The receptor interacting protein kinase 3 is a critical early mediator of acetaminophen-induced hepatocyte necrosis in mice

Acetaminophen (APAP) overdose is a major cause of hepatotoxicity and acute liver failure in the US, but the pathophysiology is incompletely understood. Despite evidence for apoptotic signaling, hepatic cell death after APAP is generally considered necrotic in mice and in humans. Recent findings suggest that the receptor interacting protein kinase 3 (RIP3) acts as a switch from apoptosis to necrosis (programmed necrosis). Thus, the aim of the current investigation was to determine if RIP3 is involved in APAP-induced liver cell death. APAP (200-300 mg/kg) caused glutathione depletion and protein adduct formation, oxidant stress, mitochondrial release of apoptosis inducing factor, and nuclear DNA fragmentation resulting in centrilobular necrosis in C57Bl/6J mice. Inhibiting RIP3 protein induction with anti-sense morpholinos in wild-type animals or using RIP3-deficient mice had no effect on protein adduct formation but attenuated all other parameters, including necrotic cell death, at 6h after APAP. In addition, cultured hepatocytes from RIP3-deficient mice showed reduced injury compared to wild-type cells after 24h. Interestingly, APAP-induced mitochondrial translocation of dynamin-related protein 1 (Drp1), the initiator of mitochondrial fission, was inhibited by reduced RIP3 protein expression and the Drp1 inhibitor MDIVI reduced APAP-induced cell death at 24h. All of these protective effects were lost after 24h in vivo or 48h in vitro. Conclusion: RIP3 is anearly mediator of APAP hepatotoxicity, involving modulation of mitochondrial dysfunction and oxidant stress. Controlling RIP3 expression could be a promising new approach to reduce APAP-induced liver injury, but requires complementary strategies to control mitochondrial dysfunction for long-term protection.