Mechanisms of H pylori-Induced Hypochlorhydria

幽门螺杆菌引起胃酸过少的机制

• 批准号: 6870783
• 负责人: ADAM J SMOLKA
• 金额: $ 25.99万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-03-15 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6870783
• 关键词: Helicobacter; achlorhydria; gastric acid; gastric mucosa; gastritis; gastrointestinal infection; gel mobility shift assay; gene deletion mutation; gene expression; genetic promoter element; genetic regulatory element; genetic strain; genetic transcription; genetic translation; human tissue; hydrogen potassium exchanging ATPase; hydrogen transport; immunocytochemistry; neoplastic cell culture for noncancer research; pathologic process; polymerase chain reaction; protein protein interaction; tissue /cell culture; transcription factor; transfection; virulence

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导的胃炎相关的低氯血症的分子机制。临床和体外证据表明胃体H。幽门螺杆菌感染伴酸分泌不足和炎症反应的产生。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、异型增生，最终发展为胃腺癌。本研究验证了急性H。幽门螺杆菌感染通过扰乱HK α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而下调H，K-ATP酶（质子泵）基因表达，从而抑制壁细胞酸分泌。在初步研究中，外源性H，K-ATP酶α亚基（HK α）启动子序列瞬时转染人胃粘膜， 腺癌（AGS）细胞对酸分泌激动剂和拮抗剂有反应，H.幽门感染该方法形成了三个具体目的的实验基础：1）在胃上皮细胞中鉴定H。pylori反应性c/s调控元件和相关转录因子参与HK基因转录调控; pylori基因型诱导HK α基因转录下调; 3）研究HK α亚基基因在H.幽门螺杆菌感染和未感染的人胃粘膜。Basal和H.将通过缺失分析、电泳迁移率变动、超变动和固相蛋白/DNA相互作用测定来研究转染的pylori细胞中的幽门应答顺式调节元件和反式激活蛋白。抑酸H. pylori基因型将通过评估感染H. pylori的转染的HKA细胞中的HKA启动子活性来鉴定。缺乏毒力相关基因的幽门螺杆菌突变株。最后，通过实时RT-PCR和定量免疫化学在大量有据可查的人类胃活检标本中测量胃H，K-ATP酶mRNA水平和质子泵表达，将在胃内pH、感染状态、H.幽门螺杆菌菌株身份和感染的解剖部位。 这些研究将为H. pylori诱导的胃低氯血症，并增加了对H. pylori病理生理学