[Mechanism of triptolide-induced hepatic cell lipotoxicity based on lipophagy]

This study aimed to investigate the effects of triptolide on lipophagy and to explore the role and underlying mechanism of lipophagy in triptolide-induced hepatocellular injury. HL7702 human normal hepatocytes and C57BL/6J mice were treated with 25, 50, or 100 nmol·L~(-1 )or 0.4, 0.6, or 0.8 mg·kg~(-1) triptolide, respectively. Additionally, HL7702 cells were co-treated with Ras-related protein 7(Rab7) small interfering RNA(siRNA) and 50 nmol·L~(-1) triptolide. Cell viability was assessed by calcein AM/PI double staining, and liver histopathological changes were observed using hematoxylin-eosin(HE) staining. Lipid droplet content was examined via oil red O and nile red staining. Commercial kits were used to measure the content of triglycerides(TG), total cholesterol(TC), free fatty acids(FFAs), glutathione(GSH), and malondialdehyde(MDA), the activities of alanine aminotransferase(ALT) and aspartate aminotransferase(AST), and the levels of mitochondrial membrane potential(MMP) and intracellular reactive oxygen species(ROS). The expression of microtubule-associated protein 1 light chain 3Ⅱ(LC3Ⅱ), selective autophagy adaptor p62, Rab7, and perilipin 2(Plin2) were detected by qRT-PCR or Western blot. The results showed that in vitro, 25,50 and 100 nmol·L~(-1) triptolide significantly reduced the cell survival rate, upregulated the expression of Rab7 and LC3Ⅱ, downregulated the expression of p62 and Plin2, reduced the content of TG, TC, GSH and MMP, and increased the content of FFAs, MDA and ROS. In vivo, 0.4,0.6 and 0.8 mg·kg~(-1) triptolide increased the activity of ALT and AST in serum, caused liver tissue damage, upregulated the expression of Rab7 and LC3Ⅱ, downregulated the expression of p62, decreased the content of TG, TC and GSH, and increased the content of FFAs and MDA. Rab7 siRNA significantly increased the levels of TG, TC and MMP, decreased the levels of FFAs and ROS, and increased the cell survival rate. In conclusion, triptolide at certain concentrations may induce Rab7-mediated lipophagy to promote lipid droplet degradation, thereby causing hepatocellular lipotoxicity, leading to mitochondrial dysfunction and oxidative stress.