AbstractBackground & AimsAcetaminophen overdose in mice is characterized by hepatocyte endoplasmic reticulum stress, which activates the unfolded protein response, and centrilobular hepatocyte death. We aimed at investigating the therapeutic potential of tauroursodeoxycholic acid, a hydrophilic bile acid known to have anti‐apoptotic and endoplasmic reticulum stress‐reducing capacities, in experimental acute liver injury induced by acetaminophen overdose.MethodsMice were injected with 300 mg/kg acetaminophen, 2 hours prior to receiving tauroursodeoxycholic acid, N‐acetylcysteine or a combination therapy, and were euthanized 24 hours later. Liver damage was assessed by serum transaminases, liver histology, terminal deoxynucleotidyl transferase dUTP nick end labelling staining, expression profiling of inflammatory, oxidative stress, unfolded protein response, apoptotic and pyroptotic markers.ResultsAcetaminophen overdose resulted in a significant increase in serum transaminases, hepatocyte cell death, unfolded protein response activation, oxidative stress,NLRP3 inflammasome activation, caspase 1 and pro‐inflammatory cytokine expressions. Standard of care, N‐acetylcysteine and, to a lesser extent, tauroursodeoxycholic treatment were associated with significantly lower transaminase levels, hepatocyte death, unfolded protein response activation, oxidative stress markers, caspase 1 expression andNLRP3 levels. Importantly, the combination of N‐acetylcysteine and tauroursodeoxycholic acid improved serum transaminase levels, reduced histopathological liver damage,UPR‐activatedCHOP, oxidative stress, caspase 1 expression,NLRP3 levels,IL‐1β levels and the expression of pro‐inflammatory cytokines and this to a greater extend than N‐acetylcysteine alone.ConclusionsThese findings indicate that a combination strategy of N‐acetylcysteine and tauroursodeoxycholic acid surpasses the standard of care in acetaminophen‐induced liver injury in mice and might represent an attractive therapeutic opportunity for acetaminophen‐intoxicated patients.