The Role And Mechanism Of Myeloid Differentiation 1 (MD-1) In Murine Experimental Colitis

Backgrounds and Aims Inflammatory bowel disease (IBD) comprises the chronic relapsing inflammatory disorders Crohn’s disease and ulcerative colitis. The precise aetiology of IBD remains unclear, but several factors that make a major contribution to disease pathogenesis have been identified. These factors mainly include:genetic factors, the host immune system, environmental factors such as the gut microbiota, epithelial barrier function, autophagy and endoplasmic reticulum stress. Available evidence suggests that both dysregulated innate and adaptive immune pathways contribute to the aberrant intestinal inflammatory response in patients with IBD. As the most effective and specialized type of antigen presenting cells (APCs), DCs can recognize and process foreign antigens, present antigens to T cells, activate them, and further influence immune response, which reveal the bridge function of DCs in connecting innate and adaptive immunity. Rencent studies in the genetics and immunology of mucosal diseases suggest that DCs from intestinal lamina propria play a vital role in the pathophysiology of IBD. Myeloid differentiation 1 (MD-1) is a secteted glycoprotein that forms a complex with RP105 restrictively expressed on DCs, B cells and macrophages, which plays an important role in Toll-like receptor 4 (TLR4) signaling pathway. Growing evidence suggests that MD-1 may be involved in the (patho) physiological regulation of the innate immune system and inflammation via modulating the functions of DCs. Previous studies suggested that TLR4/MD-2 plays an important role in regulating intestinal inflammation of IBD and experimental colitis animal models. However, no studies have been reported on MD-1/RP105 involving with IBD. Therefore, to investigate its impact on IBD, we detected the mRNA expression of MD-1 and RP105 in IBD patients and experimental colitis mice. Meanwhile, we studied the effects of the loss of MD-1 on dextran sodium sulfate (DSS)-induced colitis in mice.Methods IBD patients included 33 CD and 33 UC as well as 16 healthy control subjects wee recruited in our study. Acute experimental colitis model was established by orally receiving 3%DSS in drinking water for 7 days in wild type (WT) mice. The expressions of MD-1 and RP105 in IBD patients and experimental colitis mice were detected by quantitative real-time-PCR (qRT-PCR). WT or MD-1 deficient (MD-1-/-) mice were received 3% DSS solution in drinking water for 7 days. Body weight, morbidity, stool consistency, and the presence of gross blood in feces and at the anus were monitored daily by two independent observers. On day 7, the colon length, spleen and colon weight were calculated. H&E and immunohistochemistry (IHC)/immunofluorescence (IF) staining were performed on distal colon tissue to analyze inflammatory cells infiltration and tissue damages after induction of colitis. The mRNA expressions of cytokines, chemokines and Th cells’specific transcription factors were measured by qRT-PCR. To evaluate the intestinal permeability, DSS induced colitis mice were intragastrically administrated with FITC-conjugated dextran (0.6mg/g body weight). The FITC-dextran concentration in the plasma was detected by ELISA. Lamina propria mononuclear cells (LPMCs) were isolated by enzymic digestion method from the entire colon tissue. The number of DCs in LPMCs and the expressions of maturation molecules in LPDCs were analyzed by flow cytometry.Results MD-1 and RP105 mRNA expression were up-regulated in both human IBD patients and DSS-treated WT mice. Both MD-1-/- and WT mice successfully developed acute colitis following 7 day’s DSS administration. MD-1-/- mice were less susceptible to the development of colitis than WT mice as demonstrated by significantly reduced weight loss, disease activity index, shortening of colon length, colon histological scores, the number of inflammatory cells’ filtration and also reduced mRNA levels of pro-inflammatory cytokines (TNF-a, IFN-y, IL-6, IL-12p35, IL-13 and IL-17A), chemokines (KC, MCP-1 and MIP-2) as well as the Th-related transcription factors (GATA3). In addition, mucosal barrier function seemed to be intact in response to the loss of MD-1. Finally, LPDCs from the colon of MD-1-/-mice after DSS exposure not only decreased in number but also significantly down-regulated the expressions of surfac maturation co-stimulator molecules MHC-II, CD40 and CD86 compared with those from WT mice.Conclusions We reported for the first time that elevated MD-1 and RP105 mRNA is found in the colon of IBD patients and diseased mice, which suggests that MD-1/RP105 may be involved in the pathogenesis of IBD. Our data provided novel evidence that MD-1 deficiency has a protective effect on DSS-induced colitis by modulating the maturation of colonic LPDCs without aggravating epithelial barrier function. Taken together, our results reveal that MD-1 deficiency is of critical importance in down-regulating induction and progression of DSS induced colitis in mice, thereby suggesting that MD-1 might be a target for future interventional therapies of IBD.