Mucins in Infectious Colitis Figure 6. Tissue localization of mucins in the distal large intestine. The apical surface of the distal large intestinal epithelium and crypts are lined by the cell surface mucins Muc13 and Muc17 as well as a small amount of Muc4 (brown). The goblet cells contain Alcian blue positive material (blue = highly charged carbohydrate structures, purple = medium charged carbohydrate structures), which mainly is Muc2 (brown), although the goblet cells of the surface also contain Muc4. The photographs were taken using a 206 magnification. The quantification scores for Alcian blue, Muc1, Muc13 and Muc17 were significantly different between non-infected mice (#) and mice infected with C. rodentium for 12 days (N) (* p,0.05, Mann Whitney U test, n = 7). SCL = subcellular localization, RC = region of crypt. The mucin stains are shown in serial sections on the same tissue area, whereas the mucin stain scores represent the average stain scores for the entire specimen for each individual mouse. doi:10.1371/journal.pone.0003952.g006 response shuts down mucin production and if more prolonged or severe causes apoptosis [39,52,53]. It seems likely that the host attempts to remove pathogens by increasing secretion of mucins to trap and remove pathogens from the epithelial surface, whereas the pathogen tries to eliminate this host defence by inhibiting the mucin synthesis and causing shedding of the mucus producing epithelial cells. Interestingly, goblet cell depletion during infection does not appear to occur in the respiratory tract where goblet cell hyperplasia and mucus hypersecretion can continue for extended periods [54]. Furthemore, changes in mucin secretion is likely to affect the resident microflora due to the large change in glycosylated compounds available for bacterial degradation [55,56]. In the small intestine and proximal large intestine, C. rodentium was present, albeit in lower amounts than in the distal large intestine, and yet there were few signs of mucosal damage, inflammation or changes in mucins. Furthermore the highest concentrations of bacteria were detected in the colons of animals with the highest amount of colonic damage, mucin depletion, and systemic infection. Even within animals, damage and inflamma- tion in the distal colon was often patchy suggesting that focal bacterial penetration of the barrier leads to localised mucin release and inflammatory responses. Why these bacteria penetrate the barrier in the distal colon remains to be elucidated but could be due to an increased bacterial pathogen load in the distal colonic niche, or an ability to degrade the mucin glycoforms in this region, and/or involve induction of pathogenicity-related genes in the bacteria due to local environmental cues. We have demonstrated that MUC2 is one such cue for C. jejuni which switches on pathogenicity genes in response to mucin exposure [57]. An IgG response has been shown to be integral to clearance of C. rodentium infection [58] and it appears pertinent for our understanding of the onset of humoral immunity that the C. rodentium specific Ig production at this stage of infection was restricted to those mice in which the bacteria had penetrated the mucosal barrier. C. rodentium infection causes colonic inflammation, mucosal hyperplasia, epithelial dysfunction in association with increased permeability to luminal bacteria and a vigorous Th1 inflammatory response [59]. Thus C. rodentium infection elicits mucosal inflammation with similarities to inflammatory bowel disease, and has therefore been used to investigate the relationship between inflammation and anti-bacterial immunity in the gut [4]. MUC5AC, which is a product of normal gastric mucosa, is absent from normal colon but frequently present in colorectal adenomas and colon cancers [60,61], and expressed in patients with Figure 7. C. rodentium binding to Muc2. ELISA plates were coated with the ‘‘insoluble’’ Muc2 complex or with BSA and incubated with biotinylated C. rodentium. Statistics: Mean6SD, *** p,0.001 (t-test, n = 6) (A). Large amounts of bacteria can be found entangled in the secreted Muc2 in C. rodentium infected animals (B–D). By brightfield observation using a 206 lens (B) the Muc2 is visualized as a brown stain, and the presence of large amounts of bacteria within the secreted Muc2 can be seen by reflected light Nomarski differential interference observation (C; 406 lens and D; 1006 lens). doi:10.1371/journal.pone.0003952.g007 PLoS ONE | www.plosone.org 11 December 2008 | Volume 3 | Issue 12 | e3952