Intestinal Epithelial PTPN2 Restricts Adherent‐Invasive E. coli Colonization and Promotes Anti‐Microbial Peptide Responses in Mice

Abstract

Background In addition to a physical barrier, the intestinal epithelium generates a chemical barrier to the luminal microbiota by secreting various antimicrobial factors. Alterations in the gut microbiota and expansion of pathobionts such as adherent‐invasive Escherichia coli (AIEC) are associated with inflammatory bowel disease (IBD) pathogenesis. Our previous work showed, mice deficient for the expression of the IBD risk gene, Ptpn2, exhibit pronounced expansion of a novel murine adherent‐invasive Escherichia coli (mAIEC) strain. This study aimed to investigate how PTPN2 expression in intestinal epithelial cells restricts mAIEC colonization in vivo. Methods We generated tamoxifen‐inducible, intestinal epithelial cell‐specific knockout mice (Ptpn2∆IEC). Ptpn2∆IEC and control littermates (Ptpn2fl/fl) were infected with either PBS, non‐invasive E. coli K12, or fluorescent‐tagged mAIEC (mAIECred)for 4 consecutive days. After euthanasia, bacterial colonies were enumerated in mouse tissues. mRNA and protein expression were assayed in intestinal epithelial cells (IECs) by PCR and Western blot. Results Ptpn2∆IEC mice exhibited intestinal region‐specific higher mAIECred ‐ but not K12 ‐ bacterial load in distal colon tissue compared to Ptpn2fl/fl mice (P=0.038; n=9‐12). To identify if the higher susceptibility to mAIECred infection was associated with altered host defenses, we measured levels of anti‐microbial peptide (AMPs) in these mice. Ileal RNA expression of the alpha‐defensin AMPs, Defa5 and Defa6, were significantly lower (P=0.008, 0.0182 respectively; n=3‐5) in Ptpn2∆IEC vs. Ptpn2fl/fl mice, after mAIECred infection but not K12 infection. Further, expression of the protease matrilysin‐7 (MMP7) ‐ which is responsible for the proteolytic cleavage of alpha‐defensins – was significantly decreased in ileum and distal colon of Ptpn2∆IEC mice post mAIECred infection compared to Ptpn2fl/fllittermates (P=0.042, 0.007; n=4‐6). REG3γ, an AMP that acts largely against gram‐positive bacteria, was unchanged in all conditions. However, ileal IEC lysozyme protein expression was lower in PBS and K12, but not mAIECred, infected Ptpn2∆IECmice (P= 0.067, 0.0165 respectively; n=8) compared to control Ptpn2fl/fl mice. Conclusion Intestinal epithelial PTPN2 plays a major role in regulating intestinal bacterial defenses and gut regional variations in pathobiont colonization.