High dimensional single cell characterization of autoreactive CD8 T-cells reveals heterogeneous phenotypes that play a role in disease progression in Type 1 Diabetes

Abstract

Abstract Type 1 Diabetes (T1D) is caused by beta cell destruction, eventually resulting in loss of glycemic control. Following onset, subjects with T1D exhibit diverse amounts of residual c-peptide, indicating varying levels of beta cell function. Persistence of c-peptide is correlated with reduced risk of complications and its preservation has been used as an endpoint in clinical trials. However, the immunologic factors that differentiate subjects who retain measurable c-peptide from those who do not are not well characterized. We investigated the number and phenotype of beta cell specific CD8+ T-cells in peripheral blood samples from a cohort of T1D subjects, stratified into slow progressors (c-peptide > 0.1ng/ml) and rapid progressors (undetectable c-peptide), including longitudinal samples. Using tetramers corresponding to HLA-A2 epitopes, we enumerated epitope specific T-cells and sorted single cells for RNA-Seq. Differential gene expression between slow and rapid progressors indicated differences in effector, exhaustion, and regulatory pathways which were consistent with a recently published high dimensional CyTOF data set. Assembly of autoantigen specific TCRs revealed differences in clonal expansion for specific cell clusters. Pseudotime trajectory analysis of the scRNAseq data revealed that CD8+ T cells expressing the same TCR resided in different phenotypic end states. Furthermore, flow cytometric analyses demonstrated differential expression of lineage-specific cell surface markers, which correlated well with the transcriptional profiles within each cluster. Cumulatively, we discovered specific aspects of T cell function and repertoire that differ between slow- and rapid-progressing T1D subjects.