628 Macrophage-Epithelial Crosstalk Regulates Mucosal Regulatory T Cell Expansion Through TLR4-Mediated Peroxisome Proliferator-Activated Receptor (PPARy) Activation During Colitis

Abstract

SHP-2 (Src homology-2 domain containing protein tyrosine phosphatase-2) is a tyrosine phosphatase ubiquitously expressed. Previous works demonstrated SHP-2 implication in processes such as proliferation and migration as well as signaling pathways including the RAS-RAF-MEK-ERK and Jak/STAT pathways. Even though SHP-2 is highly expressed in intestinal epithelium, its role in this tissue is not described. Recently, polymorphisms in PTPN11 gene encoding SHP-2 have been described as potential biomarkers for ulcerative colitis susceptibility. We thus investigated the role of epithelial SHP-2 in the control of intestinal homeostasis. Methods: Using the Cre/loxP system, mice with a conditional intestinal epithelial SHP-2 deficiency were generated (mutant mice). Western blots were performed in intestinal mucosa enrichments. Tissue architecture was visualized with hematoxylin-eosin staining, Goblet cells with Alcian blue staining and Paneth cells with lysozyme immunofluorescence staining. Cytokines and chemokines were quantified by Inflammation Antibody Array. Intestinal permeability was assessed by determining the transmucosal transport of fluorescein isothiocyanate (FITC) conjugated dextran. SHP-2 mRNA expression in intestinal mucosa enrichments from inflammatory bowel disease patients was evaluated by TissueScan qPCR arrays. Results: 1Deletion of SHP-2 in conditional knockout mice was validated by the loss of SHP-2 protein expression in intestinal epithelium. 2One month aged mutant mice were smaller than their control littermates and exhibited signs of severe colitis (diarrhea and blood in stool). 3Colon architecture of mutant mice was markedly altered with infiltration of immune cells, longer colonic crypts, presence of crypt abscesses, decreased number of Goblet cells and aberrant presence of Paneth cells. 4Inflammatory transcription factors STAT3 and NFkB were rapidly hyperactivated in the colonic epithelium of mutant mice, 1 day and 2 weeks after birth, respectively. 5The epithelial chemokines CXCL1, CXCL5 and CCL5 were markedly enhanced in two week-old mutant mice. In three weekold mice, IL-1α, IL-1β, IL-6 and IL-12 levels were significantly enhanced in contrast to TNFα and IFNγ. 6Expression of claudins 1, 4, 8 and 15 as well as occludin was markedly decreased in mutant mice, two weeks after birth; this is associated with increased intestinal permeability. 7Finally, a significant decrease of SHP-2 mRNA expression was observed in patients with Crohn's disease and Ulcerative colitis. Conclusion: These results indicate that intestinal epithelial SHP-2 is a critical determinant for prevention of gut inflammation. The decreased number of Goblet cells, the hyperactivation of inflammatory transcription factors as well as the increase in intestinal permeability could explain the appearance of intestinal inflammation in absence of SHP-2.