Assessment of CD4+ T Cell Responses to Glutamic Acid Decarboxylase 65 Using DQ8 Tetramers Reveals a Pathogenic Role of GAD65 121–140 and GAD65 250–266 in T1D Development

Abstract

Susceptibility to type 1 diabetes (T1D) is strongly associated with MHC class II molecules, particularly HLA-DQ8 (DQ8: DQA1*03:01/DQB1*03:02). Monitoring T1D-specific T cell responses to DQ8-restricted epitopes may be key to understanding the immunopathology of the disease. In this study, we examined DQ8-restricted T cell responses to glutamic acid decarboxylase 65 (GAD65) using DQ8 tetramers. We demonstrated that GAD65121–140 and GAD65250–266 elicited responses from DQ8+ subjects. Circulating CD4+ T cells specific for these epitopes were detected significantly more often in T1D patients than in healthy individuals after in vitro expansion. T cell clones specific for GAD65121–140 and GAD65250–266 carried a Th1-dominant phenotype, with some of the GAD65121–140-specific T cell clones producing IL-17. GAD65250–266-specific CD4+ T cells could also be detected by direct ex vivo staining. Analysis of unmanipulated peripheral blood mononuclear cells (PBMCs) revealed that GAD65250–266-specific T cells could be found in both healthy and diabetic individuals but the frequencies of specific T cells were higher in subjects with type 1 diabetes. Taken together, our results suggest a proinflammatory role for T cells specific for DQ8-restricted GAD65121–140 and GAD65250–266 epitopes and implicate their possible contribution to the progression of T1D.

Antigen-Specific T Cell Analysis Reveals That Active Immune Responses to β Cell Antigens Are Focused on a Unique Set of Epitopes

“…To determine whether the epitopes recognized by these CD38+ T cells were presented by HLA-DR or HLA-DQ, we analyzed HLA restrictions for 10 oligoclones that we had cells available (Figure 4). We found that the reactivities of all these oligoclones were blocked by anti-HLA-DR antibody but not by anti-HLA-DQ antibody, including the reactivity to GADp32-specific peptide, which has been shown to contain a HLA-DQ restricted epitope [29] (Figure 4). These results suggest that all these oligoclones recognizes peptides presented on HLA-DR.…”

Single-Cell RNA Sequencing Reveals Expanded Clones of Islet Antigen-Reactive CD4+ T Cells in Peripheral Blood of Subjects with Type 1 Diabetes

“…The presence of antigen specific T cells in peripheral blood of T1D as well as HC subjects, even cells with a memory phenotype, has been a surprising finding (7–9, 51). The widespread detection of these islet antigen-reactive T cells may result from their expression of TCRs that cross react with pathogen derived antigens, as has been reported for the islet antigens insulin and IGRP (53, 54).…”

Autoreactive T cells in type 1 diabetes

“…Several islet cell molecules were identified as targets of autoimmunity in T1D, including native proteins and epitopes of proinsulin (9,(50)(51)(52)(53)(54)(55)(56)(57)(58)(59)(60)(61)(62)(63), GAD65 (64)(65)(66)(67)(68)(69)(70)(71)(72), tyrosine phosphatase-like insulinomaassociated antigen 2 (IA-2) (67, 73-75), islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) (76-80), the cation + T cells, the predominant immune cell type in the insulitis. As described in the main text, these may include native antigens and neoepitopes.…”