淬灭胃癌SGC-7901细胞内活性氧对维生素E琥珀酸脂诱导内质网应激的影响

Reactive oxygen species scavenging on vitamin E succinate-induced endoplasmic reticulum stress response in SGC-7901 cells

目的:探讨淬灭胃癌SGC-7901细胞内活性氧(ROS)对维生素E琥珀酸脂(VES)诱导内质网应激的影响。方法:将不同浓度(1、5、10和20 mmol/L)N-乙酰半胱氨酸(NAC)作用胃癌SGC-7901细胞2 h,再加入20μg/mL VES处理12 h,经2,7-二氢二氯荧光黄双乙酸钠(DCFH-DA)染色后,采用激光共聚焦显微镜观察细胞内ROS水平。另将20 mmol/L NAC作用SGC-7901细胞2 h,再加入20μg/mL VES处理细胞12 h,经DCFH-DA染色后,流式细胞术进一步检测细胞内ROS水平。将20 mmol/L NAC作用SGC-7901细胞2h,再加入20μg/mL VES分别处理15和24 h后,分别采用逆转录PCR(RT-PCR)和Western blotting法检测内质网应激相关分子葡萄糖调节蛋白GRP78和CHOP mRNA及蛋白的表达。结果:与单纯VES处理组相比,20 mmol/L NAC预处理组胃癌细胞内ROS下降最为明显(P<0.05)。与对照组相比,20μg/mL VES能够诱导胃癌细胞内GRP78和CHOP m RNA及蛋白的表达明显增加(P<0.05);而与单纯20μg/mL VES处理组相比,20 mmol/L NAC对VES诱导GRP78和CHOP mRNA及蛋白表达具有显著的抑制作用(P<0.05)。结论:在体外,淬灭胃癌细胞内ROS能够抑制VES诱导的内质网应激反应。

OBJECTIVE:To investigate effects of reactive oxygen species(ROS)scavenging on vitamin E succinate(VES)-induced endoplasmic reticulum stress response(ERS)in human gastric cancer SGC-7901 cells.MATERIALS:SGC-7901 human gastric cancer cells were treated with 1,5,10,20 mmol/L N-acetyl-L-cysteine(NAC)for 2 h and then with 20μg/mL VES for an additional 12 h;ROS production was measured using confocal microscopy.In addition,cells were pretreated with 20 mmol/L NAC for 2 h and then with 20μg/mL VES for an additional 12 h;there after,ROS production was measured using flow cytometry.In addition,cells were pretreated with 20 mmol/L NAC for 2 h and subsequently treated with 20μg/mL VES for 15 h and 24 h,CHOP and GRP78 were determined by RT-PCR and Western blot.RESULTS:Pretreatment with NAC drastically decreased VES-induced ROS generation in SGC-7901 cells(P<0.05).NAC significantly inhibited induction of CHOP and GRP78 mRNA and protein expression in VES-treated SGC-7901 cells(P<0.05).CONCLUSION:ER stress response is an event downstream of the oxidative stress induced by VES in SGC-7901 cells.