幽门螺杆菌通过p38MAPK信号通路上调人胃癌MKN45细胞中COX-2启动子的活性  

Helicobacter pylori infection increases the transcriptional activity of COX-2 promoter via the p38MAPK signal transduction pathway in MKN45 cells

作　　者：刘宣[1,2] 刘宁宁[1,2] 王炎[1,2] 殷佩浩[1,2] 吴琼[1,2] 周宁[1,2] 孙珏[1,2] 范忠泽[1,2] 李琦[1,2] 

机构地区：[1]上海中医药大学附属普陀医院 [2]上海中医药大学中西医结合肿瘤介入研究所,上海市200062

出　　处：《世界华人消化杂志》2010年第28期3003-3007,共5页World Chinese Journal of Digestology

基　　金：国家自然科学基金资助项目;No.81072955;No.30600844;上海市重点学科基金资助项目;No.S30302;上海市普陀区科技创新重大基金资助项目;No.2009-37-2;上海市教委青年基金科研资助项目;No.09JW44~~

摘　　要：目的:探讨H.pylori感染对人胃癌MKN45细胞环氧合酶2(COX-2)启动子荧光素酶报告基因重组质粒(pGL3-Basic-COX-2-promoter)活性的影响及p38MAPK信号通路对其的调控作用.方法:将构建的COX-2启动子pGL3-Basic-COX-2-promoter和内参质粒pRL-SV40共转染人胃癌MKN45细胞,加入100倍细胞数量的H.pylori共培养一段时间后,检测双荧光素酶的活性.Western blot检测H.pylori感染对人胃癌MKN45细胞p38MAPK信号通路的激活作用.运用p38MAPK特异性抑制剂SB203580阻断p38MAPK信号通路,观察H.pylori对MKN45细胞COX-2启动子活性的影响.结果:瞬时转染实验显示,COX-2启动子在MKN45细胞中的转录表达随时间的变化而升高,转染后40h的双报告基因活性是转染后8h的3.5倍(P<0.01);加入H.pylori共培养后,同时间点的荧光素酶活性明显升高(P<0.05或P<0.01),转染后40h的活性是转染后8h的5倍(P<0.01).H.pylori感染20min后,p38MAPK信号通路被激活,60min达峰值;加入p38MAPK信号通路特异性抑制剂SB203580抑制p38MAPK信号通路后,COX-2启动子的活性均明显下调.结论:H.pylori感染通过p38MAPK信号通路上调COX-2启动子活性,这可能是H.pylori促进胃癌发生发展的重要因素之一.AIM：To investigate the effect of Helicobacter pylori （H.pylori） infection on the transcriptional activity of cyclooxygenase-2 （COX-2） promoter in gastric cancer MKN45 cells and to explore potential mechanisms involved.METHODS：The recombinant vector pGL3Basic-COX-2-promoter and a control vector （pRL-SV40） was transiently co-transfected into MKN45 cells.Twelve hours later,the transfected cells were infected with H.pylori （100 times the number of cells） for different durations.The COX-2 promoter activity was detected using the dual luciferase assay.The activation of p38MAPK pathway was evaluated by Western blot.The p38MAPK signal transduction pathway was then blocked with a specific inhibitor SB203580 to detect the effect of H.pylori infection on COX-2 promoter activity.RESULTS：After transient transfection,the activity of COX-2 promoter in MKN45 cells was increased with time.At 40 h after transfection,the activity of dual-luciferase was 3.5 folds higher than that at 8 h （P0.05）.H.pylori infection significantly increased the activity of dualluciferase compared with control cells （all P0.05 or 0.01）.The activity of dual-luciferase in cells infected with H.pylori at 40 h after transfection was 5 folds higher than that at 8 h （P0.01）.The expression of p38MAPK was up-regulated at 20 min after infection and reached the peak at 60 min.Blockage of the p38MAPK signal transduction pathway with SB203580 significantly reduced the COX-2 promoter activity.CONCLUSION：H.pylori infection increases the transcriptional activity of COX-2 promoter via the p38MAPK signal transduction pathway,which may be one of the important mechanisms by which H.pylori infection causes gastric cancer.

关 键 词：胃癌 幽门螺杆菌 环氧合酶2 启动子 P38丝裂原活化蛋白激酶 信号转导 

分 类 号：R735.2[医药卫生—肿瘤]