Toosendanin Induces Hepatocyte Damage by Inhibiting Autophagic Flux via TFEB-Mediated Lysosomal Dysfunction

Toosendanin (TSN) is a triterpenoid from the fruit or bark of Melia toosendan Sieb et Zucc, which has clear antitumor and insecticidal activities, but it possesses limiting hepatotoxicity in clinical application. Autophagy is a degradation and recycling mechanism to maintain cellular homeostasis, and it also plays an essential role in TSN-induced hepatotoxicity. Nevertheless, the specific mechanism of TSN on autophagy-related hepatotoxicity is still unknown. The hepatotoxicity of TSN in vivo and in vitro was explored in this study. It was found that TSN induced the upregulation of the autophagy-marker microtubule-associated proteins 1A/1B light chain 3B (LC3B) and P62, the accumulation of autolysosomes, and the inhibition of autophagic flux. The middle and late stages of autophagy were mainly studied. The data showed that TSN did not affect the fusion of autophagosomes and lysosomes but significantly inhibited the acidity, the degradation capacity of lysosomes, and the expression of hydrolase cathepsin B (CTSB). The activation of autophagy could alleviate TSN-induced hepatocyte damage. TSN inhibited the expression of transcription factor EB (TFEB), which is a key transcription factor for many genes of autophagy and lysosomes, such as CTSB, and overexpression of TFEB alleviated the autophagic flux blockade caused by TSN. In summary, TSN caused hepatotoxicity by inhibiting TFEB-lysosome-mediated autophagic flux and activating autophagy by rapamycin (Rapa), which could effectively alleviate TSN-induced hepatotoxicity, indicating that targeting autophagy is a new strategy to intervene in the hepatotoxicity of TSN.