Smac过表达对食管鳞癌细胞Warburg效应和细胞凋亡的影响

Effects of Smac overexpression on Warburg effect and apoptosis in esophageal squamous cell carcinoma cells

目的:探究Smac过表达对食管鳞癌细胞Warburg效应和细胞凋亡的影响。方法:以含Smac过表达载体的慢病毒感染食管鳞癌细胞EC1和Eca109(Smac过表达组),以感染含空载体的慢病毒为阴性对照,以未感染细胞为空白对照。采用Western blot法检测3组细胞Smac蛋白的表达情况。用过氧化氢刺激Smac过表达组和阴性对照组细胞,利用线粒体膜电位检测试剂盒和Annexin V-FITC/PI双染法检测细胞线粒体膜电位和细胞凋亡情况,并用Western blot法检测细胞中凋亡信号通路相关蛋白[细胞色素C(Cyt-C)、Bcl-2、Caspase-3前体]以及Warburg效应关键蛋白[己糖激酶Ⅱ(HK2)和乳酸脱氢酶(LDH)]的表达。结果:EC1和Eca109 Smac过表达组Smac蛋白表达水平升高,提示成功构建了Smac过表达细胞株。经过氧化氢刺激后,与阴性对照组相比,Smac过表达组HK2和LDH表达水平下降,细胞线粒体膜电位降低,细胞凋亡率升高,Cyt-C的表达水平升高,Bcl-2和Caspase-3前体蛋白的表达水平下降(P<0.05)。结论:氧化应激刺激下,Smac过表达可抑制食管鳞癌细胞的Warburg效应,促进食管鳞癌细胞凋亡。

Aim:To study the effects of Smac overexpression on Warburg effect and apoptosis in esophageal squamous cell carcinoma(ESCC)cells.Methods:Eca109 and EC1 cells were infected with Smac overexpression lentivirus vector(Smac overexpression group),or mock-vehicle(negative control group),and the untreated cells were blank control group.The expression of Smac was measured by Western blot.The ESCC cells in Smac overexpression group and negative control group were stimulated by hydrogen peroxide,mitochondrial membrane potential was detected and apoptosis was observed by Annexin V-FITC/PI double staining.Finally,Western blot was employed to monitor the expressions of apoptosis signal pathway-related proteins such as cytokines cytochrome C(Cyt-C),Bcl-2,Caspase-3 precursor proteins and Warburg effect-key proteins including hexokinaseⅡ(HK2)and lactate dehydrogenase(LDH).Results:Eca109 and EC1 cells overexpressing Smac were constructed.Compared with negative control group,after hydrogen peroxide stimulation,the expressions of HK2 and LDH reduced in Smac overexpression group,the mitochondrial membrane potential was decreased,additionally,the percentage of apoptotic ESCC cells was significantly enhanced,the expression of Cyt-C was increased,while the expressions of Bcl-2 and Caspase-3 precursor proteins were decreased(P<0.05).Conclusion:Overexpression of Smac would inhibit Warburg effect and promote apoptosis of ESCC cells when treated with oxidative stress.