Paeoniflorin Ameliorates Liver Fibrosis by Inhibiting HIF ‐1α‐Mediated Mitophagy in Hepatic Stellate Cells

ABSTRACT Paeoniflorin (PF) has exhibited significant anti‐hepatic fibrosis potential, yet its precise pharmacological mechanisms and molecular targets remain to be fully elucidated. This study aims to investigate the role of PF in modulating HIF‐1α‐mediated mitophagy both in vivo and in vitro, thereby elucidating its impact on liver fibrosis. The rat model of hepatic toxicity and chronic inflammation was established via CCl 4 injection, followed by preventive administration of PF in three dosage tiers (100, 150, 200 mg/kg/d). Additionally, HSC‐T6 cells were subjected to exposure to cobalt chloride (CoCl 2 , 200 μM) to mimic hypoxic conditions. Lentiviral transfection was employed to establish stable cell lines with either overexpression or knockdown of HIF‐1α, aiming to delineate the functional significance of HIF‐1α in hepatic stellate cells (HSCs) activation. The severity of liver inflammatory response and fibrotic progression was evaluated with several pathological stainings. Levels of mRNA were detected by qPCR. Expression levels of protein were assayed with Western blot. The protein content and distribution were also visually detected through IHC and IF staining. ROS was detected by DCFH‐DA, and mitochondrial membrane potential was quantitatively assessed using JC‐1 staining. Our results demonstrated that PF treatment significantly reversed CCl 4 ‐induced liver fibrosis, effectively reducing hepatic inflammation and oxidative damage. PF also inhibited mitophagy by suppressing the HIF‐1α pathway, thereby attenuating HSCs' activation. This study uncovers a novel mechanism involving HIF‐1α‐mediated mitophagy in HSCs and provides evidence that PF alleviates hepatic fibrosis by inhibiting this process, leading to reduced inflammation and oxidative damage. These findings suggest that PF could be a natural and vital medicine to treat liver fibrosis.