Insulin-specific regulatory T cell receptor alpha clonotypes restricted to HLA-DQ6 (DQB1*06:02) are shared between blood and islets

Abstract

Abstract The major genetic determinant in susceptibility or protection from many autoimmune diseases resides in the human leukocyte antigen (HLA) region. Specific class II alleles (e.g., HLA-DQ8) increase the risk for developing type 1 diabetes (T1D), a prototypical organ specific autoimmune disease, whereas HLA-DQ6 leads to dominant protection. We hypothesize that DQ6 is protective via its ability to present insulin peptides to regulatory CD4+ T cells (Tregs), resulting in anti-inflammatory responses. We expanded insulin-specific Tregs from peripheral blood mononuclear cells (PBMCs) of DQ6+ non-diabetic individuals using an insulin peptide. Insulin-expanded Tregs were flow sorted, and paired single-cell TCR/RNA-seq (10X Genomics) was performed. Of 2,096 insulin-expanded Tregs from three DQ6+ individuals, 883 αβTCRs (42%) were present ≥2 times, and we confirmed the antigen specificity of dominant clonotypes. The single-cell RNA-seq data showed 4 clusters of Treg cells based on the expression levels of FOXP3 and IL-2RA. Additionally, we flow-sorted CD4+ T cells from the pancreatic islets of a non-diabetic DQ6+ organ donor and performed single-cell TCR sequencing. Of the 67 CD4+ cells, TCR clonotypes from three cells possessed identical TCRα chains (including CDR3 sequences) with Tregs sequenced from the PBMCs of the separate DQ6+ non-diabetic individuals. These data indicate the potential presence of insulin-specific Tregs within the pancreatic islets of non-diabetic organ donors. Our findings provide a mechanistic basis for understanding HLA-linked protection from autoimmune diabetes development and have implications for manipulating self-antigen-specific T cell responses to modify the disease course in T1D.