Bilirubin Nanoparticles Protect against Kidney Ischemia-Reperfusion Injury through Antioxidant Activity

Background: Renal ischemia-reperfusion injury (IRI) is a common complication but there was no definite treatment, except supportive care. Basic mechanisms of renal IRI is acute inflammation, where oxidative stress plays an important role. Despite potent reactive oxygen species (ROS)-scavenging effects, free bilirubin has an important limitation of water-insolubility, interfering with clinical application. Recently, Brixelle, a PEGylated bilirubin nanoparticle has been developed to improve water solubility. Methods: This study aimed to assess renoprotective effects of bilirubin nanoparticles against renal IRI. We injected bilirubin nanoparticles intravenousely 1 h before and 1.5 h after mouse renal IRI and assessed their renoprotective effects on renal IRI on day 1, 3, and 28, compared to free bilirubin. Results: Bilirubin nanoparticles suppressed in vitro ROS generation and secretion of TNF-α and IL-1β from activated neutrophil-like cells and macrophages, whereas free bilirubin did not show any suppressive effect. Moreover, Bilirubin nanoparticles increased in vitro cellular viability of renal tubular epithelial cells against ROS. Intravenous administration of bilirubin nanoparticles led to an increased uptake in the kidney with minimal migration to brain after renal IRI. Peri-IRI administration of bilirubin nanoparticles improved renal function and attenuated renal tissue injury as well as renal tubular apoptosis on day 1 after IRI to greater extent than free bilirubin. Renal infiltration of neutrophils and expression of TNF-α and MCP-1 were also suppressed by bilirubin nanoparticles. Furthermore, bilirubin nanoparticles increased renal tubular regeneration (Ki67) on day 3 and suppressed renal fibrosis as well as renal expression of αSMA, fibronectin, and type IV collagen on day 28 after IRI to greater extent than free bilirubin. Bilirubin nanoparticles suppressed expression of nicotinamide adenine dinucleotide phosphate oxidase (NOX) and iNOS, increased expression of nuclear factor erythroid-2-related factor 2 (Nrf2) and HO-1, and attenuated ROS generation after IRI to greater extent than free bilirubin. Conclusion: Bilirubin nanoparticles effectively attenuates ROS generation and acute renal injury, facilitates subacute renal recovery, and suppressed chronic fibrosis after renal IRI, having a potential as a new therapy for renal IRI. Funding: Government Support – Non-U.S.