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The objective of this study was to explore the therapeutic effects and underlying mechanisms of Guizhi Fuling Wan (GZFLW) in the treatment of hepatocellular carcinoma (HCC). Firstly, the therapeutic effect of GZFLW on HCC was evaluated by constructing a subcutaneous xenograft mouse model of H22 hepatoma cell line and combining with cell experiments. Liquid chromatography tandem mass spectrometry (LC-MS/MS) was used to analyze the drug components of GZFLW, and network pharmacology and transcriptomics methods were combined to screen out the potential core targets and signaling pathways of GZFLW in the treatment of HCC, and the core targets were verified by molecular experiments. Bioinformatics analysis results showed that, GZFLW may regulate p53, vascular endothelial growth factor A (VEGF-A) and hypoxia inducible factor-1α (HIF-1α), which may play an important role in the regulation of GZFLW. HIF-1α and other key targets, which have a certain impact on angiogenesis and ferroptosis in HCC. The results of in vitro and in vivo experiments showed that GZFLW could significantly inhibit the growth of subcutaneous transplanted tumor, and the serum containing GZFLW could inhibit the proliferation of H22 cells in vitro. The results of immunofluorescence and immunohistochemistry showed that GZFLW could down-regulate the expression of vascular endothelial markers cluster designation 31 (CD31) and cluster designation 34 (CD34). It also reduced the levels of VEGF-A and HIF-1α. Western blot analysis further showed that after GZFLW treatment, the expression of ferroptosis-related proteins solute carrier family 7 member 11 (SCL7A11) and glutathione peroxidase 4 (GPX4) was significantly decreased. However, the expression of p53 was significantly increased. The comprehensive analysis results suggest that GZFLW may induce ferroptosis in HCC by regulating the p53/SLC7A11/GPX4 signaling pathway, and inhibit tumor angiogenesis by inhibiting the expression of HIF-1α and VEGF-A, thereby effectively inhibiting the occurrence and development of HCC. This experiment has been approved by the Animal Experiment Ethics Committee of Shaanxi University of Chinese Medicine (ethics approval number: SUCMDL20230915001).
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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