1779-P: Lipotoxicity-Induced Beta-Cell Small Extracellular Vesicles Promote Beta-Cell Functional Failure through Activation of the TGFß/Smad3 Pathway

Lipotoxicity is a pathological process that occurs in several metabolic disorders including obesity-driven Type 2 diabetes (T2D). Chronically elevated circulating free fatty acids have been shown to exert detrimental effects on pancreatic islet β-cells through reduced glucose-stimulated insulin secretion (GSIS), alterations in β-cell transcriptional identity, and apoptosis. β-cell-derived small extracellular vesicles (sEV) have been shown to contribute to β-cell failure in T2D, however their role in lipotoxicity-mediated β-cell failure has yet to be elucidated. We hypothesize that lipotoxicity-induced β-cell functional failure is mediated in part by aberrant release of altered β-cell-derived sEV. To test this, MIN6 β-cells were exposed to free fatty acid, palmitate (PAL) (0.5 mM, 24h) and EV were isolated using differential ultracentrifugation to yield PAL EV (vs. control (CTL) EV). β-cell functional assessment using GSIS on mouse islets exposed to PAL EV (48h) resulted in significant suppression of GSIS (~80% decrease in stimulation index). RNA-Seq was conducted on islets exposed to PAL EV (vs. untreated (UT)). De novo motif analysis from this data identified enriched transcription factor binding motifs associated with several SMAD genes in TGFβ signaling. Gene Set Enrichment Analysis revealed significant enrichment of genes associated with TGFβ receptor binding suggesting its potential role in lipotoxic β-cell EV-mediated β-cell failure. To confirm, analysis of p-Smad3 expression was conducted by immunofluorescence staining on mouse islets exposed to PAL EV (48h; vs. UT). The results showed an ~60-fold increase in β-cell p-Smad3 expression (p<.05). Moreover, mouse islets were treated with a TGFβ receptor inhibitor in the presence of PAL EV (48h; vs. UT). GSIS results showed a significant improvement in β-cell function (p<.05) suggesting the role of the TGFβ/Smad3 pathway in lipotoxic β-cell sEV induced β-cell failure. Disclosure A.Roy: None. A.Hoff: None. T.K.Her: None. K.Sankaran rajagopalan: None. D.D.Maltais: None. A.Matveyenko: None. N.Javeed: None. Funding National Institute of Diabetes and Digestive and Kidney Diseases (DK129208-01); JDRF (2-SRA-2022-1272-S-B)