Silibinin (silybin) is the major active compound of silymarin used to treat several chronic liver diseases including metabolic dysfunction-associated steatotic liver disease (MASLD). However, the molecular mechanism of hepatic protection offered by silibinin remains still incompletely understood. In this study, we aimed to investigate whether silibinin could ameliorate hepatic steatosis by regulating mitochondrial function in Western diet (WD)-fed MASLD mice and free fatty acid-treated HepG2 cells. WD-fed mice and oleic acid/palmitic acid (OA/PA; 2:1)-treated HepG2 cells were established to evaluate the protection of silibinin against hepatocyte steatosis. Mitochondrial quality was detected using transmission electron microscope, confocal microscope, and cell analyzer for energy metabolism. Silibinin effectively attenuated WD-fed steatotic liver in mice and decreased lipid accumulation in hepatocytes, proved to be associated with stabilization of mitochondrial networked areas and inhibition of mitochondrial swelling. Functionally, silibinin at concentrations of 100 ng/μL was found to enhance mitochondrial respiratory capacity in the OA/PA-treated cells. RNA transcriptome analysis showed that, following silibinin administration, the expressions of numerous mitochondria-associated signaling molecules including AMP-activated protein kinase and mitophagy were upregulated. Among them, optic atrophy (OPA)1 expression increased prominently, which coincided with not only elevated mitochondrial fusion but also declined mitochondrial fragmentation in mouse steatotic livers and OA/PA-treated hepatocytes. In contrast, knockdown OPA1 abolished the protective effect offered by silibinin against lipid accumulation and deteriorated hepatocyte steatosis. Our findings suggest that silibinin attenuating hepatic steatosis is potentially attributed to stabilizing mitochondrial homeostasis via upregulation of OPA1. SIGNIFICANCE STATEMENT: The study results revealed that silibinin maintains mitochondrial homeostasis by upregulating optic atrophy 1 expression in hepatocytes to attenuate lipid accumulation and oxidative stress. Therefore, silibinin is a potential therapeutic candidate for the treatment of metabolic dysfunction-associated steatotic liver disease.
