Fisetinidin chloride ameliorates carbon tetrachloride-induced hepatotoxicity in HepaRG cells

Liver diseases poses a significant global health burden. This study investigated the hepatoprotective effects of fisetinidin chloride (FC), a natural flavonoid, against carbon tetrachloride (CCl₄)-induced hepatotoxicity in HepaRG cells, a model mimicking human hepatocyte responses. Co-treatment with FC restored cell viability and reduced cellular steatosis, and minimized lactate dehydrogenase leakage, demonstrating membrane stabilization. FC mitigated oxidative stress by reducing mitochondrial reactive oxygen species (ROS) and lipid peroxidation, while enhancing antioxidant defenses through upregulating mitochondrial superoxide dismutase and glutathione. FC preserved mitochondrial function, as evidenced by restored mitochondrial membrane potential (ΔΨm), and modulated apoptosis by upregulating anti-apoptotic BCL2 mRNA and downregulating pro-apoptotic BAX and caspase-3. Flow cytometry analysis confirmed FC's anti-apoptotic effects, reducing apoptotic cell populations. Additionally, FC attenuated CCl₄-induced elevations in aspartate aminotransferase and alanine aminotransferase, markers of hepatocellular injury. Treatment with FC significantly upregulated choline, citric acid, cis-aconitic acid, L-carnitine, L-tryptophan, and gamma-Linolenic acid in CCl₄-induced cells. Conversely, it significantly downregulated glutamate, xanthine, indole acetic acid, succinic acid, hypotaurine, and other metabolites. Pathway enrichment and network analysis of the metabolome demonstrated that FC's protective effects were mediated through the modulation of mitochondrial energy metabolism. Collectively, these findings highlight FC's multifaceted hepatoprotective effects, including attenuation of cellular steatosis, ROS scavenging, mitochondrial stabilization, and apoptosis inhibition. This study underscores FC's potential as a therapeutic candidate worthy of further mechanistic studies, bridging in vitro efficacy with clinical relevance. Further in vivo studies are warranted to validate its pharmacokinetics and translational potential.