P011 Dysbiosis and Goblet cells depletion triggers early intestinal barrier dysfunction that precedes gut inflammation in IL-10 deficient mice (IL-10−/−)

Background Interleukin-10 deficient mouse (IL-10−/−) is a widely used model of spontaneous ileocolitis that resembles human inflammatory bowel disease (IBD); intestinal barrier dysfunction is an early pathophysiological event, but its underlying mechanisms are still unknown. The objective of this work is to study the natural history of ileocolitis in IL-10−/−, and unravel the influence of intestinal barrier dysfunction and dysbiosis in the development of overt inflammation. Methods Wild-type (WT) and IL-10−/− mice were followed until sacrifice at 3, 5, 10, 20, 57 and 70 weeks of life. Bodyweight, colonic weight/length ratio and in vivo intestinal permeability (measured by rectal administration of FITC-dextran) were registered. After the sacrifice, the colon was harvested and the evaluation of the expression of inflammatory (interleukin-1β (IL-1β), tumour necrosis factor-α (TNF-α), inducible nitric synthase (iNOS) and cyclooxygenase-2 (COX-2) and epithelial permeability (ZO-1, E-cadherin, Occludin, Claudins 2 and 7, and Reticulon-4B (RTN-4B) markers was performed by qPCR; expression of mucin-2 (MUC-2) and molecules involved in goblet cell maturation such as interleukin-18 (IL-18) and WAP Four-Disulphide Core Domain 2 (WFDC2), as well as the endoplasmic reticulum stress marker X-box-binding protein (Xbp)-1) by qPCR were also analysed. We also used colon slices for histologic evaluation with haematoxylin-eosin and alcian blue stainings. The microbiota composition was studied by sequencing of the V3-V4 regions of ribosomal 16S from faecal samples of all these mice. Results Compared with WT, IL-10−/− mice showed lower weight gain at all ages and a higher colonic weight/length ratio and histological evidence of inflammation at weeks 20 and 57. iNOS and IL-1b gene expression in the colon were significantly higher in IL-10−/− mice at weeks 10 and 20, respectively. Nevertheless, increased intestinal permeability was observed from week 10; the number of goblet cells and expression of MUC-2, IL-18, WFDC2 and XBP-1 were significantly lower in knockout mice from week 10. Moreover, dysbiosis in IL-10−/− mice began at week 5, increasing at 10 and showing the lowest diversity and appearance of pathogenic families at 20 weeks of age. Conclusion Dysbiosis and goblet cell depletion in the colon of IL-10−/− mice are associated with early intestinal barrier dysfunction, and precede overt gut inflammation in this animal model of IBD.