By day 12 most mice in this study had an increased Ig response to (Determine 2)

By day 12 most mice in this study had an increased Ig response to (Determine 2). The majority of the infected mice designed systemic contamination and colitis in the mid and distal colon by day 12. bound to the major secreted mucin, Muc2, and induced MUC1 in the colon. Conclusion Major changes in both the cell-surface and secreted mucins occur in response to intestinal contamination. Introduction Bacteria such as enteropathogenic (EPEC), enterohaemorrhagic (EHEC), and are major causes of infectious diarrhea in humans worldwide. EPEC and EHEC develop a commensal, rather than pathogenic, interaction within the mouse host [1], [2]. is usually a natural mouse pathogen that is related to, and uses the same molecular mechanisms of type III secretion and attaching and effacing lesions as human EPEC and EHEC, to colonise the epithelial cells of the gut, hence providing a good model for gastroenteritis due to these bacteria [3], [4]. The gastrointestinal tract is usually lined by a constantly secreted mucus layer created by high molecular mass oligomeric mucin glycoproteins. This mucus layer moves to obvious trapped material. In the healthy human intestine, MUC2 is the main secreted mucin making up the mucus layer, whereas in the belly MUC5AC and MUC6 are produced. Colistin Sulfate Under the mucus layer, the cell-surface mucins are a dominant feature of the apical surface of all mucosal epithelial cells. Cell-surface mucins are likely to play an important role in mucosal defense since they may provide both a barrier and reporting function, and we have demonstrated increased pathology following gastrointestinal contamination in mice lacking the Muc1 cell surface mucin [5], [6]. These filamentous molecules extend further than most other cell surface structures [7]. The intestine produces the MUC1, MUC3, MUC4 MUC12, MUC13 and MUC17 cell surface mucins [8], [9]. Microbial products and inflammatory cytokines stimulate increased production of mucins by mucosal epithelial cells, which can effect a massive discharge of mucin in response to stimuli [10], [11], [12], [13], [14], [15], [16], [17], [18], [19], [20], [21]. Stimulated mucin release occurs rapidly and is accompanied by a hundredfold growth of the secretory granules upon hydration. Several pathogens have been shown to interact with mucins [22], [23], [24], [25], [26], including EPEC and EHEC which bind to bovine mucins [27]. Upregulation of MUC3 expression in colonic cells has been correlated with decreased binding of EPEC [28], [29]. The ability of human milk to limit gastrointestinal bacterial and viral infections has been attributed in part to the presence of large amounts of cell surface mucins, chiefly MUC1 and MUC15 [30], [31], [32]. Consistent with an important role for Muc1 upregulation in the intestine limiting contamination, Muc1?/? mice have a higher rate of systemic contamination in a murine model of gastroenteritis [6]. Many pathogens have evolved mechanisms to subvert the mucin barrier, for example the StcE zinc metalloprotease secreted by EHEC is usually a mucinase [33]. Goblet cell depletion in contamination has been reported previously [34], however, no comprehensive study of expression of all mucins in an animal contamination model has been performed. We map the localization along the intestinal tract of all murine mucins for which antibodies are Colistin Sulfate available: Muc1, Muc2, Muc4, Muc5AC, Muc13 and Muc3 (orthologue of human MUC17, MTRF1 therefore hereafter referred to as Muc17. There is currently no Muc3 orthologue annotated in the mouse genome. However, a peptide identical to a peptide from your human MUC3 mucin has been recognized using proteomics on mouse mucins, suggesting the presence of the mouse orthologue to this mucin [35]). In a murine contamination model, we demonstrate substantial changes in the amount of virtually all intestinal mucins after 12 days of contamination. We further showed mucin binding to strain ICC169 was produced on Lauria-Bertani agar for 20 h at 37C. Bacteria harvested from plate cultures were suspended in warmed Lauria-Bertani broth. 8 mice were orally inoculated with 107 colony forming models (CFU) and sacrificed after 12 days by cervical dislocation. Duplicate samples of duodenum, caecum, small intestine and proximal, mid and distal colon were dissected and collected in either broth or 10% formalin. To assess the quantity of CFU/g tissue, tissue and fecal samples (collected as defecated samples) were homogenized in broth, serially diluted, plated onto McConkey’s selective agar, and produced for 20 h at 37C. CFU were enumerated by counting 1 mm diameter fuschia-coloured colonies. The mice were weighed and diarrhea was scored every second day. Histological assessment For analysis of colitis, formalin fixed Colistin Sulfate tissue sections of the small intestine, caecum, proximal and distal colon stained Colistin Sulfate with hematoxylin/eosin were coded to blind the analysis, and the entire section was systematically scored: aberrant crypt architecture (0C5), Colistin Sulfate increased crypt length (0C3), goblet cell depletion (0C3), general leukocyte infiltration (0C3),.