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| 茅莓水提物抗肝纤维化的作用及机制研究 |
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王果,谷英敏,王军伟,茅雪莉,季梦漂
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杭州医学院附属天台县人民医院
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| 摘要: |
| 目的：探究茅莓(Rubus parvifolius L.,RPL)水提取物(ERPL)对肝纤维化的作用及机制。方法： 取40只雄性SD大鼠随机分为对照组、模型组、模型+75 mg/kg ERPL 组和模型+225 mg/kgERPL组,每组10只。模型组、模型+75 mg/kg ERPL组、模型+225 mg/kg ERPL组大鼠,分别予CCl4溶液(溶于橄榄油)每周皮下注射2次,持续8周,以诱导肝纤维化。造模后第5周开始进行对应剂量ERPL灌胃给药。对照组和模型组大鼠则给予等量的0.9 % 氯化钠,持续8周。HE染色和Masson染色观察肝脏组织病理状态,检测大鼠肝组织中ALT、AST和ALP的含量,Western Blot检测大鼠肝脏中铁死亡相关蛋白FER、TFR-1、TF和ACSL4以及p53/SLC7A11/GPX4通路蛋白的水平。体外通过PDGF-BB诱导肝星状细胞(HSC)LX-2细胞活化,CCK8法检测细胞活性,qRT-PCR和Western Blot法检测细胞中α-SMA、collagen 1、fibronectin的表达,试剂盒检测细胞ROS和Fe2+的表达,Western Blot检测细胞中铁死亡相关蛋白FER、TFR-1、TF和ACSL4以及p53/SLC7A11/GPX4通路蛋白的水平。结果： 在体内,ERPL能减轻CCl4诱导的大鼠肝损伤和肝纤维化,降低了肝组织中ALT、AST、ALP以及纤维化相关蛋白LN、PIIINP和Col-IV的表达水平。ERPL调节p53/SLC7A11/GPX4信号并诱导肝组织纤维细胞产生铁死亡。在体外,ERPL降低PDGF-BB诱导的LX-2细胞的活力以及α-SMA、collagen 1、fibronectin的表达,抑制LX-2细胞的活化。ERPL增强了经PDGF-BB处理的LX-2细胞的氧化应激和铁死亡反应,表现为ROS、MDA、Fe2+水平的升高,GSH-Px水平的降低以及铁死亡蛋白FER、TF、ACSL4和TFR-1表达的增强。此外,ERPL能调节p53/SLC7A11/GPX4通路。然而p53信号通路抑制剂PFT-α逆转了ERPL的作用。结论：ERPL通过调节p53/SLC7A11/GPX4通路诱导铁死亡反应,从而改善肝纤维化。 |
| 关键词: 茅莓(Rubus parvifolius L.) 肝纤维化 肝硬化 铁死亡 p53/SLC7A11/GPX4通路 |
| DOI： |
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| 基金项目:浙江省卫生健康委551卫生人才培养工程(浙卫办[2021]40号) |
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| Exploration of the anti-hepatic fibrosis effect and mechanism of aqueous extracts of Rubus parvifolius L.Wang Guo 1, Gu Yingmin1, Wang Junwei 2*, Mao Xueli1, Ji Mengpiao1 |
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wangguo1,2,3, guyingmin1,2,3, wangjunwei1,2,3, maoxueli1,2,3, jimengpiao1,2,3
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1.Tiantai People'2.'3.s Hospital affiliated to Hangzhou Medical College
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| Abstract: |
| Objective: To investigate the effect and mechanism of aqueous extract of Rubus parvifolius L. (ERPL) on liver fibrosis. Methods: 40 male SD rats were randomly divided into control, model, model+75 mg/kg ERPL and model+225 mg/kg ERPL groups, with 10 rats in each group. Rats in the model group, model+75 mg/kg ERPL group and model+225 mg/kg ERPL group were given CCl4 solution (dissolved in olive oil) subcutaneously twice a week for 8 weeks to induce liver fibrosis. Corresponding doses of ERPL were administered by gavage starting at week 5 after modeling. The rats in the control and model groups were given equal amounts of 0.9% NaCl for 8 weeks. HE staining and Masson staining were used to observe the histopathological status of liver tissues. The levels of ALT, AST and ALP were detected in the liver tissues of rats. Western Blot was used to detect the levels of Ferroptosis-associated proteins FER, TFR-1, TF and ACSL4 and p53/SLC7A11/GPX4 pathway proteins in the livers of rats. Hepatic stellate cell (HSC) LX-2 cell activation was induced by PDGF-BB in vitro, and cell activity was detected by CCK8 assay. qRT-PCR and Western Blot assays were performed to detect the expression of α-SMA, collagen 1 and fibronectin in the cells. Cellular ROS and Fe2+ expression were detected by kits. Western Blot was used to detect the levels of Ferroptosis-associated proteins FER, TFR-1, TF, and ACSL4, as well as p53/SLC7A11/GPX4 pathway proteins in cells. Results: In vivo, ERPL attenuated CCl4-induced liver injury and hepatic fibrosis in rats, and reduced the expression levels of ALT, AST, ALP, and fibrosis-associated proteins LN, PIIINP, and Col-IV in liver tissues. ERPL regulated p53/SLC7A11/GPX4 signaling and induced ferroptosis in liver tissue fibroblasts. In vitro, ERPL decreased the viability as well as the expression of α-SMA, collagen 1, fibronectin in PDGF-BB-induced LX-2 cells and inhibited LX-2 cell activation. ERPL enhanced oxidative stress and ferroptosis responses in PDGF-BB-treated LX-2 cells, as evidenced by elevated levels of ROS, MDA and Fe2+, decreased levels of GSH-Px, and enhanced expression of the ferroptosis proteins FER, TF, ACSL4, and TFR-1. In addition, ERPL had a regulatory effect on the p53/SLC7A11/GPX4 pathway. However the p53 signaling pathway inhibitor PFT-α reversed the effects of ERPL. Conclusion: ERPL induced ferroptosis response by regulating the p53/SLC7A11/GPX4 pathway, thereby ameliorating liver fibrosis. |
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