Type 1 diabetes (T1D) is a polygenic T cell mediated autoimmune disease with strong genetic linkages to the MHC class II region and insulin gene promoter region. Murine model studies indicated that insulin B:9-23 (InsB:9-23 is a crucial epitope for diabetes initiation in NOD mouse. The diabetogenic T cells have been shown to recognize InsB:9-23 peptide bound to I-Ag7 with different registers. However, in humans, the existence of DQ8 restricted InsB:9-23 specific T cells has not been conclusively demonstrated and the register that the peptide is bound and recognized in the context of HLA-DQ8 has not been established. In this study, we developed a HLA-DQ8/InsB:11-23R22E tetramer to detect insulin specific T cells. We found that tetramer positive T cells are more frequently detected in T1D patients (6/16 subjects) than in healthy controls (0/12 subjects, Fisher exact test, p=0.02). Although these T cells were isolated with tetramer prepared with modified peptide, they responded vigorously to stimulation by wild type Ins:B11-23 peptide as well as denatured insulin – with maximal observed stimulation indexes of 70.3±60.1 and 86.7±75.4 for wild type peptide and denatured insulin, respectively. T cell clones were isolated from several different patients, and they recognize the InsB:11-23R22E and InsB:11-23 peptides bound to HLA-DQ8 with Register 3. Furthermore, these T cells were also activated by homologous peptides derived from several microbial antigens that are present in the environment.
In summary, we demonstrated that insulin specific T cells are commonly present in T1D patients. These cells recognize the InsB:9-23 peptide bound to DQ8 in register 3. These T cells can be activated by homologous peptides derived from environmental microbes and are ready to respond to stimulation by antigen-presenting cells primed with denatured insulin protein. Thus, InsB:11-23 presented on DQ8 can play an important role in the initiation of T1D in human.