Arrb1 Ameliorates Liver Ischemia/Reperfusion Injury Via Interacting with TRAF6 in Hepatocytes

Background: The hallmarks of hepatic ischemia/reperfusion (I/R) injury, a common clinical problem that occurs during liver surgical procedures, include severe cell death and inflammatory responses that contribute to early graft failure and a higher incidence of organ rejection. Unfortunately, effective therapeutic strategies are limited. β-arrestins are multifunctional proteins that mediate receptor desensitization and serve as important signaling scaffolds in numerous physiopathological processes. However, the role of β-arrestins in hepatic I/R remains largely unknow. Methods: We overexpressed Arrb1 in the hepatocyte of C57BL/6 mice using an Arrb1 adeno-associated virus. Wide-type, Arrb1 overexpressing and knockout mice were subjected to a partial (70%) hepatic ischemia for 60 minutes, followed by various periods of reperfusion. Isolated hepatocytes from different group of mice were subjected to hypoxia-reoxygenation (H/R) injury to determine the in vitro effects of Arrb1. The molecular mechanisms underlying Arrb1-regulatd hepatic I/R injury were further explored and verified in mice. Findings: Here, we showed that only Arrb1 was downregulated during I/R-injured liver injury. Hepatocyte-specific overexpression of Arrb1 significantly ameliorated liver damage, as demonstrated by decreases in serum aminotransferases, hepatocellular necrosis and apoptosis, infiltrating inflammatory cells and secretion of pro-inflammatory cytokines relative to control mice. Whereas, experiments with Arrb1 knockout mice reveled opposite effects. Mechanistically, Arrb1 directly interacts with TRAF6 in hepatocyte, and inhibits the ubiquitination of TRAF6, which then prevents the activation of the downstream MAPK and NF-κB signaling pathways during hepatic I/R injury. Interpretation: The Arrb1-TRAF6 interaction inhibited the ubiquitination of TRAF6 and the activation of MAPK and NF-κB signaling, and then protected against cell death and inflammation, suggesting Arrb1 is a potential therapeutic target for hepatic I/R injury. Funding: This research was supported by grants from the 481 National Key Research and Development Program of China (2016YFC0905900), The State Key Program of National Natural Science Foundation of China (81430062), and the Innovative Research Groups of the National Natural Science Foundation (81521004 to B.S.). The authors declare no conflicts of interest. Declaration of Interest: The authors declare no potential conflicts of interest. Ethical Approval: Animal protocols were approved by the Animal Care and Use Committee of Nanjing Medical University, and experiments were conducted in adherence with the Guide for the Care and Use of Laboratory Animals (National Institutes of Health, Bethesda, MD, USA). We gained permission from all the patients and healthy volunteers, and the procedures of our study on the basis of the basic principles of Helsinki Declaration were approved by the Ethics Committee of Nanjing Drum Tower Hospital.