h后3组细胞分别加入10、25、50μmol／LLA，正常组和模型组细胞加入等量培养液。复氧12h后MTT检测5组细胞存活率：2’，7’-二氯荧光黄双乙酸盐（DCFH-DA）荧光探针检测细胞内活性氧自由基（ROS）含量；流式细胞术、Western blotting分别检测细胞凋亡率和活化caspase-3蛋白的表达。结果与正常组比较，模型组、10、25、50μmol／LLA组细胞存活率降低，凋亡率增高；模型组细胞ROS含量增高，50μmol／LLA组细胞ROS含量降低；模型组、10μmol／LLA组细胞caspase-3蛋白表达增高，差异均有统计学意义（P〈0．05）。与模型组比较，25、50μmol／LLA组细胞存活率升高；10、25、50μmol／LLA组细胞ROS含量降低，凋亡率[（19．2±2．2）％vs（15．2±0．9）％、（13．1±0．7）％、（10．7±1．4）％]降低，caspase-3蛋白表达降低，差异均有统计学意义（P〈0．05）。结论LA对缺氧复氧诱导损伤的C6细胞具有保护作用，其机制可能与清除细胞内过量的ROS、抑制caspase-3途径依赖的细胞凋亡有关。
Objective To study the protective effects of α-lipoic acid （LA） on rat C6 glioma cells （C6 cells） exposed to hypoxia/reoxygenation and its mechanism. Methods Rat glioma cells C6 were conventionally in vitro cultured and cells at logarithmic growth phase were used in the experiment. Normal control group, oxygen-glucose deprivation/reoxygenation （OGD/ROG） model group, and OGD/ROG plus 10, 25 and 50 μmol/L LA treatment groups were chosen. C6 cells were exposed to OGD/ROG to simulate ischemiaJreperfusion in vitro. Different concentrations of LA （10, 25 and 50μmol/L） were added to the culture medium in the later three groups 12 h before hypoxia. Twelve h after C6 cells reperfusion, cell viability was analyzed by MTT assay; flow cytometry was used to evaluate the level of cellular reactive oxygen species （ROS） and Annexin V/PI staining to examine cell apoptosis rate; the expression of cleaved Caspase-3 protein was determined by Western blotting. Results As compared with normal control group, OGD/ROG model group, and OGD/ROG plus 10, 25 and 50 μmol/L LA treatment groups had significantly decreased cell viability and increased cell apoptosis rate （P〈0.05）; as compared with that in the normal control group, ROS level in the model group was statistically increased, while that in the 50 μmol/L LA treatment group was significantly decreased （P〈0.05）; as compared with that in the normal control group, Caspase-3 expression in the model group and 10 μmol/L LA treatment group was significantly increased （P〈0.05）. As compared with the model group, the 25 and 50 μmol/L LA treatment groups had significantly higher cell viability, the 10, 25 and 50μmol/L LA treatment groups had significantly lower ROS level, cell apoptosis rate and Caspase-3 expression （P〈0.05）. Conclusion LA has a protective effect on injury in C6 cells induced by hypoxiaJreoxygenation, whose mechanism may be related to LA eliminating excess cellular ROS, preventing occurrence of oxidative damage and thereby inhibiting Caspase-3 pathway-dependent apoptosis in C6 cells.
出处
《中华神经医学杂志》
CAS
CSCD
北大核心
2014年第6期581-585,共5页
Chinese Journal of Neuromedicine
基金
国家自然科学基金（81370449）
关键词
Α-硫辛酸
胶质瘤细胞
缺氧复氧
细胞凋亡
Alpha-lipoic acid
Glioma cell
Hypoxia/reoxygenation
Cell apoptosis
