Investigation of rifampicin-induced hepatotoxicity in rat hepatocytes maintained in gel entrapment culture.

Rifampicin-induced hepatotoxicity has been well recognized in animals and patients. However, it is undetectable in cultured hepatocyte monolayers in vitro at the equivalent toxic concentration in vivo. This study investigated the rifampicin-induced toxicity on rat hepatocytes in gel entrapment vs. in monolayer culture. Thiazolyl tetrazolium reduction and albumin secretion were routinely detected to identify the toxic responses of rat hepatocytes to rifampicin, while reactive oxygen species (ROS) accumulation and intracellular glutathione (GSH) content were assayed as biomarkers of oxidative stress. In addition, Nile red staining and malondialdehyde (MDA) generation were, respectively, used as endpoints for lipid accumulation and peroxidation. After treatment of hepatocytes for 96 h at a serum rifampicin concentration (12 microM), gel-entrapped rat hepatocytes showed significant cellular damage indicated by alternations of all parameters indicated above, while hepatocyte monolayers did not show severe responses. In contrast to a lack of protections by cytochrome P 450 inhibitors, the ROS scavenger (glycyrrhizic acid) and thiol compounds (N-acetylcysteine and GSH) significantly reduced rifampicin toxicity in gel-entrapped hepatocytes. It appears that gel-entrapped rat hepatocytes reflected significant hepatotoxicity of rifampicin in vivo, and this toxicity was most possibly associated with oxidative stress and lipid accumulation.