CAJ | 학술논문

目的：观察幽门螺杆菌感染人胃上皮细胞后自噬的发生及变化情况，并探索上调幽门螺杆菌感染后人胃上皮细胞GES-1自噬反应对细胞是否具有一定的保护作用，为进一步研究幽门螺杆菌感染与细胞自噬奠定基础。方法使用一定浓度的幽门螺杆菌感染人胃上皮细胞GES-1，通过Western blot检测感染不同时间点GES-1中微管相关蛋白轻链3Ⅰ／Ⅱ（ LC3Ⅰ／Ⅱ）的转换与Beclin 1的表达，免疫荧光染色观察自噬体的出现。确定Xestospongin B处理人胃上皮细胞GES-1的浓度后，将培养细胞分为三组：幽门螺杆菌感染组、幽门螺杆菌感染＋Xestospongin B组、正常培养组。首先确定诱导剂对感染后自噬的上调作用，然后使用微毛细管式细胞分析仪检测细胞凋亡和活性，比较三组细胞存活以及凋亡的差异。结果在GES-1细胞被感染后2 h即可观察到自噬的发生，且呈时间依赖性，于8 h到达高峰，24 h开始恢复基础水平。加诱导后细胞自噬水平上调。单纯感染组细胞活性先迅速降低，2 h之后细胞活性平稳降低，凋亡率逐渐增加。加诱导剂组细胞活性在感染后的6 h内细胞活性稍有降低，6 h后细胞活性开始增加，凋亡率降低。结论幽门螺杆菌能够诱导人胃上皮GES-1发生自噬，Xestospongin B能够上调幽门螺杆菌感染后人胃上皮细胞GES-1的自噬水平，并且上调自噬反应能够对幽门螺杆菌感染的人胃上皮细胞GES-1起到一定保护作用。
목적：관찰유문라간균감염인위상피세포후자서적발생급변화정황，병탐색상조유문라간균감염후인위상피세포GES-1자서반응대세포시부구유일정적보호작용，위진일보연구유문라간균감염여세포자서전정기출。방법사용일정농도적유문라간균감염인위상피세포GES-1，통과Western blot검측감염불동시간점GES-1중미관상관단백경련3Ⅰ／Ⅱ（ LC3Ⅰ／Ⅱ）적전환여Beclin 1적표체，면역형광염색관찰자서체적출현。학정Xestospongin B처리인위상피세포GES-1적농도후，장배양세포분위삼조：유문라간균감염조、유문라간균감염＋Xestospongin B조、정상배양조。수선학정유도제대감염후자서적상조작용，연후사용미모세관식세포분석의검측세포조망화활성，비교삼조세포존활이급조망적차이。결과재GES-1세포피감염후2 h즉가관찰도자서적발생，차정시간의뢰성，우8 h도체고봉，24 h개시회복기출수평。가유도후세포자서수평상조。단순감염조세포활성선신속강저，2 h지후세포활성평은강저，조망솔축점증가。가유도제조세포활성재감염후적6 h내세포활성초유강저，6 h후세포활성개시증가，조망솔강저。결론유문라간균능구유도인위상피GES-1발생자서，Xestospongin B능구상조유문라간균감염후인위상피세포GES-1적자서수평，병차상조자서반응능구대유문라간균감염적인위상피세포GES-1기도일정보호작용。
Objective To observe the autophagy induced by Helicobacter pylori ( H.pylori ) infection,and to investigate the protective role of upregulating autophagy of human gastric epithelium cell GES-1 induced by H.pylori infection, and to lay the foundation for the study on H.pylori infection and autophagy.Methods Infected human gastric epithelium cell GES-1 by a certain concentration of H.pylori. Western blot was utilized for detecting the level of LC3-I/II conversion and the level of Beclin 1, which indicated the autophagy induction.The formations of autophagosomes were observed via immunofluorescence with microscopy.GES-1s were divided into three groups:infection group,infection+Xestospongin B group and control group.Microcapillary cell analyzer ( Millipore Guava easyCyte ) was utilized to measure the cell viability and apoptosis rate,and the differences among the three groups were analyzed.Results At 2 h post infection,autophagy induction became detectable.It reached a peak level at 8 h p.i,while it returned to the basic level at 24 h p.i.Autophagy activities of infected cells were upregulated with induction of Xestospongin B.Cell viability of infection group rapidly decreased firstly,and steadily decreased after 2 hours,and the rate of apotosis was increasing with duration of infection.Cell viability of infection +Xestospongin B group decreased sightly within 6 hours post infection and increased after that untill 24 hours,and the rate of apotosis decreased.Conclusions H.pylori is capable of inducing autophagy in human gastric epithelium cell GES-1, which can be modulated by autophagic regulators Xestospongin B.The upregulated autophagy may play a role in protecting GES-1 from H.pylori infection.