Type 1 diabetes (T1D) results from T cell-mediated destruction of insulin-producing β-cells. Insulin represents a key self-antigen in disease pathogenesis, as recent studies identified proinsulin-responding T cells from inflamed pancreatic islets of organ donors with recent-onset T1D. These cells respond to an insulin B-chain (InsB) epitope presented by the HLA-DQ8 molecule associated with high T1D risk. Understanding insulin-specific T-cell frequency and phenotype in peripheral blood is now critical. We constructed fluorescent InsB10-23:DQ8 tetramers, stained peripheral blood lymphocytes directly ex vivo, and show DQ8+ patients with T1D have increased tetramer+ CD4+ T cells compared with HLA-matched control subjects without diabetes. Patients with a shorter disease duration had higher frequencies of insulin-reactive CD4+ T cells, with most of these cells being antigen experienced. We also demonstrate that the number of insulin tetramer+ effector memory cells is directly correlated with insulin antibody titers, suggesting insulin-specific T- and B-cell interactions. Notably, one of four control subjects with tetramer+ cells was a first-degree relative who had insulin-specific cells with an effector memory phenotype, potentially representing an early marker of T-cell autoimmunity. Our results suggest that studying InsB10-23:DQ8 reactive T-cell frequency and phenotype may provide a biomarker of disease activity in patients with T1D and those at risk.
Bibliographical noteFunding Information:
Funding. This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases DK-108868 (A.W.M.); National Institute of Allergy and Infectious Diseases P01-AI-35296 (M.K.J. and B.T.F.), R01-AI-106791 (B.T.F.), and U24-AI-118635 (B.T.F.); Regenerative Medicine Minnesota RMM-11215-TR002 (B.T.F.); and JDRF 3-2014-215 (J.A.S.) and 17-2014-3 (B.T.F.). Duality of Interest. No potential conflicts of interest relevant to this article were reported. Author Contributions. J.A.S., N.L.S., J.C.W., T.M., and B.T.F. designed and performed the experiments and analyzed the data. J.A.S., T.D., M.K.J., and B.T.F. designed, generated, and validated the Ins:DQ8 tetramer. J.A.S., A.W.M., and B.T.F. directed the studies, prepared the figures, and wrote the manuscript. A.L.B., E.B.F., A.W.M., and A.M. procured the human samples. E.B.F. and B.R.B. provided critical feedback on the manuscript. All authors reviewed the manuscript. B.T.F. is the guarantor of this work and, as such, had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis.