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Objective To investigate the interventional effect of anwulignan on pulmonary fibrosis and the underlying mechanism. Methods Sixty-five male ICR mice were randomly divided into control group, bleomycin (BLM) model group, anwuzhisu low dose group (1 mg/kg), anwuzhisu high dose group (4 mg/kg) and N-acetylcysteine group (150 mg/kg), with 13 mice in each group. Mouse pulmonary fibrosis model was induced by intratracheal perfusion of 5 mg/kg BLM. The serum and lung tissues were collected, and the pathological changes of lung tissues were observed by HE staining and Masson trichrome staining; serum oxidative stress index and hydroxyproline (HYP) content in lung tissue were measured by kit; the expression level of iron death pathway related genes was detected by Western blotting and qRT-PCR. Take the logarithmic growth phase HFL-1 cells, (1) set the control group and anwuzhisu administration group (0.3125, 0.625, 1.25, 2.5, 5, 10, 20,40, 80 μmol/L), and CCK-8 method was used to detect the toxic effect of anwuzhi on HFL-1 cells. (2) Set the control group, transforming growth factor (TGF)-β1 model group, anwuzhisu low dose group (5 μmol/L) and anwuzhisu high dose group(10 μmol/L), with TGF-β1 induced pulmonary fibrosis cell model. The oxidative stress index and reactive oxygen species(ROS) level were measured by kit; the expression level of iron death pathway related genes was detected by Western blotting and qRT-PCR. Results HE staining, Masson trichrome staining and the increase of HYP content indicated that the BLM-induced pulmonary fibrosis model was successfully constructed, and the pulmonary fibrosis phenotype was significantly improved after the administration of anwulignan. CCK-8 assay showed that the concentration of anwulignan <20 μmol/L had no significant effect on the proliferation activity of HFL-1 cells (P>0.05). Compared with model group, the activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT) and the levels of glutathione (GSH) in lung tissues of mice and HFL-1 cells were significantly increased after the administration of anwulignan, while the levels of malondialdehyde(MDA) were significantly decreased (P<0.01 or P<0.001). Compared with TGF-β1 model group, 10 μmol/L anwulignan could decrease the level of ROS in HFL-1 cells (P<0.01). Western blotting and qRT-PCR results showed that anwulignan could significantly up-regulate the expressions of ferroptosis pathway related genes in lung tissues of mice and HFL-1 cells including glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11) and transferrin (TF), and significantly down-regulate the expression of transferrin receptor 1 (TFR1) (P<0.05, P<0.01 or P<0.001). Conclusion Anwulignan can improve pulmonary fibrosis by inhibiting oxidative stress and ferroptosis, laying a foundation for the development of clinical drugs for pulmonary fibrosis.
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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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