A25 INVESTIGATING THE ROLE OF TOLL-INTERACTING PROTEIN DURING CITROBACTER RODENTIUM INFECTION

TL;DR

This study investigates how a protein called Tollip helps protect against a gut infection in mice, showing it plays a key role in clearing the infection and maintaining the protective mucus layer in the intestines.

Contribution

The study reveals a new protective role for Tollip in resolving Citrobacter rodentium infection through its impact on mucin production and glycosylation.

Findings

Tollip deficiency delayed bacterial clearance during the resolution phase of infection.

Tollip-/- mice showed a thinner mucin barrier and altered mucin glycosylation patterns.

Lack of Tollip did not affect initial infection kinetics or colitis severity.

Abstract

Diarrheal diseases are a threat to human health, being the second leading cause of death in children. Diarrheic enterocolitis can be caused by attaching/effacing (A/E) pathogens like enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC). Citrobacter rodentium (CR), a mouse A/E pathogen, mimics EPEC and EHEC infections, causing goblet cell depletion, crypt hyperplasia, and leukocyte infiltration. Autophagy, a critical immune pathway linked to goblet cell mucus secretion, is involved in the defence against CR. Toll-interacting protein (Tollip) is an intracellular mediator of autophagy and immunity, but its role in A/E infections remains elusive. This study aims to test the hypothesis that Tollip plays a protective role in CR infection.

- Assess CR infection kinetics in WT and Tollip-/- mice.

- Assess colitis in CR-infected WT and Tollip-/- mice.

Bacterial load was assessed by fecal colony forming unit (CFU) count in CR-infected WT and Tollip-/- mice. Histological damage was evaluated by scoring epithelial damage, leukocyte infiltration and crypt hyperplasia in H&E-stained distal colon sections. Apoptotic cells were identified by TUNEL. Acid and neutral mucin production was evaluated by Alcian-PAS staining. Glycosylation patterns were determined by immunofluorescence with lectins specific to sialic acid- and fucose-bound mucins.

Tollip
-/- and WT mice showed similar infection kinetics during the establishment and expansion phases (2-6 days post-infection (dpi)) and steady-state (7-14 dpi), with a peak of bacterial counts at 10 dpi. WT mice cleared CR by day 15, but Tollip-/- still shed bacteria by 17 dpi. The colitis grade was similar between WT and Tollip-/- mice, with marked epithelial damage at 8 dpi and increased leukocyte infiltration at 17 dpi. Scarce apoptotic cells (<4 per field) were detected in WT and Tollip-/- mice during infection. Tollip-/- showed a thinner colonic mucin barrier and less mucin-filled goblet cells at 17 dpi compared to WT mice. CR-infected Tollip-/- mice showed altered mucin glycosylation patterns, with lower fucosylated mucins in the upper crypt and lumen and increased sialic acid-bound mucins in the luminal barrier and lower crypt. The mucin barrier, goblet cells, and mucin glycosylation were similar in uninfected Tollip-/- and WT mice.

This study demonstrates a new protective role for Tollip, particularly during the resolution phase, as a lack of Tollip delayed the clearance of CR. This phenotype could be explained by deficiencies in the mucin layer observed in Tollip-/- mice, evidenced by lower mucin production and anomalies in mucin glycosylation. By enhancing our understanding of host-pathogen interactions during enteric infections, this research serves as a step in the development of new treatments and prevention strategies.

None