“Deep phenotyping of islet autoreactive and immunoregulatory T-cells in type 1 diabetes”
Abstract:
Auto‐reactive CD8 T‐cells play an important role in the destruction of pancreatic β‐cells resulting in type 1 diabetes (T1D). However the phenotype of these auto‐reactive cytolytic CD8 T‐cells has not yet been extensively described. We used high‐dimensional mass cytometry to phenotype auto‐ (pre‐proinsulin), neo‐ (insulin‐DRIP) and virus‐ (Cytomegalovirus) reactive CD8 T‐cells in peripheral blood mononuclear cells (PBMCs) of T1D patients. A panel of 33 monoclonal antibodies was designed to further characterize these cells at the single‐cell level. HLA‐A2 class I tetramers were used for the detection of antigen‐specific CD8 T‐cells. Using a novel Hierarchical Stochastic Neighbor Embedding (HSNE) tool (implemented in Cytosplore), we identified 42 clusters within the CD8 T‐cell compartment of three T1D patients and revealed profound heterogeneity between individuals, as each patient displayed a distinct cluster distribution. Single‐cell analysis of pre‐proinsulin, insulin‐DRIP and cytomegalovirus‐specific CD8 T‐cells showed that the detected specificities were heterogeneous between and within patients. These findings emphasize the challenge to define the obscure nature of auto‐reactive CD8 T‐cells. Using a different panel of markers we were able to identify regulatory and effector T‐cells induced by priming with tolerogenic dendritic cells pulsed with proinsulin peptide, which led to the discovery of biomarkers to be monitored in our clinical trial.