Abstract

Background Cholestatic liver diseases, such as primary biliary cholangitis (PBC), primary sclerosing cholangitis (PSC), and biliary atresia (BA), are characterized by bile accumulation and frequently progress to liver fibrosis, cirrhosis, and organ failure. Objective Given its well-documented hepatoprotective and anti-inflammatory properties, this study aimed to investigate the therapeutic potential and underlying mechanisms of gastrodin (4-(hydroxymethyl) phenyl β-D-glucopyranoside, C 13 H 18 O 7 ), a primary bioactive compound from Gastrodia elata , for the treatment of cholestatic liver fibrosis. Methods Two established mouse models of cholestatic fibrosis were used: the 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet model and the bile duct ligation (BDL) model. An integrated analysis of single-nucleus RNA sequencing (snRNA-seq) data was performed, combining an in-house dataset from the DDC model with a public dataset. Results Gastrodin treatment significantly alleviated liver injury, reduced serum levels of total bilirubin (TBIL) and total bile acids (TBA), decreased collagen deposition, and prolonged survival in mouse models. snRNA-seq analysis revealed that gastrodin promoted hepatocyte repair by regulating key sulfotransferases (Sult2a1 and Sult1e1), enhanced detoxification, reduced the proportion of Kupffer cells, and suppressed their production of inflammatory mediators (e.g., TNF-α, IL-6), potentially via upregulation of immunomodulatory genes ( Pilrb1, Ifi27l2a, Rtp4 ). Furthermore, gastrodin modulated hepatic stellate cell (HSC) heterogeneity by expanding a novel HSC subgroup characterized by Serpina expression and associated with anti-fibrotic traits. Conclusion Gastrodin alleviates cholestatic liver fibrosis through coordinated multi-cellular mechanisms involving hepatocytes, Kupffer cells, and HSCs. It represents a promising natural therapeutic c