Mechanisms of H pylori-Induced Hypochlorhydria

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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.