Purpose: Islets undergo enormous stress during isolation and peri-transplant periods which causes a significant release of stressomes (exosomes released during stress) . Uncovering the biological role of such vesicles is vital for long-term graft function and survival. Here, we sought to identify the global protein content and significance of stressomes isolated during isolation in autologous islet transplantation (AIT) .
Methods: Post-transplant stress was mimicked in vitro by exposing human islets to hypoxia and cytokines (Cyt+Hyp) . Stressome protein contents were characterized by mass spectrometry and immunoblotting. Similar stressomes were also isolated from the islet infusion bag samples of AIT recipients (N=20) . The stressome protein levels were correlated with 1-year AIT outcomes. Finally, these islet stressomes were treated to another set of purified islets and macrophages to find their uptake and immune response.
Results: The mass spectrometry analysis has revealed the islet stressomes contain DAMP proteins like histones (H2B, H3, and H4) . The canonical ER stress signal plays a crucial role in the biogenesis of islet stressomes induced by (Cyt+Hyp) . Exosomes isolated from the infusion bags showed an elevated histone level in the insulin-dependent (ID) group. Moreover, high levels of DAMP proteins were positively correlated with increased post-transplant recipients insulin requirements (p=0.029) . The fluorescent confocal microscopy analysis has revealed these stressomes were easily uptaken by the macrophages and also conferred an inflammatory phenotype. Instead, the stressome treated islets showed elevated expression of innate immune receptors like TLR2 and AGTR1.
Conclusion: The study has identified the islet stressome global protein content and briefs its prospective to induce islet inflammation and graft loss in AIT.
P.Saravanan: None. J.Kalivarathan: None. M.F.Levy: Research Support; Vertex Pharmaceuticals Incorporated, Speaker's Bureau; AbbVie Inc., Sanofi Genzyme. M.A.Kanak: None.