Imidazole Alkaloids Epiisopilosine and Epiisopiloturine Attenuate Acetaminophen-Induced Liver Toxicity in Mice via Autophagy Modulation and Anti-Inflammatory Effects.

There is increasing interest in natural metabolites, such as alkaloids, due to their potential in treating liver diseases, including acetaminophen (APAP)-induced hepatotoxicity. Alkaloids are known to modulate autophagy, a mechanism associated with liver protection. Epiisopilosine (EPIIS) and epiisopiloturine (EPI), imidazole alkaloids derived from Pilocarpus microphyllus (Jaborandi), exhibit anti-inflammatory and hepatic immunomodulatory effects. Therefore, this study aimed to compare the hepatoprotective and autophagy-modulating effects of EPI and EPIIS in a murine model of APAP-induced hepatotoxicity. In the experimental design, male BALB/c mice received APAP (750 mg/kg, i.p.) to induce hepatotoxicity, followed by phosphate-buffered saline (PBS), N-acetylcysteine (NAC-318 mg/kg, i.p.), or alkaloids (0.3, 1, or 3 mg/kg, i.p.) 30 min later. To assess the involvement of autophagy, hydroxychloroquine (HCQ; 80 mg/kg, i.p.), an autophagy inhibitor, was administered 2 h before APAP. Treatment with EPI, EPIIS, or NAC significantly reduced liver toxicity (p < 0.05). APAP-treated mice exhibited marked centrilobular necrosis, which was markedly reduced following the treatment with Jaborandi alkaloids. Furthermore, EPIIS and EPI significantly reduced the inflammatory and oxidative markers. Administration of HCQ 2 h before APAP abolished these effects. Western blotting analysis revealed increased LC3B expression, a marker of autophagy, in the hepatic tissues of the treated mice, indicating that EPIIS and EPI modulate autophagy. Molecular docking analysis suggested potential interactions between the alkaloids and CXCL10, a chemokine linked to inflammation and autophagy inhibition. These findings demonstrate that EPIIS and EPI protect against APAP-induced hepatotoxicity in mice, potentially by modulating autophagy and reducing inflammation.