幽门螺杆菌刺激胃癌细胞株SGC-7901分泌白细胞介素-8过程中核因子κB的作用

目的　探讨胃上皮细胞在幽门螺杆菌 (Hp)刺激下分泌白细胞介素 8(IL 8)的过程中 ,核因子 κB(NF κB)的作用。方法　电穿孔法将I κBαM(NF κB抑制蛋白 )基因转染入SGC 790 1中 ,β半乳糖苷酶分析转染效率 ,Western印迹分析I κB基因的表达。将不同浓度的Hp活菌、灭活菌及液体培养上清分别与稳定转染I κB的胃癌细胞株SGC 790 1及其空白质粒对照共同孵育 ,凝胶电泳迁移率变迁分析 (EMSA)和NF κB荧光素酶报告基因分析检测不同时间细胞内NF κB的激活 ,ELISA法检测不同时间细胞上清中IL 8的水平。结果　得到了稳定表达I κB的SGC 790 1细胞 ,命名为SGC790 1 I κB细胞 ,其空白质粒对照命名为SGC790 1 neo。Hp活菌刺激的SGC790 1 neo在 2h和 4h时可检测到明显的NF κB激活 ,灭活菌和液体培养上清刺激下无激活 ;Hp活菌、灭活菌和培养上清刺激下SGC 790 1 I κB内均无NF κB激活。 4h时 ,2× 10 7Hp/ml的Hp活菌刺激下即可检测到SGC790 1 neo内明显的IL 8分泌 ,并有时间 剂量依赖效应 ;灭活菌和液体培养上清无此作用。Hp活菌、灭活菌和液体培养上清均不能刺激SGC790 1 I κB产生IL 8。结论　Hp诱导胃上皮细胞分泌IL 8依赖于NF κB的激活。

The role of nuclear factor kappa B in secretion of interleukin-8 by gastric cancer cell line SGC 7901 induced by Helicobacter pyiori

OBJECTIVE: To investigate the role of nuclear factor kappaB (NF-kappaB) in the gastric inflammation induced by Helicobacter pylori (H. pylori). METHODS: SGC-7901 was transfected with IkappaB (NF-kappaB inhibitor gene) by electroporation, (beta)lacZ activity assay was used to examine transfected efficacy. Expression of IkappaB was assessed by Western-blot. Different concentration of live and heat-killed Hp (ATCC 43504) and supernatant of liquid culture were cocultured with SGC7901-IkappaB and its negative control SGC7901- neo. Activation of intracellular NF-kappaB was examined by electrophoretic mobility shift analyses (EMSA) and luciferase report gene assay at different time point. IL-8 levels were measured by ELISA at different time point. RESULTS: IL-8 release was evident 4 hours after infection of SGC-7901-neo with H. pylori, and this effect was dose-time dependent. SGC-7901-IkappaB in which NF-kappaB has not been activated could not secret IL-8 after infection with H. pylori. CONCLUSION: Secretion of IL-8 by gastric epithelial cell upon H pylori infection is dependent on activation of NF-kappaB.