Despite the utility of cyclooxygenase (COX) inhibition as an antiinflammatory strategy, prostaglandin (PG) products of COX-1 and -2 provide important regulatory functions in some pathophysiological states. Scattered reports suggest that COX inhibition may also promote adverse drug events. Here we demonstrate a protective role for endogenous COX-derived products in a murine model of acetaminophen (APAP)-induced acute liver injury. A single hepatotoxic dose caused the selective induction of COX-2 mRNA and increased PGD2 and PGE2 levels within the livers of COX^+/+ male mice suggesting a role for COX-2 in this model of liver injury. APAP-induced hepatotoxicity and lethality were markedly greater in COX-2^-/- and ^-/+ mice in which normal PG responsiveness is altered. The significantly increased toxicity linked to COX-2 deficiency could be mimicked using the selective COX-2 inhibitory drug, celecoxib, in COX^+/+ mice and was not due to alterations in drug−protein adduct formation, a surrogate for bioactivation and toxicity. Microarray analyses indicated that increased injury associated with COX-2 deficiency coincided, most notably, with a profoundly impaired induction of heat shock proteins in COX-2^-/+ mice suggesting that PGs may act as critical endogenous stress signals following drug insult. These findings suggest that COX-2-derived mediators serve an important hepato-protective function and that COX inhibition may contribute to the risk of drug-induced liver injury, possibly through both nonimmunological and immunological pathways.