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Mechanisms of H pylori-Induced Hypochlorhydria

幽门螺杆菌引起胃酸过少的机制

基本信息

批准号：
7367046
负责人：
ADAM J SMOLKA
金额：
$ 24.58万
依托单位：
MEDICAL UNIVERSITY OF SOUTH CAROLINA
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-03-15 至 2010-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7367046
关键词：
5&apos Flanking Region Acids Acute Acute Clinical Course Agonist Ammonium Anatomic Sites Antral Archives Atrophic Gastritis Biological Assay Biopsy Body of uterus Cell Count Cells Chimera organism Chronic Classification Clinical DNA-Protein Interaction Data Deletion Mutation Down-Regulation Dysplasia Epithelial Cells Functional disorder Gastric Adenocarcinoma Gastric Parietal Cells Gastric mucosa Gastrins Gastritis Gel Gene Expression Generations Genes Genetic Transcription Genotype Gland Goals H(+)-K(+)-Exchanging ATPase Helicobacter Helicobacter Infections Helicobacter pylori Human Hypochlorhydria Immunochemistry Immunohistochemistry In Vitro Infection Inflammatory Response Ingestion Intestinal Metaplasia Ions Measures Messenger RNA Modeling Molecular Patients Phase Proteins Proton Pump Pylorus Range Regulation Regulatory Element Reporter Reverse Transcriptase Polymerase Chain Reaction Solid Stomach Techniques Testing Time Transcriptional Regulation Transfection Translations Urease Virulence base deletion analysis innovation mutant promoter response transcription factor

项目摘要

DESCRIPTION (provided by applicant): The goal of this study is to define the molecular mechanisms underlying the hypochlorhydria associated with Helicobacter pylori-induced gastritis. Clinical and in vitro evidence associates gastric corpus H. pylori infection with acid hyposecretion and generation of an inflammatory response. In a significant subset of patients, the ensuing gastritis progresses to atrophic gastritis, intestinal metaplasia, dysplasia, and eventually gastric adenocarcinoma. This study tests the hypothesis that acute H. pylori infection down-regulates H,K-ATPase (proton pump) gene expression by perturbing normal interactions of cis-regulatory elements on the HKalpha gene promoter and cellular trans-activating proteins, thereby inhibiting parietal cell acid secretion. In preliminary studies, exogenous H,K-ATPase alpha subunit (HKalpha) promoter sequences transiently transfected into human gastric adenocarcinoma (AGS) cells were responsive to acid secretory agonists and antagonists, and were inhibited by H. pylori infection. This approach forms the experimental basis for three Specific Aims: 1) To identify in gastric epithelial cells H. pylori-responsive c/s-regulatory elements and cognate transcription factors involved in regulation of HK( gene transcription; 2) To investigate the mechanisms whereby specific H. pylori genotypes induce down-regulation of HKalpha gene transcription; and 3) To investigate HKalpha subunit gene transcription and translation within H. pylori.infected and uninfected human gastric mucosa. Basal and H. pylori-responsive cis-regulatory elements and transactivating proteins in transfected ACTS cells will be investigated by deletion analysis, electrophoretic mobility shift, supershift, and solid-phase protein/DNA interaction assays. Acid-inhibitory H. pylori genotypes will be identified by assessing HKa promoter activity in transfected ACTS cells infected with H. pylori mutant strains deficient in virulence-associated genes. Finally, gastric H,K-ATPase mRNA levels and proton pump expression, as measured by real-time RT-PCR and quantitative immunochemistry in a large, well documented archive of human gastric biopsies, will be investigated in the context of intragastric pH, infection status, H. pylori strain identity, and anatomic site of infection. These studies will establish the mechanistic basis of H. pylori-induced gastric hypochlorhydria, and add to the understanding of H. pylori pathophysiology.

描述（由申请人提供）：本研究的目的是确定与幽门螺杆菌诱发的胃炎相关的胃酸过少的分子机制。临床和体外证据表明胃体幽门螺杆菌感染与胃酸分泌不足和炎症反应的产生有关。在相当一部分患者中，随后的胃炎进展为萎缩性胃炎、肠上皮化生、发育不良，并最终发展为胃腺癌。本研究检验了以下假设：急性幽门螺杆菌感染通过干扰 HKalpha 基因启动子上的顺式调节元件和细胞反式激活蛋白的正常相互作用，下调 H,K-ATPase（质子泵）基因表达，从而抑制壁细胞酸分泌。初步研究中，外源 H,K-ATPase α 亚基 (HKα) 启动子序列瞬时转染人胃腺癌细胞（AGS）对酸分泌激动剂和拮抗剂有反应，并受到幽门螺杆菌感染的抑制。该方法构成了三个具体目标的实验基础：1) 鉴定胃上皮细胞中幽门螺杆菌反应性顺/反式调节元件和参与HK(基因转录)调节的同源转录因子；2) 研究特定幽门螺杆菌基因型诱导HKα基因转录下调的机制； 3)研究幽门螺杆菌感染和未感染的人胃粘膜中HKα亚基基因的转录和翻译。将通过缺失分析、电泳迁移率变化、超移和固相蛋白/DNA 相互作用测定来研究转染的 ACTS 细胞中的基础和幽门螺杆菌响应顺式调节元件和反式激活蛋白。通过评估感染了缺乏毒力相关基因的幽门螺杆菌突变株的转染ACTS细胞中的HKa启动子活性，可以鉴定酸抑制性幽门螺杆菌基因型。最后，胃 H,K-ATP 酶 mRNA 水平和质子泵表达（通过实时 RT-PCR 和定量免疫化学在大量记录良好的人类胃活检档案中测量）将在胃内 pH 值、感染状态、幽门螺杆菌菌株身份和感染解剖部位的背景下进行研究。这些研究将建立幽门螺杆菌引起的胃酸过少的机制基础，并增加对幽门螺杆菌病理生理学的理解。