幽门螺杆菌脂多糖作用于胃癌细胞后刺猬蛋白信号通路中相关蛋白Ptch-1、Gli的表达及意义  

Effect of Helicobacter pylori lipopolysaccharide on expression of Ptch-1 and Gli proteins in gastric cancer AGS cells

作　　者：周慧[1] 张军[1] 张国新[2] 邰婷[1] 周晓颖[2] 苏静[2] 

机构地区：[1]南京医科大学附属南京医院南京市第一医院消化科,江苏省南京市210006 [2]南京医科大学第一附属医院消化科,江苏省南京市210029

出　　处：《世界华人消化杂志》2014年第24期3557-3564,共8页World Chinese Journal of Digestology

摘　　要：目的:通过用幽门螺杆菌(Helicobacter pylori,H.pylori)脂多糖(lipopolysaccharide,LPS)、环巴胺(cyclopamine)分别或联合作用于胃癌细胞AGS后,观察刺猬蛋白(sonic hedgehog,Shh)信号通路中相关蛋白Ptch-1、Gli的表达变化,并探讨其意义.方法:(1)不同浓度H.pylori LPS作用于AGS细胞24 h,Western blot检测Ptch-1、Gli的表达,找到最佳刺激浓度.以最佳刺激浓度分别作用于AGS细胞24、48、72 h,Western blot法检测Ptch-1、Gli的表达;(2)不同浓度环巴胺作用于AGS细胞24 h后,MTT法检测对AGS的抑制率,求得半数抑制浓度(50%concentration of inhibition,IC50).以环巴胺IC50作用AGS细胞24、48、72 h后,Western blot检测Ptch-1、Gli的表达;(3)AGS细胞培养后分3组,分别用H.pylori LPS、环巴胺、H.pylori LPS+环巴胺作用AGS细胞24 h,另设阴性对照组,应用Western blot法对各组细胞中Shh信号通路相关蛋白Ptch-1、Gli的表达水平进行检测.结果:不同浓度H.pylori LPS作用于AGS细胞24 h后Ptch-1、Gli的表达升高,且随着浓度增加,表达逐渐增高(P<0.05),最终达到一个平台期,以平台期浓度作为最佳刺激浓度,分别刺激AGS细胞24、48、72 h,Western blot结果显示,Ptch-1、Gli的表达逐渐升高,呈时间依赖性(P<0.05);不同浓度环巴胺作用于AGS细胞24 h后,MTT结果显示环巴胺对AGS的抑制作用呈浓度依赖性(P<0.05);以环巴胺IC50作用AGS细胞24 h后,Western blot结果显示随着时间的延长,Gli的表达逐渐减低,而Ptch-1的表达无显著性差异.H.pylori LPS、环巴胺、H.pylori LPS+环巴胺组分别作用于AGS细胞24 h后,H.pylori LPS+环巴胺联合作用组的Ptch-1表达高于对照组及环巴胺组(P<0.05),而与H.pylori LPS组无统计学差异;H.pylori LPS+环巴胺联合作用组Gli的表达低于对照组和H.pylori LPS组(P<0.05),与环巴胺组无统计学差异.结论:H.pylori通过其毒力因子LPS能够影响胃癌细胞AGS的Shh信号通路相关蛋白的表达.环巴胺可通过抑制Shh信号通路抑�AIM： To explore the expression of Ptch-1 andGli, two members in the sonic hedgehog（Shh）signaling pathway, in gastric cancer AGS cellsafter stimulation with Helicobacter pylori（H. py-lori） lipopolysaccharide（LPS） and cyclopamine.METHODS： Different concentrations of H. py-lori LPS were used to stimulate AGS cells for24 h, and the expression of Ptch-1 and Gli wasassayed by Western blot to find the optimalstimulation concentration. AGS cells were thentreated with H. pylori LPS at the optimal concen-tration for 24, 48 and 72 h, and the expression of Ptch-1 and Gli was detected by Western blot. MTT assay was applied to explore the effect of cyclopamine on AGS cell proliferation, and the half maximal inhibitory concentration（IC50） was calculated. AGS cells were then treated with cyclopamine at IC50 for 24, 48 and 72 h, and the expression of Ptch-1 and Gli was detected by Western blot. AGS cells were divided into three groups and stimulated with H. pylori LPS, cyclo-pamine and H. pylori-LPS plus cyclopamine for 24 h, respectively, and the expression of Ptch-1 and Gli was then detected by Western blot.RESULTS： After different concentrations of H. pylori LPS were used to stimulate AGS cells for 24 h, the expression of Ptch-1 and Gli was increased（P〈0.05） and eventually reached a plateau. After treatment of AGS cells with H. pylori LPS at the optimal concentration for 24, 48 and 72 h, the expression of Ptch-1 and Gli was increased in a time dependent manner（P〈0.05）. After treatment of AGS cells with different concentrations of cyclopamine for 24 h, the pro-liferation of AGS cells was inhibited in a concen-tration dependent manner（P〈0.05）. Treatment with cyclopamine at IC50 decreased the expres-sion of Gli in a time dependent manner（P〈0.05）, but showed no significant effect on expression of Ptch-1（P 〉0.05）. The expression of Ptch-1 in the H. pylori LPS plus cyclopamine group was sig-nificantly higher than that in the control group and cyclopamine group（P〈0.05）, but wa

关 键 词：幽门螺杆菌 脂多糖 胃癌细胞 刺猬蛋白 环巴胺 

分 类 号：R735.2[医药卫生—肿瘤]