Objective:To investigate the impact of lipotoxic hepatocyte-derived exosome(LTH-ex)on ferroptosis and activation of hepatic stellate cell(HSC).Methods:Lipotoxic damage was induced in the human liver cell line LO2 using oleic acid and palmitic acid. The supernatants of the lipotoxic damaged livercells were collected and LTH-ex was extracted through ultracentrifugation. The characterization of LTH-ex was identified using transmission electron microscopy and Nanosight nanoparticle analysis system, including assessing morphology, particle size and concentration, and LTH-ex surface markers through Western blotting. The human HSC cell line LX-2 was co-cultured with LTH-ex or the ferroptosis activator Erastin for 48 h, and then all cells were divided into three groups: the PBS control group, the LTH-ex treatment group, and the co-treatment group consisting of LTH-ex and the ferroptosis inducer Erastin. Cell viability and mitochondrial membrane potential of LX-2 were evaluated using FDA and JC-1 staining. The iron ion concentration kit was used to detect the iron level in LX-2, the expression of mRNA and protein of death suppressor genes GPX4, xCT, and HSC activation gene α-SMA, were measured through qRT-PCR and Western blotting. Additionally, a mouse model of non-alcoholic fatty liver disease was established by feeding a high-fat diet, followed by injection of LTH-ex through the tail vein. Liver tissue structure was observed using HE staining, while GPX4 and α-SMA protein expression was assessed through immunohistochemistry staining. Sirius red staining was employed to detect collagen deposition in the liver tissue.Results:The LTH-ex particles were approximately 110 nm in size, displaying the typical lipid bilayer structure of exosomes and containing exosome marker proteins TSG101, Alix, and CD63. In comparison to the PBS control group, the LTH-ex treatment group demonstrated enhanced viability of LX-2 cells, an increased mitochondrial membrane potential, decreased iron levels, and upregulation of the ferroptosis-inhibiting genes GPX4 and xCT, as well as HSC activation gene α-SMA. Conversely, when compared to the LTH-ex treatment group, the co-treatment group of LTH-ex and the ferroptosis inducer Erastin exhibited reduced LX-2 cell viability and mitochondrial membrane potential, alongside downregulation of GPX4, xCT, and α-SMA. HE staining, immunohistochemistry, and Sirius red staining demonstrated that LTH-ex could exacerbate vacuolar degeneration of hepatocytes in the liver tissue of mice with non-alcoholic fatty liver disease. Additionally, LTH-ex promoted the expression of GPX4 and α-SMA proteins and increased collagen deposition in the liver tissue. expression.Conclusion:LTH-ex increase the cell viability of hepatic stellate cells and the expression of ferroptosis inhibitory genes GPX4 and xCT, and it may be implicated that LTH-ex may promote hepatic stellate cell activation by inhibiting ferroptosis, and exacerbate collagen deposition in the liver tissue of non-alcoholic fatty liver disease mice.