Aberrant immune regulation and enrichment of stem-like CD8+ T cells in the pancreatic lymph node during type 1 diabetes development

Effector CD8+ T cells are key cellular drivers of type 1 diabetes (T1D) pathogenesis, yet questions remain regarding the molecular defects leading to altered cytotoxicity, their signature in peripheral tissues, and their receptor specificity. Thus, we analyzed human pancreatic lymph nodes (pLN) using mass cytometry and single cell RNA sequencing (scRNAseq) with combined proteomic and T cell receptor (TCR) profiling. Cytometric analysis revealed an enriched population of T stem-cell memory (TSCM)-like cells (CD8+CD45RA+CD27+CD28+CCR7+CXCR3+ T cells) in T1D pLNs. scRNAseq profiling indicated an elevated inflammatory cytokine gene signature (IFITM3, LTB) along with regulators of terminal differentiation (BCL6, BCL3), coupled with reduced expression of exhaustion-associated genes (DUSP2, NR4A2, TSC22D3) in CD8+ T cells in T1D pLN. Additionally, effector CD8+ T cells expressed features of progenitor exhausted cells (BCL2) in T1D pLN. Immune Response Enrichment Analysis (IREA) indicated IL-15 signaling as a significant driver of these phenotypes. Integrated TCR and transcriptomic analysis revealed a cluster of diverse naive-like CD8+ T cell clones in T1D pLN. When comparing pLN and pancreatic slice cellular isolates, we observed sharing of effector CD8+ T cells, with upregulation of terminal effector signatures detected within the pancreas relative to paired pLN samples. Multiplex imaging revealed differential localization of TCF1 and TOX expressing T cells in the pancreas, with TCF1+TOX+ cells located in closer proximity to the islets and displaying a mixture of activation and exhaustion-associated phenotypes. Thus, we provide multimodal cellular profiles enriched in T1D tissues for consideration in therapeutic targeting.