Involvement Of CD4~+T Cell Subsets Imbalance And Abnormal Expression Of TIM-3in The Regulation Of Inflammatory Bowel Diseases

Inflammatory bowel disease (IBD) including Crohn’s disease and ulcerative colitis is an intestinal chronic inflammatory disease whose pathogenesis mechanism is unknown. It is currently believed that genetics, atmosphere, microbiology and immune factor are involved in pathogenesis of IBD. Researches indicate immune imbalance among cell subsets may be the most direct and vital factors, thus unveiling the relationship between CD4+T cell subsets imbalance and IBD may shed light on new direction to clinical therapy.T cell immunoglobulin and mucin domain (Tim)-3is a protein first identified on terminally differentiated murine Thl cells. Engagement of Tim-3by its specific ligand galectin-9in Th1/Th17cell may lead to down regulation of immunological reaction and tolerance by inducing apoptosis. Currently, it is clear that Gal-9/Tim-3signaling pathways play an important role by inducing apoptosis of T cells in the induction of immune tolerance and autoimmune disease prevention, such as autoimmune diabetes, inflammatory bowel disease, autoimmune encephalomyelitis, rheumatoid arthritis, systemic lupus erythematosus. We prepared recombinant Gal-9protein and anti-human Tim-3monoclonal antibody to investigate its role in models of inflammatory bowel disease and regulation mechanism.Tim-3antibody was cloned and expressed successfully, and its function was identified by ELISA in vitro. DSS-induced mouse colitis model was established by DSS drinking. The model was evaluated from the weight, colon, gross and histological changes and changes in cytokines and other aspects. DSS-colitis mice were injected intraperitoneally (i. p) with rGal-9protein or anti-Tim-3monoclonal antibody to examine its role on colitis from changes of body weight and organizations. FACS and real-time PCR were used to detect the expression of T cell subsets in mouse spleen lymph cells. Anti-human Tim-3mAb genes were successfully cloned, and got the correct gene sequence. Activity of rGal-9protein and anti-human Tim-3monoclonal antibody were identified in vivo.Administration of DSS lead to mice apathetic, deprivation, diarrhea, weight loss, death and other symptoms, colon shortening, wall thickening, edema, inflammatory exudate, and multifocal ulcers changes, structural damage and loss, a large number of infiltrated inflammatory cells. The secretion level of IL-17was up-regulated, but IFN-y, Foxp3were down-regulated. The result showed that rGal-9administration prevented DSS-induced colitis. Administering anti-human Tim-3mAb, which blocked the Tim-3/Gal-9signal pathway, increased weight loss, decreased survival rate and aggravated tissue injury, which exacerbated DSS-induced colitis. The secretion of IL-17, IFN-y was up-regulated and Foxp3was down-regulated. Our data suggest that the Tim-3/Gal-9pathway is actively involved in the negative regulation of autoimmune colitis, which might impact polarization and function of different T cell subsets. The data suggest that malfunction of T cell subsets may be one of the important mechanism related to pathogenesis of IBD, rGal-9protein and neutralizing antibody of Tim-3are highly involved in intervention effect on IBD.