Mechanisms of gut epithelial DUOX-mediated intestinal homeostasis

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PROJECT SUMMARY/ABSTRACT A monolayer of gut epithelial cells covers the intestinal lumen preventing an overt immune response against the normal gut microbiota, while at the same time controlling infection with potentially life-threatening pathogens. Breakdown of this homeostatic relationship can lead to foodborne infections and chronic intestinal inflammation. Reactive oxidative species (ROS) play a critical role in mucosal host defense but also contribute to the inflammatory milieu. The protective host defense role of the classical NOX2/gp91phox NADPH oxidase, responsible for the oxidative burst reaction of macrophages, has been extensively researched, but the role of another NADPH oxidase, dual oxidase (DUOX) is less clear. Our long-term objective is to define the role of DUOX2, the only form of DUOX expressed in the gut epithelium, in epithelial defense to maintain mucosal homeostasis. The objective here is to determine how DUOX2 can protect epithelial cells from invasive bacteria that produce antioxidative enzymes to shield themselves from H2O2. Our hypothesis is that activation of DUOX2 by bacterial attachment is critical for proper handling of engulfed bacteria to maintain microbe- host homeostasis. The rationale for the proposed research is that, once it is known how DUOX2 orchestrates mucosal host defense in the intestinal epithelium, specific pharmacotherapeutics can be selected to enhance DUOX-mediated bacterial handling, resulting in new approaches to the prevention and treatment of a variety of GI disorders associated with increased bacterial translocation. We will test our central hypothesis and, thereby, accomplish the objective of this application by pursuing the following specific aims: 1. Investigate the mechanisms of DUOX2 induction by mucosa-associated bacteria, 2. Define the role of DUOX2 in the cell-autonomous containment of intestinal pathogens. 3. Study the implications of DUOX2 deficiency for susceptibility to IBD. The expected outcomes, the proposed work is expected to elucidate a mechanistic framework how DUOX2 is activated in response to bacteria-epithelial contact, supports cell- autonomous inactivation of intracellular bacteria, and, thereby, maintains immune homeostasis. Such results are expected to have an important positive impact because this immune mechanism is highly likely to provide new targets for preventative and therapeutic interventions for diseases associated with dysregulated microbe- intestinal interaction (e.g., IBD, IBS, colon cancer) in addition to fundamentally advancing the fields of gut mucosal immunity. It will also provide much-needed insight into the pathogenesis of IBD linked to DUOX2 LOF and thus the foundation to better strategize a treatment plan for these patients.