NOD2 induces autophagy to control AIEC bacteria infectiveness in intestinal epithelial cells

Abstract

Objective: The importance of autophagy in mechanisms underlying inflammation has been highlighted. Downstream effects of the bacterial sensor NOD2 include autophagy induction. Recently, a relationship between defects in autophagy and adherent/invasive Escherichia coli (AIEC) persistence has emerged. The present study aims at investigating the interplay between autophagy, NOD2 and AIEC bacteria and assessing the expression level of autophagic proteins in intestinal biopsies of pediatric patients with inflammatory bowel disease (IBD).

Methods: A human epithelial colorectal adenocarcinoma (Caco2) cell line stably over-expressing NOD2 was produced (Caco2NOD2). ATG16L1, LC3 and NOD2 levels were analysed in the Caco2 cell line and Caco2NOD2 after exposure to AIEC strains, by western blot and immunofluorescence. AIEC survival inside cells and TNFα, IL-8 and IL-1βmRNA expression were analysed by gentamicin protection assay and real time PCR. ATG16L1 and LC3 expression was analyzed in the inflamed ileum and colon of 28 patients with Crohn's disease (CD), 14 with ulcerative colitis (UC) and 23 controls by western blot.

Results: AIEC infection increased ATG16L1 and LC3 in Caco2 cells. Exposure to AIEC strains increased LC3 and ATG16L1 in Caco2 overexpressing NOD2, more than in Caco2 wild type, while a decrease of AIEC survival rate and cytokine expression was observed in the same cell line. LC3 expression was increased in the inflamed colon of CD and UC children.

Conclusions: The NOD2-mediated autophagy induction is crucial to hold the intramucosal bacterial burden, especially towards AIEC, and to limit the resulting inflammatory response. Autophagy is active in inflamed colonic tissues of IBD pediatric patients.

Keywords: AIEC; Autophagy; Inflammation; Inflammatory bowel disease; NOD2.