OXIDATIVE INJURY TO GASTRIC EPITHELIAL CELLS BY H. PYLORI

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DESCRIPTION (provided by applicant): The long-term goal of our work is to gain insight into mechanisms by which oxidative stress responses to H. pylori infection and other injury to the gastric epithelium lead to human disease including peptic ulcer disease and gastric carcinoma. In response to infection, phagocytes recruited to the gastric mucosa, become activated and generate reactive oxygen species (ROS) and H. pylori infection induces oxidative stress in gastric epithelial cells directly through the generation of ROS. This project has focused on ROS-induced activation of apurinic/apyrimidinic endonuclease (APE1), a multifunctional protein that is the rate-limiting enzyme in DNA base excision repair of oxidative lesions. It is also known as redox factor (Ref)-1 due to its ability to control gene expression by reductively activating transcription factors including activator protein (AP)-1, NF-¿B, and p53. Our recent preliminary studies build on the understanding that ROS arises from activation of NADPH oxidase during H. pylori infection. During the course of preliminary studies, we observed that APE1 is not only activated by ROS, but its induction provides a negative feedback on the accumulation of ROS. Data suggest that this feedback is based on novel molecular interactions between APE1 and Rac1. Rac1 is an important regulator of cell function, implicated in the control of bacterial infections and the pathogenesis of chronic inflammatory diseases including IBD. One of its functions is to activate NADPH oxidase which leads to the accumulation of ROS. The hypothesis that APE1 modulates gastric epithelial cell responses by regulating the accumulation of ROS will be tested in the following Specific Aims: Aim 1: Define the role of APE1 in ROS accumulation in epithelial cells during H. pylori infection. Aim 2: Determine the mechanisms whereby APE1 regulates Rac1 activity. Aim 3: Determine the mechanisms whereby APE1 regulates NAPH oxidase. Despite the advances in our understanding of the pathogenesis of H. pylori, the mechanisms by which this infection leads to epithelial cell injury and disease such as malignancy remain poorly understood. The broad objective for the proposed studies is to define a novel molecular mechanism whereby APE1 regulates Rac activation and the generation of ROS through NADPH oxidase. Using H. pylori as a model system, these studies will provide new knowledge of the control of oxidative stress. By the end of 5 years, the expected outcomes and milestones include a definition of the molecular basis for the regulation of ROS accumulation by APE1 including its interaction with Rac1 and NADPH oxidase. This information will have an important positive impact by advancing our understanding of the molecular mechanisms regulating Rac activation that is relevant to diseases in the human GI tract.