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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。
Objective To investigate the role of nuclear factor kappa B (NF κB) in the secretion of interleukin 8 (IL 8) from gastric epithelial cells stimulated by Helicobacter pylori (Hp). Methods IκBαM (NF κB inhibitory protein) gene was transfected into SGC7901 by electroporation. The transfection efficiency was analyzed by β-galactosidase, and the expression of IκB gene was analyzed by Western blotting. Different concentrations of live Hp bacteria, inactivated bacteria, and liquid culture supernatants were incubated with IκB-transfected gastric cancer cell line SGC7901 and its blank plasmid control, respectively, and gel electrophoretic mobility shift analysis (EMSA) and NF were performed. The κB luciferase reporter gene assay was used to detect the activation of NF κB in cells at different times. ELISA was used to detect the level of IL-8 in the cell supernatant at different times. Results SGC 790 1 cells stably expressing IκB were obtained and designated as SGC790 1 I κB cells. The blank plasmid control was named as SGC790 1 neo. SGC790 1 neo stimulated by Hp live bacteria can detect obvious NF κB activation at 2h and 4h, and the inactivated bacteria and liquid culture supernatants have no stimulation. Spirulina, live bacteria, and culture supernatants stimulate SGC790. There was no activation of NFκB in 1 IκB. At 4 h, significant IL-8 secretion in SGC790 1 neo could be detected under the stimulation of 2×10 7 Hp/ml Hp viable bacteria, and there was a time-dependent effect; the inactivated bacteria and the liquid culture supernatant had no such effect. Neither live Hp bacteria, inactivated bacteria, nor liquid culture supernatants stimulated IL-8 production by SGC790 1 I κB. Conclusion The secretion of IL-8 by gastric epithelial cells induced by Hp is dependent on the activation of NF-κB.