The acquisition of Helicobacter pylori and its clinical sequelae of peptic ulcer disease and gastric cancer remain substantial health concerns for our Veterans. H. pylori infects half of the world's population, causes peptic ulcers in 10% and gastric cancer in 1% of those infected, and it is the second leading cause of cancer death worldwide. American Servicemen/women are exposed to H. pylori in regions of the world where infection rates are very high and strains associated with high cancer risk are prevalent. Thus, improved understanding of the defective immune response is crtical. We have gained new insights into why the sustained immune response to H. pylori fails to eradicate the organism. Our work has focused on the role of L-arginine (L-Arg) in the macrophage response to H. pylori. L-Arg is the common substrate for two divergent enzymatic pathways: inducible nitric oxide (NO) synthase (iNOS), that generates high output NO;and arginase, consisting of two forms, arginase I (Arg1) and arginase II (Arg2) that generate L-ornithine, which is the substrate for polyamine synthesis by ornithine decarboxylase (ODC). We have shown that the regulator of L-Arg uptake in macrophages, cationic amino acid transporter-2 (CAT2), is required for protein expression of iNOS and that its function is inhibited by polyamines in a process that restricts NO generation and leads to sustained bacterial colonization and inflammation. In addition, we found that Arg2 is at the center of the ineffective response to H. pylori. In the current paradigm, M1 macrophages express iNOS and produce pro-inflammatory mediators involved in host defense against extracellular infections, while M2 cells express Arg1, are involved in host response to parasitic infection, and are implicated as tumor-associated macrophages. However, in response to H. pylori, macrophages are not M1 or M2 type. They express iNOS and Arg2, but not Arg1. Upon inhibition or knockdown of Arg2, or in cells from Arg2-/- mice, there is enhanced iNOS protein translation/expression and NO generation, indicating that H. pylori-stimulated cells have sub-optimal M1 response. Arg2-/- mice exhibit increased immune responses to H. pylori infection that are correlated with decreased colonization, and we have linked enhanced host defense to more gastric macrophages (GMacs), which undergo less apoptosis and have more iNOS/NO production, and an enhanced Th1/Th17 response. We now show that a substantial portion of GMacs in the H. pylori-infected stomach are regulatory macrophages (Mregs), which may contribute to persistence of the infection. We hypothesize that the competition between iNOS and Arg2 for L-Arg in GMacs leads to a futile cycle and a compromise of host defense that results in H. pylori persistence, inflammation, and cancer risk, and is a site for therapeutic intervention.
Our specific aims are: 1 To determine if immunosuppressive effects of Arg2 are due to inhibition of iNOS. We will generate Arg2-/-iNOS-/- double knockout mice that will be compared to wild-type (WT), Arg2-/-, and iNOS-/- mice, and bone marrow chimeras will be used, to examine A) H. pylori colonization and gastritis;B) GMac phenotype and function;and C) Adaptive immunity in gastric tissues, splenocytes, and lymphocytes. 2.) To determine if effects of Arg2 are mediated by downstream polyamines. We will generate Arg2-/-ODC+/- mice, and administer DFMO (ODC inhibitor) to WT and Arg2-/- mice, and assess: A) H. pylori colonization and gastritis;B) GMac phenotype and function;and C) Adaptive immunity. 3) To determine if Arg2 facilitates H. pylori-induced gastric carcinogenesis, is a target for intervention, and is a biomarker for identifying gastric cancer ris in Veterans. We will utilize: A) Mice that develop dysplasia/cancer, crossed with Arg2-/- mice. B) Gerbils that develop dysplasia/cancer, treated with an arginase inhibitor, BEC. C) Gastric tissues and H. pylori isolates from human subjects in two biorepositories: from areas of low and high gastric cancer risk in Colombia, and from Veterans at the Nashville VAMC. These studies will have a major impact on our understanding of H. pylori immunopathogenesis and gastric carcinogenesis, and will lead to new approaches for risk assessment and treatment.

Public Health Relevance

The gastric bacterial pathogen Helicobacter pylori infects half of the world's population. It causes peptic ulcer disease in 10% and gastric carcinoma in 1% of those infected. Gastric cancer is the second leading cause of cancer-related deaths worldwide. American servicemen/women are frequently exposed to H. pylori at sites of duty where prevalence of this infection is high, and they are at risk of acquiring virulent strains that are harder to treat and associated with carcinogenesis. Our studies will investigate mechanisms by which H. pylori survives in the stomach and incites a futile inflammatory response that leads to ongoing tissue damage. We will focus on how alterations in arginine metabolism cause macrophages to be unable to generate an antimicrobial response that is needed to kill H. pylori, and instead contribute to immunologic tolerance to H. pylori. Our investigations will lead to new understanding of innate and adaptive immunity, strategies for identifying biomarkers for gastric cancer risk, and future therapies involving inhibition of the arginase pathway.

Agency
National Institute of Health (NIH)
Institute
Veterans Affairs (VA)
Type
Non-HHS Research Projects (I01)
Project #
5I01BX001453-02
Application #
8528323
Study Section
Infectious Diseases B (INFB)
Project Start
2012-04-01
Project End
2016-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
2
Fiscal Year
2013
Total Cost
Indirect Cost
Name
Veterans Health Administration
Department
Type
DUNS #
156385783
City
Nashville
State
TN
Country
United States
Zip Code
37212