Inflammatory and hypoxic stress-induced islet exosomes released during isolation are associated with poor transplant outcomes in islet autotransplantation

Abstract

Islets experience enormous stress during the isolation process, leading to suboptimal endocrine function after total pancreatectomy with islet autotransplantation (TPIAT). Our investigation focused on inducing isolation stress in islets ex vivo, where proinflammatory cytokines and hypoxia prompted the release of stress exosomes (exo^S) sized between 50 and 200 nm. Mass spectrometry analysis revealed 3 distinct subgroups of immunogenic proteins within these exo^S: damage-associated molecular patterns (DAMPs), chaperones, and autoantigens. The involvement of endosomal-sorting complex required for transport proteinsincluding ras-associated binding proteins7A, ras-associated binding protein GGTA, vacuolar protein sorting associated protein 45, vacuolar protein sorting associated protein 26B, and the tetraspanins CD9 and CD63, in exo^S biogenesis was confirmed through immunoblotting. Next, we isolated similar exo^S from the islet infusion bags of TPIAT recipients (N = 20). The exosomes from infusion bags exhibited higher DAMP (heat shock protein family A [Hsp70] member 1B and histone H2B) levels, particularly in the insulin-dependent TPIAT group. Additionally, elevated DAMP protein levels in islet infusion bag exosomes correlated with increased insulin requirements (P = .010) and higher hemoglobin A1c levels 1-year posttransplant. A deeper exploration into exo^S functionality revealed their potential to activate monocytes via the toll-like receptor 3/7: DAMP axis. This stimulation resulted in the induction of inflammatory phenotypes marked by increased levels of CD68, CD80, inducible nitric oxide synthase, and cyclooxygenase-2. This activation mechanism may impact the successful engraftment of transplanted islets.

Keywords: DAMPs; TPIAT; cytokines; exosomes; hypoxia; innate immunity; islets; monocytes; stress; toll-like receptors; transplantation.