CAJ | 학술논문

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。
목적：탐토유문라간균(Hp)대위점막상피세포(GES-1)자서화조망적영향。방법：Hp여GES-1체외공배양，동시설립뢰파매소(RAPA)、3-갑기선표령(3-MA)간예조급공백대조조，우2、4、6、12급24 h종지감염，통과면역단백인적、형광염색、투사전경、류식세포술급세포내활균계수등방법관찰Hp대GES-1자서화조망적영향。결과：Hp+GES-1조재2 h출현자서표지단백LC3-Ⅱ표체급세포내형광자서과립，투사전경관찰도세포내출현쌍층혹다층막포과적자서소체양결구，세포자서재4 h체고봉，감염중후기축점감약병소실；RAPA+Hp+GES-1조세포자서정도진일보증강，세포조망솔급세포내활균수균강저(P<0.05~P<0.01)；3-MA+Hp+GES-1조세포자서강도현저강저，이기세포조망솔화세포내활균수균명현승고(P <0.01)。결론：Hp가재감염조기유도GES-1자서；RAPA화3-MA가증강혹강저자서발생정도，종이감경혹촉진세포손상。
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.