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目的:研究Smac表达下调对吲哚美辛诱导的食管癌EC109细胞凋亡的影响,并探索其内在分子机制。方法:使用Smac-siRNA脂质体法转染食管癌EC109细胞,将细胞分为空白对照组,siRNA-control阴性对照组和siRNA-Smac组。使用不同浓度的吲哚美辛处理各组细胞,MTT检测细胞活力变化,流式细胞术检测其对细胞凋亡的影响,Western blot法检测Caspase-9、3以及XIAP、survivin表达情况。结果:采用不同浓度吲哚美辛处理后,细胞活力明显受到抑制;Smac siRNA可明显降低Smac在RNA水平和蛋白水平的表达;吲哚美辛可引起各组凋亡率明显增加,单纯导入siRNA-Smac片段并不能引起细胞凋亡变化,siRNASmac联合药物能明显降低EC109细胞凋亡率(P<0.05)。在蛋白水平,siRNA-Smac组的survivin表达无明显变化,XIAP表达明显增强;Caspase-9及Caspase-3的活性片段明显表达减弱(P<0.05)。结论:NSAIDs药物吲哚美辛可诱导食管癌EC109细胞凋亡,这种作用一定程度上依赖于Smac的正常表达;Smac表达降低后其对XIAP的抑制减弱,Caspase-9和Caspase-3的活化受到抑制,而survivin在其中并不起决定性作用。
AIM: To investigate the effect of Smac expression down-regulation on indomethacin-induced esophageal cancer EC109 cell apoptosis and explore its intrinsic molecular mechanism. Methods: Smac-siRNA liposomes were transfected into esophageal carcinoma EC109 cells and divided into blank control group, siRNA-control negative control group and siRNA-Smac group. The cells were treated with different concentrations of indomethacin. The changes of cell viability were detected by MTT assay. The apoptosis of cells was detected by flow cytometry. The expressions of Caspase-9, 3, XIAP and survivin were detected by Western blot. Results: After treated with different concentrations of indomethacin, cell viability was significantly inhibited; Smac siRNA significantly reduced the expression of Smac at the RNA level and protein level; indomethacin significantly increased the apoptosis rate in each group. -Smac fragment did not induce apoptosis, siRNASmac combined with drugs can significantly reduce the apoptosis rate of EC109 cells (P <0.05). At the protein level, there was no significant change in the expression of survivin in siRNA-Smac group, and the expression of XIAP was significantly increased. The active fragments of Caspase-9 and Caspase-3 were significantly decreased (P <0.05). Conclusion: Indomethacin, a NSAIDs drug, induces apoptosis of esophageal carcinoma EC109 cells. This effect depends to a certain extent on the normal expression of Smac. The decreased expression of Smac attenuates the inhibition of XIAP and the activation of Caspase-9 and Caspase-3 Inhibited, and survivin in which does not play a decisive role.