Introduction and Objective: We have shown that beta cell-derived extracellular vesicle (EV) programmed death ligand-1 (PD-L1) reduces CD8+ T-cell activity and may protect against beta cell killing in T1D. However, underlying mechanisms linking differences in islet EV cargo to heterogeneity in beta cell survival are poorly defined. Methods: EVs were isolated using size exclusion chromatography from nondiabetic human islets, plasma and pancreas slice perifusates from donors with or without a history of T1D or islet autoantibody-positivity (AAB+). EV cargo was quantified using ELISA or single-EV analysis (ExoView). PD-L1 transcript variants were assessed by RT-qPCR and Basescope. Results: Plasma EV PD-L1 was elevated in AAB+ individuals compared to euglycemic controls: but within the AAB+ group, EV PD-L1 was lower in those who developed clinical T1D during follow-up compared to non-progressors. Interferon-α exposure (2000 U/mL, 24h) induced heterogenous increases in human islet EV PD-L1 (average 4-fold) but also EV HLA class I (average 2.5-fold). Notably, 0.5% of EVs co-expressed PD-L1 and HLA-I, indicating segregation into distinct EV subpopulations. In pancreas slice perifusates, both EV PD-L1 and HLA-I were increased in donors with T1D. However, EV PD-L1 positively correlated with glucose-stimulated insulin secretion (r=0.89, p0.001), while EV HLA class I showed a strong inverse correlation (r=-0.88, p0.001). IFN-α or Coxsackievirus B exposure also induced an exon-3-skipped PD-L1 transcript variant. Transgenic cell lines overexpressing this variant exhibited 80% reduced EV incorporation. Conclusion: IFN-α activates EV trafficking programs directing immunomodulatory cargo into discrete vesicle populations. Differences in EV cargo sorting could ultimately be linked to differential beta cell survival in T1D. Disclosure C. Rao: None. M.C. Austin: None. S. Roy: None. A.K. Linnemann: None. J. Piganelli: None. E. Sims: Consultant; Current; Sanofi. Advisory Panel; Current; Diamond, Wink Therapeutics. Funding Breakthrough T1D (3-PDF-2024-1496-A-N) to CR NIDDK (DK133881) to EKS
Rao et al. (Sun,) studied this question.