Huanglian wendan decoction attenuates hepatic inflammation and lipogenesis via inhibition of the NF-κB/HDAC1/SREBP-1c axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Metabolic dysfunction-associated steatohepatitis (MASH) is characterized by hepatic steatosis accompanied by persistent inflammation and early fibrotic remodeling. In traditional Chinese medicine, Huanglian Wendan Decoction (HLWDD) is prescribed for phlegm-heat and damp-heat syndromes affecting the gallbladder and stomach and is traditionally used to clear heat, dry dampness, and resolve phlegm. It is commonly applied in the treatment of phlegm-heat-related metabolic disorders, including fatty liver disease. However, the therapeutic effects of HLWDD and the contributions of its key constituents to MASH remain to be further elucidated.

AIMS OF THE STUDY: This study aimed to evaluate the anti-inflammatory and lipid-regulatory effects of HLWDD and its key components in MASH and to explore the underlying molecular mechanisms.

MATERIALS AND METHODS: Male C57BL/6 J mice were given a methionine-choline-deficient (MCD) diet and received HLWDD in either low or high doses through oral gavage, with fenofibrate serving as a positive control. Body weight, liver index, serum levels of alanine aminotransferase and aspartate aminotransferase, serum lipid profiles, and hepatic triglyceride and total cholesterol contents were among the evaluated parameters. H&E, Oil Red O, and Masson's trichrome staining were used to evaluate histopathological changes. Hepatic macrophage infiltration was examined by immunofluorescence, inflammatory cytokines were measured by ELISA, and key signaling and lipid metabolism-related proteins were analyzed by western blotting. UPLC‒MS/MS was used to characterize the chemical profile of the HLWDD granules and identify their major constituents. Network pharmacology analysis integrating multiple databases, together with GO and KEGG enrichment analyses, was performed to predict potential targets and pathways. Molecular docking and molecular dynamics simulations were further used to investigate compound‒target interactions. Cell viability in vitro was measured with CCK-8 assays, protein levels were confirmed through western blotting, and intracellular lipid buildup was assessed using Oil Red O staining.

RESULTS: UPLC‒MS/MS analysis revealed that berberine (BBR), an isoquinoline alkaloid, is a major bioactive component of HLWDD. Network pharmacology analysis suggested that HLWDD and BBR may exert anti-MASH effects by modulating multiple targets and pathways, including IL-6, PPARα, and the NF-κB/HDAC1/SREBP-1c axis. These predictions were supported by in vivo experiments, which confirmed the protective effects of HLWDD against MASH. Both in vivo and in vitro studies further revealed that BBR markedly ameliorated MASH-related phenotypes by suppressing the NF-κB/HDAC1/SREBP-1c axis. Additionally, simulations of molecular docking and dynamics revealed stable interactions between BBR and important proteins within this axis. Microscale thermophoresis (MST) assays further demonstrated direct binding of BBR to HDAC1. Collectively, these findings suggest that HLWDD and its key active constituent BBR alleviate MASH, at least in part, by inhibiting the NF-κB/HDAC1/SREBP-1c axis, which is closely associated with inflammatory responses and dysregulated lipogenesis.

CONCLUSION: HLWDD markedly ameliorated the MASH phenotype by attenuating hepatic inflammation and lipogenesis, with the NF-κB/HDAC1/SREBP-1c axis emerging as a key mechanism linking inflammatory signaling to aberrant lipid synthesis. BBR, identified by UPLC‒MS/MS as a major active constituent of HLWDD, largely recapitulated these effects and directly bound to HDAC1, supporting its important contribution to the protective effects of HLWDD against MASH.