Acetaminophen-induced Mitochondrial Oxidative Stress in Murine J774.2 Monocyte Macrophages

Abstract

The cytotoxic potential of an antipyretic and analgesic drug, acetaminophen (APAP), was evaluated in mouse J774.2 monocyte macrophages. The cytotoxicity of APAP was evaluated by MTT cell viability and apoptosis assays. Based on the cell viability and apoptosis assays, further experiments were designed with a low (1 µmol/ml) and a high (10 µmol/ml) dose treatment of APAP in J774.2 cells. Mitochondrial oxidative stress, reactive oxygen species (ROS), mitochondrial glutathione (GSH) metabolism, lipid and protein peroxidation were measured in the drug treated cells. An increase in mitochondrial oxidative stress and ROS production was observed. A decrease in the mitochondrial GSH pool, accompanied by an increase in lipid and protein peroxidation appeared to be the main cause of mitochondrial oxidative stress. GSH pool and GSH metabolizing enzymes were differentially affected in the mitochondria and extramitochondrial compartments. Increased nuclear translocation of NF-kB-p65, a marker of redox metabolism was also observed in the drug treated cells. In addition, we have demonstrated, for the first time that the mitochondrial aconitase enzyme is a potential ROS-sensitive target in J774.2 cells, which might be used as a marker for APAP-induced cytotoxicity. These results have clearly suggested that APAP induced cytotoxicity in macrophages is mediated by increased mitochondrial oxidative stress and altered redox metabolism. This might have implications in determining the role of circulating macrophages against APAP induced toxicity and cellular defenses in tissues.