Involvement of Diclofenac Acyl-β-d-glucuronide in Diclofenac-induced Cytotoxicity in Glutathione-depleted Isolated Murine Hepatocytes Co-cultured with Peritoneal Macrophages

Direct hepatotoxic effects of drugs can occur when a parent drug and/or its reactive metabolites induces the formation of reactive oxygen species. Reactive metabolites of diclofenac (DIC) such as DIC acyl-β-d-glucuronide (DIC-AG) bind covalently to proteins, potentially decreasing protein function or inducing an immune response. However, it is unclear whether the macrophages and GSH depletion participate in DIC-induced cytotoxicity. Mouse hepatocytes (Hep) co-cultured with peritoneal macrophages (PMs) were used to clarify the effects of presence of PM with GSH depletion on DIC-induced cytotoxicity in Hep. DIC-AG but not hydroxy-DIC concentrations in medium were significantly increased in Hep co-cultured with PM with GSH depletion. Depletion of GSH resulted in significantly higher LDH leakage. Interestingly, LDH leakage in Hep/PM (1:0.4) with GSH depletion was significantly higher than in Hep/PM (1:0 and 1:0.1) with BSO. It is likely that macrophages with GSH depletion could facilitate DIC-induced cytotoxicity.