H. pylori induced DNA Damage and Immune Dysregulation

Vanderbilt University, Nashville TN

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Abstract

DESCRIPTION (provided by applicant): H. pylori remains a major public health problem, since half of the world's population is infected and the associated gastric cancer is the second leading cause of cancer deaths. We have elucidated mechanisms of immune system failure that result in persistence of the organism and sustained oxidative stress that leads to DNA damage. H. pylori induces ornithine decarboxylase (ODC), the rate limiting enzyme for polyamine synthesis, in macrophages. In both in vitro and in vivo models, ODC-derived spermine synthesis results in 1.) apoptosis; and 2.) inhibition of nitric oxide (NO) synthesis, by inhibition of L-arginine (L-Arg)-dependent inducible nitric oxide (NO) synthase (iNOS) protein translation. Inhibition of ODC with oral administration of a- difluoromethylornithine (DFMO) in mice restores L-Arg uptake, macrophage NO production, and cytokine balance, resulting in reduction in both gastritis and H. pylori colonization. H. pylori also up-regulates the enzyme spermine oxidase (SMO), thus generating H2O2, and acting as a novel mechanism for the generation of oxidative stress-associated DNA damage in gastric epithelial cells. We have also discovered another target of H. pylori, heme oxygenase (HO)-1, known for its anti-inflammatory and anti-oxidant properties. We show that H. pylori blocks NO-induced HO-1 in gastric epithelial cells, and this results in unrestrained chemokine synthesis. We hypothesize that H. pylori infection results in failure of antimicrobial and immunoregulatory NO generation and overproduction of polyamines and H2O2, leading to dysregulation of immune responses and survival of gastric epithelial cells with DNA damage. These events lead to persistence of the bacterium, chronic inflammation, and risk for neoplastic transformation. Our Specific Aims are as follows. 1.) To determine the mechanism of persistence of H. pylori colonization and gastritis due to ODC. We will compare WT and ODC+/-, myeloid-specific ODC-/-, iNOS-/- mice, and appropriate bone marrow chimeras, each ¿ DFMO, and assess: A.) gastritis and colonization; B.) immune responses; C.) DNA damage and carcinogenesis by comparing WT and ODC mutant x INS-GAS mice, each ¿ DFMO. 2.) To determine the role of SMO in H. pylori infection, gastritis, and carcinogenesis. We will use SMO-/- mice and assess: A.) colonization, gastritis, advanced lesions, and effects on iNOS; B.) gastric epithelial cell apoptosis and DNA damage; C.) role in carcinogenesis by comparing WT and SMO-/- INS-GAS mice. 3.) To determine the role of HO-1 as a regulator of H. pylori immunopathogenesis and carcinogenesis in vivo by assessing: A.) modulation of HO-1 by polyamines and NO in ODC, SMO, and iNOS KO mice; B.) in mice with normal versus increased NO production and HO-1 expression treated with a pharmacologic inhibitor of HO-1, or in HO-1-/- and epithelial-specific HO-1-/- mice, effects on colonization, gastritis, immune responses, apoptosis, DNA damage, and carcinogenesis. The studies proposed seek new strategies for amelioration or prevention of H. pylori-induced disease by clarifying targets for intervention in the pathogenic immune response that lead to disease persistence and cancer risk.