Elucidation of Cortex Dictamni hepatotoxicity mechanism by integrated serum pharmacochemistry and metabolomics

OBJECTIVE: To provide a panoramic analysis of Cortex Dictamni (DIC) and to elucidate the potential toxic components and mechanism of action of DIC-induced hepatotoxicity. METHODS: Liquid chromatography‒tandem mass spectrometry (LC‒MS/MS) was used to identify serum components, and network toxicology, molecular docking and molecular dynamics simulations were combined to identify potentially toxic components and targets. In vivo experiments were performed using rats, which were divided into a control group (Con group) and groups treated with DIC-l (0.9 g/kg), DIC-M (2.7 g/kg), DICH (8.1 g/kg), and isoniazid (INH) (100 mg/kg) via gavage for 14 d Biochemical indices, hepatic tissue pathology, hepatic function parameters, and apoptosis markers were systematically evaluated to assess hepatic toxicity. Hepatotoxicity was also assessed by metabolomics analysis. The changes in the serum metabolic profiles during the administration period and after drug withdrawal were analyzed by metabolomics to determine the mechanism of metabolic pathway perturbation in DIC-induced liver injury. Finally, BRL 3A cells were used to verify the in vitro toxicity of DIC, and the mechanism of DIC hepatotoxicity was comprehensively explained by a combination of in vivo and in vitro experiments. RESULTS: Thirty-four prototypical components, including alkaloids, terpenoids, flavonoids and other compounds, were successfully identified from DIC. Through network toxicology analysis, molecular docking and molecular dynamics simulation, dictamnine and fraxinellone were identified as potential key components mediating the hepatotoxicity of DIC; this toxicity may be mediated through the PI3K/AKT signaling pathway. Animal experimental data revealed that after 14 d of treatment, the rats in all the dosage groups presented different degrees of hepatic injury, and the serum levels of AST, ALT and ALP were significantly increased, confirming the hepatotoxicity of DIC. Metabolomic studies further revealed that DIC induced liver injury by interfering with bile acid metabolism, lipid metabolism and sphingolipid metabolism. Mechanistic validation experiments revealed that dictamnine and fraxinellone activated the PI3K/AKT signaling pathway and promoted the apoptosis of hepatocytes, thus mediating the hepatotoxicity. CONCLUSION: This study is the first to analyze the components of DIC using serum medicinal chemistry to identify the main toxicants and to elucidate the potential molecular mechanism of DIC-induced hepatotoxicity. The results of this study provide an important theoretical basis for further investigations of the mechanism of DIC toxicity and the development of toxicity reduction strategies. These results are also important for promoting the safe use of DIC and the modernization of traditional Chinese medicine.