T1212 Upregulation of E3 Ubiquitin Ligases Related to T Cell Anergy in CD4+ T Cells Isolated from Patients with Ulcerative Colitis in Remission

Abstract

Background and Aims Insufficient tolerance against luminal antigens is suggested to cause inflammatory bowel diseases including ulcerative colitis (UC). T cell anergy is one of the mechanisms regulating immune tolerance. Recently, some E3 ubiquitin ligases are reported to downregulate downstream signals of T cell receptor and induce T cell anergy. The aim of this study was to investigate the state of T cell anergy in healthy subjects and UC patients by quantitating the expression of E3 ubiquitin ligases, GRAIL, c-Cbl, Itch, all of which are reported to induce anergy on T cells. Patients and Methods Peripheral mononuclear cells were obtained from 20 UC patients (active UC: 12, UC in remission: 8) and 10 healthy volunteers (HV) by either centrifugal leukocytapheresis or sampling of peripheral blood. CD4+ T cells were isolated by magnetic cell sorter and expressions of GRAIL, c-Cbl, and Itch mRNA in these cells were measured by real-time quantitative RT-PCR. The level of GRAIL, c-Cbl, and Itch mRNA expression were compared between in active UC patients, UC patients in remission, and HV. The expression of these ubiquitin ligases were followed in 6 active UC patients who were treated by corticosteroid and centrifugal leukocytapheresis. Results Anergic CD4+ T cells obtained by ionomycin treatment In Vitro revealed higher GRAIL, c-Cbl, and Itch expression when compared with non-anergic T cells. GRAIL was predominantly expressed on CD4+ T cells whereas c-Cbl and Itch were also expressed on non-CD4+ T cell subsets. The GRAIL expression in CD4+ T cells of UC patients in remission was significantly higher than that of active UC patients or that of HV (p<0.01). The c-Cbl and Itch expressions in CD4+ T cells of UC patients in remission were relatively higher than that of active UC or that of HV, but there were no significant differences. Remarkably the levels of GRAIL expression were significantly increased after the treatment in UC patients whose clinical activities ewre improved by the treatment. However, the c-Cbl and Itch expression did not increase after the treatment. Conclusion Among the 3 different E3 ubiquitin ligases which induce T cell anergy, upregulation of GRAIL expression seems to be associated with the induction of remission in UC patients. These results indicate that GRAIL is a potential therapeutic target of inflammatory bowel disease.