Activation of antigen-specific T cells in vivo induced by tolerogenic encounter with antigen-coupled splenocytes

Intravenous injection of ethylene carbodiimide-fixed, antigen-coupled spienocytes is a potent method of inducing T cell tolerance in vitro and in vivo. We have shown that injecting neuroantigen-coupled splenocytes i.v. blocks disease induction and treats ongoing experimental autoimmune encephalomyelitis. To explore the mechanisms involved in in vivo tolerance, we utilized D011.10 T cell receptor transgenic mice and BALB/c recipients of D011.10 T cells. To mimic the R-EAE tolerance model, we examined the effects of i.v. administered OVA 323-339-coupled splenocytes (OVA323-SP) before or after priming with ovalbumin 323-339 (OVA323) in CFA. Tolerance induction prior to priming results in diminished antigen-specific T cell proliferative and IFN-g production responses without altering the number of clonotypic T cells. Tolerance induction following peptide priming resulted in diminished IL-2 and IFN-g production and proliferative responses as compared to controls. Unlike pre-tolerance, the percentage of transgenic cells in the spleen and lymph nodes was double that of controls. Examination of the direct effects of OVA323 injection in the DO11.10 mouse revealed that specific T cells are rapidly activated as assessed by changes in CD25, CD44, CD69 and CD62L expression on clonotypic cells. These cells failed to proliferate or become activated in vitro when cultured with peptide. In addition, tolerized cells rapidly upregulated CTLA-4. Thus, ECDI-fixed, antigen-coupled splenocyte treatment results in antigen-specific cell-activation leading to unresponsiveness.