Nano-Encapsulation of Bilirubin in Pluronic F127–Chitosan Improves Uptake in β Cells and Increases Islet Viability and Function after Hypoxic Stress

Pancreatic islet transplantation is the only curative, noninvasive treatment for type 1 diabetes mellitus; however, high rates of cell death in the immediate postimplantation period have limited the success of this procedure. Bilirubin, an endogenous antioxidant, can improve the survival of murine pancreatic allografts during hypoxic stress but has poor solubility in aqueous solutions. We hypothesized that nano-encapsulation of bilirubin in pluronic 127-chitosan nanoparticle bilirubin (nBR) would improve uptake by murine pancreatic islet cells and improve their viability following hypoxic stress. Nano-bilirubin was synthesized, and drug release characteristics were studied in vitro. Cellular uptake of nBR was compared to free bilirubin (fBR) in an insulinoma cell line (INS-R3) model using confocal-like structured illumination microscopy. Next, C57BL/6 mouse islets were treated with concentrations of 0 to 20 μM of nBR, fBR, or empty nanoparticle (eNP), prior to incubation under standard or hypoxic conditions. Islet viability and function were compared between treatment groups. Release of bilirubin was greatest from nBR suspended in protein-rich solution. Increased, selective uptake of nBR by INS-R3 cells was demonstrated. Cell death after hypoxic stress was significantly decreased in murine islets treated with 5 μM nBR (18.5% ± 14.1) compared to untreated islets (33.5% ± 17.5%; P = 0.019), with reduction in central necrosis. Treatment group had a significant effect on glucose stimulation index [SI], ( P = 0.0137) and islets treated with 5 μM nBR had the highest SI overall. Delivery of bilirubin using pluronic F127-chitosan NP improves uptake by murine islets compared to fBR and offers dose-dependent protective effects following hypoxic stress.