An improved understanding of the pathophysiologic mechanisms contributing to insulin deficiency in type 1 diabetes (T1D) is crucial to optimize approaches to disease prevention and treatment. Multiple groups have described increases in beta cell prohormones in individuals at different stages of T1D development, both at the level of the islet and in circulation. These findings have highlighted the concept that dysfunctional beta cell prohormone processing may contribute to insulin deficiency and T1D progression. Because of historical limitations of access to human pancreatic tissue, prior human analyses have been unable to combine analysis of prohormone secretory dynamics with tissue–level phenotyping of prohormone expression and processing. Our preliminary analysis identified heterogeneity in proinsulin secretion among the donors with a history of T1D. Extracellular vesicles (EVs) and their cargo contribute to islet intercellular communication. However, the relationship between the beta cell EV secretome and beta cell prohomone secretion has never been described. For this addendum to the proposal, our overarching hypothesis is that heterogeneity in islet proinsulin release could be associated with differences in islet EV content. We will specifically test EV proinsulin, EV programmed death ligand–1 (PD–L1), and EVs linked to autophagy (LC3+EVs). To test this, we propose analysis of EVs using the remaining perifusate samples already at our possession at IU, using the Exoview platform, with quantification of PD–L1+ and LC3+ positive EVs (absolute and as a percentage of total EVs) as well as bead–based pulldown and permeablization to identify proinsulin+ EVs. These results will determine if heterogeneity islet proinsulin secretion is linked to altered EV composition, yielding critical insight into association of altered prohormone processing and release with insulin deficiency in T1D.
