Acetaminophen (APAP) is a widely used analgesic and antipyretic drug, which is safe at therapeutic doses but can cause severe liver injury and even liver failure after overdoses. The mouse model of APAP hepatotoxicity recapitulates closely the human pathophysiology. As a result, this clinically relevant model is frequently used to study mechanisms of drug-induced liver injury and even more so to test potential therapeutic interventions. However, the complexity of the model requires a thorough understanding of the pathophysiology to obtain valid results and mechanistic information that is translatable to the clinic. However, many studies using this model are flawed, which jeopardizes the scientific and clinical relevance. The purpose of this review is to provide a framework of the model where mechanistically sound and clinically relevant data can be obtained. The discussion provides insight into the injury mechanisms and how to study it including the critical roles of drug metabolism, mitochondrial dysfunction, necrotic cell death, autophagy and the sterile inflammatory response. In addition, the most frequently made mistakes when using this model are discussed. Thus, considering these recommendations when studying APAP hepatotoxicity will facilitate the discovery of more clinically relevant interventions.

Acetaminophen (APAP) overdose can induce liver injury and is the most frequent cause of acute liver failure in the United States. We investigated the role of p62/SQSTM1 (referred to as p62) in APAP-induced liver injury (AILI) in mice. We found that the hepatic protein levels of p62 dramatically increased at 24 h after APAP treatment, which was inversely correlated with the hepatic levels of APAP-adducts. APAP also activated mTOR at 24 h, which is associated with increased cell proliferation. In contrast, p62 knockout (KO) mice showed increased hepatic levels of APAP-adducts detected by a specific antibody using Western blot analysis but decreased mTOR activation and cell proliferation with aggravated liver injury at 24 h after APAP treatment. Surprisingly, p62 KO mice recovered from AILI whereas the wild-type mice still sustained liver injury at 48 h. We found increased number of infiltrated macrophages in p62 KO mice that were accompanied with decreased hepatic von Willebrand factor (VWF) and platelet aggregation, which are associated with increased cell proliferation and improved liver injury at 48 h after APAP treatment. Our data indicate that p62 inhibits the late injury phase of AILI by increasing autophagic selective removal of APAP-adducts and mitochondria but impairs the recovery phase of AILI likely by enhancing hepatic blood coagulation.

Background Acetaminophen (APAP) overdose can cause liver injury and liver failure, which is one of the most common causes of drug-induced liver injury in the United States. Pharmacological activation of autophagy by inhibiting mechanistic target of rapamycin (mTOR) protects against APAP-induced liver injury likely via autophagic removal of APAP-adducts and damaged mitochondria. In the present study, we aimed to investigate the role of genetic ablation of mTOR pathways in mouse liver in APAP-induced liver injury and liver repair/regeneration. Methods Albumin-Cre (Alb-Cre) mice, mTORf/f and Raptorf/f mice (C57BL/6J background) were crossbred to produce liver-specific mTOR knockout (L-mTOR KO, Alb Cre+/-, mTORf/f) and liver-specific Raptor KO (L-Raptor, Alb Cre+/-, Raptor f/f) mice. Alb-Cre littermates were used as wild-type (WT) mice. These mice were treated with APAP for various time points for up to 48 h. Liver injury, cell proliferation, autophagy and mTOR activation were determined. Results We found that genetic deletion of neither Raptor, an important adaptor protein in mTOR complex 1, nor mTOR, in the mouse liver significantly protected against APAP-induced liver injury despite increased hepatic autophagic flux. Genetic deletion of Raptor or mTOR in mouse livers did not affect APAP metabolism and APAP-induced c-Jun N-terminal kinase (JNK) activation, but slightly improved mouse survival likely due to increased hepatocyte proliferation. Conclusions Our results indicate that genetic ablation of mTOR in mouse livers does not protect against APAP-induced liver injury but may slightly improve liver regeneration and mouse survival after APAP overdose.