Cantharidin Causes Hepatotoxicity by Disrupting GSH‐Mediated Redox Homeostasis

Abstract

ABSTRACT Cantharidin (CTD) possesses potent anticancer activity, whereas its prominent hepatotoxicity severely limits its clinical application. Because the detailed molecular mechanisms responsible for CTD‐triggered hepatotoxicity have not been fully elucidated, additional investigations are necessary to uncover its toxicological pathways. This study investigated the mechanism of CTD‐triggered hepatotoxicity and the protective effects of exogenous glutathione (GSH) supplementation. In vitro results showed that CTD reduced the GSH/glutathione disulfide (GSSG) ratio and the activities of catalase (CAT) and glutathione peroxidase (GPX) in hepatocytes, induced a compensatory upregulation of superoxide dismutase (SOD), and promoted malondialdehyde (MDA) accumulation. Additionally, CTD downregulated glutamate‐cysteine ligase modifier subunit (GCLM) expression, decreased mitochondrial membrane potential (MMP), suppressed B‐cell lymphoma 2 (Bcl‐2) expression, and upregulated Bcl‐2‐associated X protein (Bax), cysteine‐dependent aspartate‐directed protease (caspase‐9), downstream caspase‐3, and cytochrome c (Cyt c) levels, thereby triggering hepatocyte apoptosis, with no significant changes in cellular cysteine (Cys) content and glutamate‐cysteine ligase catalytic subunit (GCLC) expression. In vivo experiments verified that CTD markedly decreased hepatic GSH and Cys levels and downregulated GCLC expression, whereas GCLM expression and hepatic Cyt c levels showed only a slight upward trend without statistical significance. Glutathione ethyl ester (GSH‐OEt) pretreatment in vitro and GSH intervention in vivo effectively prevented CTD‐caused oxidative damage. Collectively, CTD induces divergent expression patterns of glutamate‐cysteine ligase (GCL) subunits in vitro and in vivo, disrupts GSH metabolic homeostasis, and provokes oxidative stress and mitochondrial dysfunction, ultimately leading to hepatocellular injury.