幽门螺杆菌对胃黏膜上皮细胞自噬和凋亡的影响  

Effect of Helicobacter pylori infection on autophagy and apoptosis of gastric epithelial cells

作　　者：吕杰[1] 李柏青[2] 刘勇[1] 马丽娜[1] 刘婷婷[1] 刘丛森[1] 

机构地区：[1]蚌埠医学院微生物学教研室安徽省感染与免疫重点实验室,安徽蚌埠233030 [2]蚌埠医学院免疫学教研室安徽省感染与免疫重点实验室,安徽蚌埠233030

出　　处：《蚌埠医学院学报》2014年第6期705-710,共6页Journal of Bengbu Medical College

基　　金：安徽省高等学校省级优秀青年人才基金重点资助项目(2013SQRL049ZD);蚌埠医学院科技发展基金重点项目(bykf13A07)

摘　　要：目的：探讨幽门螺杆菌（Hp）对胃黏膜上皮细胞（GES-1）自噬和凋亡的影响。方法：Hp与GES-1体外共培养，同时设立雷帕霉素（RAPA）、3-甲基腺嘌呤（3-MA）干预组及空白对照组，于2、4、6、12及24 h终止感染，通过免疫蛋白印迹、荧光染色、透射电镜、流式细胞术及细胞内活菌计数等方法观察Hp对GES-1自噬和凋亡的影响。结果：Hp＋GES-1组在2 h出现自噬标志蛋白LC3-Ⅱ表达及细胞内荧光自噬颗粒，透射电镜观察到细胞内出现双层或多层膜包裹的自噬小体样结构，细胞自噬在4 h达高峰，感染中后期逐渐减弱并消失；RAPA＋Hp＋GES-1组细胞自噬程度进一步增强，细胞凋亡率及细胞内活菌数均降低（P〈0.05-P〈0.01）；3-MA＋Hp＋GES-1组细胞自噬强度显著降低，而其细胞凋亡率和细胞内活菌数均明显升高（P 〈0.01）。结论：Hp可在感染早期诱导GES-1自噬；RAPA和3-MA可增强或降低自噬发生程度，从而减轻或促进细胞损伤。Objective：To investigate the effects of Helicobacter pylori（Hp） on autophagy and apoptosis of gastric epithelial cells. Methods：Human gastric epithelial cell line GES-1（GES-1） were infected by Hp in vitro culture,the intervention groups of Hp-infected GES-1 were treated with autophagy inducer-rapamycin（RAPA） and autophagy inhibitor-3-methyladenine（3-MA）,and uninfected cell was set as the blank control group. Cells were collected at 2,4,6,12 and 24 h after infection. The effects of Hp infection on autophagy and apoptosis of GES-1 were observed by Western Blotting,fluorescent staining,transmission electron microscopy,flow cytometry and intracellular bacterial survival counting. Results：Autophagy marker protein microtubule associated protein 1 light chain 3-Ⅱexpression and cellular autophagy fluorescent particles were found at 2 h after treatment,the typical autophagic vacuoles with double or multilayer membrane in the cytoplasm were observed by transmission electron microscopy. Autophagy of infected cells peaked at 4 h, gradually decreased and disappeared. Autophagy of RAPA-treated cells infected by Hp further enhanced,while the apoptotic rate and intracellular bacterial survival were lower than those in RAPA-untreated infected cells（P〈0. 05 to P〈0. 01）. The autophagy of 3-MA-treated cells infected by Hp attenuated,and apoptosis and bacterial survival in this group were higher than those in RAPA-treated and -untreated infected cells（P〈0. 01）. Conclusions：Hp can induce the autophagy in gastric epithelial cells in the early infection. The autophagy inducer-RAPA and autophagy inhibitor-3-MA can promote or reduce the autophagy in Hp-infected cells,which can lessen or enhance injury of infected cells.

关 键 词：幽门螺杆菌 自噬 细胞凋亡 雷帕霉素 3-甲基腺嘌呤 

分 类 号：R378.99[医药卫生—病原生物学]