P-135 YI IL-22 Promotes Helicobacter Hepaticus-Induced H2AX Phosphorylation and Dysbiosis in the Large Bowel

Abstract

Interleukin-22 (IL-22) plays an essential role in the innate immunity to enteric pathogens, at least in part through induction of anti-microbial peptides (AMP). Helicobacter hepaticus (Hh) induces chronic colitis and colon cancer in susceptible mouse strains. Previous studies have described high levels of IL-22 mRNA within the lower bowel of Rag2−/− mice chronically infected with Hh. However, the role of IL-22 in modulating the early innate immune response to Hh infection has not been fully explored. Female and male 129S6/SvEvTac-Rag2tm1Fwa mice were infected with H. hepaticus and the inflammatory response in the lower bowel examined 2 weeks later. Inflammation was evaluated by scoring of H&E stained slides of the cecum and colon, and immune cell populations within the intestine were evaluated using flow cytometry. Pro-inflammatory cytokine expression within the lower bowel was evaluated by quantitative-PCR, and DNA damage within colonic epithelial was assessed by immunostaining for γH2AX, a marker of DNA double-stranded breaks. To evaluate the role of IL-22 in this system, mice were treated with anti-IL-22 antibody following Hh infection. The influence of Hh infection and IL-22-depletion on the colonic microbiome was evaluated by sequencing of bacterial 16S rRNA genes. We found that Hh infection rapidly induced inflammation that was significantly more severe in the cecocolic junction than either the proximal or distal colon, associated with increases in the percentages of CD45+CD11b+CD64+Ly6c+MHCII+ inflammatory monocytes and Ly6G+Ly6c+ neutrophils, as well as increases in absolute numbers of ThyHigh type-3 innate lymphoid cells (ILCs). Infection was associated with increase expression of IL-22 and AMP RegIIIβ and RegIIIγ, and AMP expression was inhibited by treatment with an anti-IL-22 antibody. Depletion of IL-22 did not influence the severity of inflammation within the cecum, although it did mildly reduce inflammation in the colon. Unexpectedly, mice infected with Hh exhibited accumulation of γH2AX+ intestinal crypt epithelial cells suggesting that Hh infection may acutely induce DNA damage, and remarkably γH2AX staining was inhibited by depletion of IL-22. Interestingly, Hh infection induced dysbiosis characterized by reduced species diversity and expansion of the phylum Proteobacteria, which was partially reversed by depletion of IL-22. These results indicated that IL-22 has a limited role in regulating Hh-induced inflammation within the cecum. However, the observation that Hh infection acutely induces γH2AX staining which is inhibited by depletion of IL-22, raises the possibility that IL-22 mediates the accumulation of epithelial cells that contain damaged DNA following Hh infection. These observations could have important implications for understanding of the relationship between inflammation, dysbiosis, and neoplasia within the lower bowel.