Isoliensinine from Plumula Nelumbinis Attenuates Hepatic Fibrosis in Metabolic Associated Fatty Liver Disease through Regulating Lipid Droplet Metabolism

• Isoliensinine, from the food - medicine homologous Plumula Nelumbinis, improves MAFLD - related fibrosis in vivo. • In vitro, Isoliensinine inhibits hepatic stellate cells (HSCs) activation in a dose - dependent manner, restoring lipid droplets in HSCs. • Isoliensinine activates TRPV1, enhancing its co - localization with the endoplasmic reticulum and restoring calcium homeostasis in HSCs. • Isoliensinine activates the TRPV1 - AMPK pathway, which is essential for restoring lipid droplets and inhibiting HSCs activation in MAFLD - related fibrosis. Metabolic associated fatty liver disease (MAFLD) is a multifactorial disorder driving liver fibrosis progression. Hepatic stellate cell (HSC) activation represents a central event in fibrogenesis, with evidence indicating that replenishing lipid droplets suppresses HSCs activation. Although clinical studies suggest isoliensinine from Plumula Nelumbinis ameliorates lipid metabolism, its role in liver fibrosis remains unexplored. In this study, methionine-choline-deficient diet- induced and Western diet combined with carbon tetrachloride injection-induced MAFLD model were established. Isoliensinine (10/20/40 mg/kg) were given by oral administration, and the possible effect on liver fibrosis were assessed in vivo. The underlying mechanisms were validated in LX2 cells in vitro. And the experiments employing molecular biology methods including Western blot, PCR, CESTA and immunofluorescence were used in vivo and in vitro. Results demonstrated that isoliensinine exerts anti-fibrotic effects in MAFLD models both in vitro and in vivo. Mechanically, isoliensinine significantly upregulated the expression of TRPV1 and activated AMPK/ACC signaling pathway to enhance Ca 2+ homeostasis in activated HSC-LX2, which ultimately promoted lipid droplet replenishment and suppress HSCs activation to result in the attenuate of MAFLD and liver fibrosis.