Objective To investigate the mechanism by which Scutellaria barbata D. Don regulates ferroptosis in hepatocellular carcinoma (HCC) cells.

Methods Network pharmacology was used to screen the intersecting targets of Scutellaria barbata D. Don, HCC, and ferroptosis regulation. Molecular docking was then performed to validate the binding between the main active components of Scutellaria barbata D. Don and the screened key targets. Huh7 cells were cultured in vitro, and the cell counting kit-8 (CCK-8) assay was used to detect cell proliferation after treatment with Scutellaria barbata D. Don at different concentrations (0, 3.15, 6.3, 12.5, 25, and 50 mg/mL) for 24 h, so as to determine the optimal drug concentration. Huh7 cells were divided into a control group and a Scutellaria barbata D. Don group (20 mg/mL). Lactate dehydrogenase (LDH) release, Fe²⁺ content, reactive oxygen species (ROS) levels, and mitochondrial membrane potential were measured. Quantitative polymerase chain reaction (qPCR) and Western blot were used to detect the mRNA and protein expression levels of ferroptosis-related indicators, including glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11), iron-responsive element-binding protein 2 (IREB2), and long-chain acyl-CoA synthetase 4 (ACSL4), as well as key targets, including mitogen-activated protein kinase 14 (MAPK14), tumor protein p53 (TP53), hypoxia-inducible factor-1α, NF-κB subunit (p65) gene (RELA), and MAPK1. A Huh7 subcutaneous xenograft model was established in male BALB/c nude mice, which were randomly divided into a control group and a Scutellaria barbata D. Don group. The control group was given normal saline by gavage, while the Scutellaria barbata D. Don group was given Scutellaria barbata D. Don granules at 140 mg/kg by gavage. Tumor volume and weight were monitored regularly. After tissue collection, hematoxylin-eosin (HE) staining was performed to evaluate pathological changes in tumor tissues.

Results Network pharmacology identified the top five active components of Scutellaria barbata D. Don as quercetin, luteolin, wogonin, baicalein, and eriodictyol. These components showed strong binding affinities with the core targets MAPK14, TP53, HIF-1α, RELA, and MAPK1. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that Scutellaria barbata D. Don-induced ferroptosis in HCC cells was associated with the HIF-1 signaling pathway, cancer-related signaling pathways, and the IL-17 signaling pathway. In vitro experiments confirmed that Scutellaria barbata D. Don inhibited Huh7 cell proliferation in a concentration-dependent manner. Compared with the control group, the Scutellaria barbata D. Don group showed increased intracellular ROS and Fe²⁺ levels and decreased mitochondrial membrane potential. Among ferroptosis-related indicators, the mRNA and protein expression levels of GPX4 and SLC7A11 were downregulated, while those of IREB2 and ACSL4 were upregulated. The mRNA and protein expression levels of the network pharmacology-screened targets MAPK1, TP53, MAPK14, and HIF-1α were increased, whereas RELA expression was decreased. In vivo experiments showed that tumor volume and weight in mice in the Scutellaria barbata D. Don group were lower than those in the control group. Tumor tissue cells were arranged irregularly, with obvious necrotic foci observed.

Conclusion Scutellaria barbata D. Don can effectively inhibit the malignant progression of HCC both in vivo and in vitro, and its mechanism may be associated with the induction of ferroptosis in HCC cells.