Koike Takero, Shimada Tadahito, Fujii Yoichiro, Chen Guozhong, Tabei Kyoko, Namatame Takashi, Yamagata Michiko, Tajima Akihiro, Yoneda Masashi, Terano Akira, Hiraishi Hideyuki
Department of Gastroenterology and Institute for Medical Science, Dokkyo Medical University, Mibu, Tochigi, Japan.
J Gastroenterol Hepatol. 2007 Jun;22(6):936-42. doi: 10.1111/j.1440-1746.2007.04861.x.
TFF1 (pS2) is expressed at a high level in gastric epithelial cells and plays an important role in protecting the gastric mucosa. However, the regulatory mechanisms of TFF1 expression are not fully understood. The aim of this study was to investigate the effect of TNF-alpha, a representative proinflammatory cytokine, on TFF1 expression.
MKN45 and AGS cells, derived from human gastric carcinoma, were used. Endogenous TFF1 mRNA expression was analyzed by real-time quantitative RT-PCR. The sequences of the human TFF1 promoter were cloned into the pGL3-basic vector and reporter gene assays were performed. Nuclear factor (NF)-kappaB activity was monitored using a reporter vector that contained multiple copies of NF-kappaB responsive element upstream of the luciferase gene. Interaction between NF-kappaB and TFF1 cis-element was examined by electophoretic mobility shift assay (EMSA).
TNF-alpha activated NF-kappaB and up-regulated endogenous TFF1 mRNA expression as well as the transcription of the TFF1 reporter genes in a dose-dependent manner. IL-1beta, another proinflammatory cytokine, also up-regulated TFF1 expression. TNF-alpha responsive element was mapped between -342 and -147 of the human TFF1 promoter and a putative NF-kappaB binding site was identified at -231. When this element was deleted, the reporter genes became almost insensitive to TNF-alpha treatment. EMSA showed binding of NF-kappaB to this element.
Inflammatory stimuli that activate NF-kappaB appear to up-regulate TFF1 expression in gastric epithelial cells. This mechanism may aid in the protection of the gastric mucosa under inflammatory conditions.
三叶因子1（TFF1，即pS2）在胃上皮细胞中高表达，对胃黏膜具有重要的保护作用。然而，TFF1表达的调控机制尚未完全明确。本研究旨在探讨具有代表性的促炎细胞因子肿瘤坏死因子-α（TNF-α）对TFF1表达的影响。
采用源自人胃癌的MKN45和AGS细胞。通过实时定量逆转录聚合酶链反应（RT-PCR）分析内源性TFF1 mRNA表达。将人TFF1启动子序列克隆至pGL3-基本载体中，并进行报告基因检测。使用在荧光素酶基因上游包含多个核因子（NF）-κB反应元件拷贝的报告载体监测NF-κB活性。通过电泳迁移率变动分析（EMSA）检测NF-κB与TFF1顺式元件之间的相互作用。
TNF-α以剂量依赖方式激活NF-κB，上调内源性TFF1 mRNA表达以及TFF1报告基因的转录。另一种促炎细胞因子白细胞介素-1β（IL-1β）也上调TFF1表达。TNF-α反应元件定位于人TFF1启动子的-342至-147之间，在-231处鉴定出一个假定的NF-κB结合位点。当该元件缺失时，报告基因对TNF-α处理几乎不敏感。EMSA显示NF-κB与该元件结合。
激活NF-κB的炎性刺激似乎上调胃上皮细胞中TFF1的表达。该机制可能有助于在炎症条件下保护胃黏膜。
