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MUC2-DEFICIENT MICE EXHIBIT DEFECTS IN TIGHT JUNCTION PROTEINS AND COLONIC PERMEABILITY
E Trusevych, C Hirota, F Moreau, K Tran, J Meddings, W MacNaughton, K ChadeeUniversity of Calgary, Calgary, AB
Aims: The intestinal mucosal barrier serves as the first line of host defence in the gastrointestinal (GI) tract, and has two main constituents: the mucus-gel and the underlying single layer of epithelial cells connected by tight junctions. MUC2 produced by goblet cells is the main structural component of the mucus layer. It is unknown if MUC2 regulates epithelial barrier function. During inflammatory bowel diseases, MUC2 levels are diminished and epithelial permeability is increased. In this study we investigated whether mice lacking Muc2 mucin (Muc2-/-) have a compromised epithelial barrier that increases their susceptibility to intestinal inflammation.
Methods: Full thickness segments of ileum and colon from Muc2-/- and Wt mice were mounted in Ussing chambers to access ion transport, transepithelial resistance (TER) and mucosal to serosal flux of FITC-dextran (3-5kDa). In other experiments, protein was isolated from intestinal tissues and processed for western blotting for tight junction proteins. To determine site-specific and temporal permeability changes in vivo, Muc2-/- and Wt mice were gavaged once per week for three months with a solution containing sucrose (permeability marker: gastric), lactulose (small intestine), mannitol (small intestine) and sucralose (colon). Urine samples collected for 22 h post-gavage were analyzed for the presence of the oligosaccharides using HPLC.
Results: Baseline short-circuit current was similar in Muc2-/- and Wt mice, however Muc2-/- mice had increased TER in the proximal and distal colon compared to Wt mice. Additionally, the mucosal to serosal flux of FITC-dextran was lower in Muc2-/- mice than Wt mice, indicating that Muc2-/- mice have reduced paracellular permeability. Of the tight junction proteins tested, barrier-forming occludin was increased at every age (3 wk, 2 mo and 5 mo) in Muc2-/- mice, whereas cation-selective claudin-2 was only increased in Muc2-/- mice after 2 mo of age. In vivo permeability measurements indicate no change in gastric permeability, decreased small intestinal permeability in Muc2-/- mice, and an age-dependent increase in Muc2-/- colonic permeability.
Conclusions: Surprisingly colonic epithelial barrier functions were enhanced in the absence of Muc2 mucin. However, in vivo permeability measurements show temporal changes in colonic permeability that were not identified with ex vivo techniques suggesting that multiple barrier function defects may occur in the absence of Muc2.