维生素E琥珀酸酯诱导人胃癌细胞发生未折叠蛋白反应的研究.docxVIP

维生素E琥珀酸酯诱导人胃癌细胞发生 未折叠蛋白反应的研究 黄晓莉吴坤赵莎莎 山东大学齐鲁医院营养科哈尔滨医科大学营养与食 品卫生教研室 摘要： 目的:探讨维生素E琥珀酸酯(VES)对人胃癌细胞未折叠蛋白反应的影响。方法: 采用Western blot法分析VES不同剂量(0、5、10和20 u g/mL)处理人胃癌 SGC-7901 细胞 24 h 和 20 u g/mL VES 处理 SGC-7901 细胞不同时间(0、6、12、 18和24 h)对内质网膜上的跨膜蛋A PKR样内质网蛋白激酶(PERK)磷酸化和 活化转录因子6 (ATF6)活化的影响;釆用RT-PCR法检测VES不同剂量(0、5、 10 和 20 u g/mL)处理 SGC-7901 细胞 24 h 和 20 u g/mL VES 处理 SGC-7901 细胞 不同时间(0、3、6、9、12、15、18和24 h)对活化转录因子4 (ATF4) mRNA 表达和X盒结合蛋白1 (XBP1) mRNA剪切的影响。结果:在翻译水平上VES对磷 酸化PERK的诱导作用具有时间依赖关系，与阴性对照组比较，10 u g/mL VES的 诱导作用最强(P0. 05);在翻译水平上VES对ATF6活化片段的诱导作用具有时 间依赖关系和剂量依赖关系;在转录水平上，随着作用时间的延长，VES对ATF4 mRNA的诱导作用也不断增强，18 h达到高峰，而XBPlmRNA从VES作用6 h即开 始出现剪切。结论:VES诱导人胃癌SGC-7901细胞发生内质网应激过程中，未折 叠蛋口反应相关信号分子被活化。 关键词： 未折叠蛋白反应;维牛素E琥珀酸酯;胃癌细胞;内质网应激; 作者简介：黄晓莉，E-ma订:hxl0251010@163. com 收稿日期：2016-09-20 基金：国家自然科学基金资助项FI Vitamin E succinate induced unfolded protein response of human gastrie cancer SGC-7901 cells HUANG Xiaoli WU Kun ZHAO Shasha Department of Nutrition, Qilu Hospital of Shandong University; Department of Nutrition and Food Hygiene, Harbin Medical University; Abstract： OBJECTIVE: To investigate the effect of vitamin E succinate (VES) on unfolded protein response (UPR) in human gastrie cancer SGC-7901 cells. METHODS:SGC-7901 cells were treated with VES at 5, 10 and 20 p g/mL and with tunicamycin (TM) at 3 U g/mL for 24 h or at 20 u g/mL for up to 24 h. RNA-dependent protein kinase (PKR) -like ER kinase (PERK) and activating transcription factor 6 (ATF6) were evaluated using Western blotting. X-boxbinding protein 1 (XBP1) and activating transcription factor 4 (ATF4) were evaluated using RT-PCR. RESULTS:VES induced the phosphorylation of PERK in a time-dependent manner and reached the top level at 10 P g/mL. In add ition, VES induced the gen eration of p50ATF6 in SGC-7901 cells in a dose-and time-dependent manner. Like the positivc control, VES treatment at 20 u g/mL induced the spliced form of XBP1 from 6 h up to 24 h. Meanwhile, ATF4 mRNA was also up-regulated in cells exposed to VES