Synergistic hepatotoxicity of N,N-dimethylformamide with carbon tetrachloride in association with endoplasmic reticulum stress

N,N-Dimethylformamide (DMF) is an organic solvent extensively used in industries such as synthetic leather, fibers and films, and induces liver toxicity and carcinogenesis. Despite a series of experimental and clinical reports on DMF-induced liver failure, the mechanism of toxicity is yet unclear. This study investigated whether DMF in combination with a low dose of hepatotoxicant enhances hepatotoxicity, and if so, on what mechanistic basis. Treatment of rats with either DMF (50–500 mg/kg/day, for 3 days) or a single low dose of CCl₄ (0.2 ml/kg) alone caused small increases in plasma transaminases and lactate dehydrogenase activities. However, combinatorial treatment of DMF with CCl₄ markedly increased blood biochemical changes. Histopathology confirmed the synergism in hepatotoxicity. Moreover, DMF + CCl₄ caused PARP cleavage and caspase-3 activation, but decreased the level of Bcl-xL, all of which confirmed apoptosis of hepatocytes. Consistently, DMF + CCl₄ treatment markedly increased lipid peroxidation. By contrast, treatment of DMF in combination with lipopolysaccharide, acetaminophen or d-galactosamine caused no enhanced hepatotoxicity. Given the link between endoplasmic reticulum (ER) dysfunction and cell death, ER stress response was monitored after DMF and/or CCl₄ treatment. Whereas either DMF or CCl₄ treatment alone marginally changed the expression levels of glucose-regulated protein 78 and 94 and phosphorylated PKR-like ER-localized eIF2α kinase, concomitant treatment with DMF and CCl₄ synergistically induced them with increases in glucose-regulated protein 78 and C/EBP homologous protein mRNAs. Our results demonstrate that DMF treatment in combination with CCl₄ synergistically increases hepatocyte death, which may be associated with the induction of severe ER stress.