3021-LB: NKG2D Signaling as a Critical Modulator of Autoreactive CD8+ T Cell Differentiation and ß-Cell Destruction in Type 1 Diabetes

Introduction and Objective: Type 1 diabetes (T1D) is an autoimmune disease driven by T cell mediated destruction of pancreatic β cells. However, the signals that regulate early activation and differentiation of pathogenic CD8+ T cells remain incompletely understood. The activating immune receptor NKG2D has been implicated in T1D pathogenesis, yet its role remains controversial due to conflicting reports. We investigated whether NKG2D signaling regulates early autoreactive cytotoxic T lymphocyte (CTL) priming during autoimmune diabetes. Methods: To study the role of NKG2D, we generated multiple genetically modified mouse strains on the non-obese diabetic (NOD) background. Diabetes incidence, insulitis, spatial proteomic profiles, and islet-specific CD8+ CTL differentiation were compared between NKG2D deficient and wild type mice housed under specific pathogen free and germ free conditions. Results: NKG2D deficient NOD mice exhibited reduced diabetes incidence, decreased insulitis, and diminished pancreatic CD8+ T cell infiltration. The protective phenotype was most pronounced in germ free and male mice, suggesting modulation by microbiome dependent signals. Mechanistic studies demonstrated that NKG2D signaling does not occur within the pancreas but instead during early immune priming in the pancreatic lymph node, where NKG2D ligands are enriched. Deletion of NKG2D specifically in CD8+ T cells delayed disease onset and impaired differentiation of islet specific naïve CD8+ T cells into CTLs. Transcriptomic profiling revealed altered CTL differentiation programs in NKG2D deficient cells. Pharmacologic blockade of NKG2D signaling similarly reduced human CTL differentiation. Conclusion: These findings identify NKG2D signaling as a critical signaling pathway in early autoreactive CTL priming in T1D and provide a potential explanation for conflicting reports by revealing a microbiome and sex influenced compensatory pathway that buffers CTL differentiation. Disclosure A. Dudhate: None. A. Manning: None. H.E. Poague: None. E.M. Puckett: None. Z. Bedrosian: None. M.A. Markiewicz: Consultant; Current; Design-Zyme, LLC. Funding Breakthrough T1D (3-SRA-2023-1412-S-BDOD: HT9425-23-1-0630), Emilie Rosebud Diabetes Research Foundation, University of Kansas Diabetes Institute