Recommendations for the use of the acetaminophen hepatotoxicity model for mechanistic studies and how to avoid common pitfalls

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.

Mitochondrial damage and biogenesis in acetaminophen-induced liver injury

Liver injury and acute liver failure caused by acetaminophen(APAP)overdose is the clinically most important drug toxicity in Western countries.Mechanistic investigations have revealed a central role of mitochondria in the pathophysiology.Excess formation of the reactive metabolite N-acetyl-p-benzoquinone imine(NAPQI)after an overdose leads to hepatic glutathione depletion,mitochondrial protein adducts formation and an initial oxidant stress,which triggers the activation of mitogen activated protein(MAP)kinase cascade ultimately leading to c-jun N-terminal kinase(JNK)phosphorylation.Phospho-JNK translocates to the mitochondria and amplifies the oxidative and nitrosative stress eventually causing the mitochondrial membrane permeability transition pore opening and cessation of adenosine triphosphate(ATP)synthesis.In addition,mitochondrial matrix swelling ruptures the outer membrane and releases endonucleases,which cause nuclear deoxyribonucleic acid(DNA)fragmentation.Together,the nuclear DNA damage and the extensive mitochondrial dysfunction result in necrotic cell death.However,the procell death signaling events are counteracted by adaptive responses such as autophagy and mitochondrial biogenesis.The improved mechanistic insight into the pathophysiology leads to better understanding of the mechanisms of action of the existing antidote N-acetylcysteine and justifies the clinical testing of novel therapeutics such as 4-methylpyrazole and calmangafodipir.