南方医科大学学报

南方医科大学学报 ›› 2025, Vol. 45 ›› Issue (6): 1307-1316.doi: 10.12122/j.issn.1673-4254.2025.06.20

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复方积雪草减轻小鼠日本血吸虫引起的肝纤维化：通过调控TLR4/MyD88通路抑制炎症-纤维化级联反应

管丽萍1(), 颜燕1, 卢心怡1, 李智峰1, 高晖2, 曹东1, 侯晨曦1, 曾靖宇1, 李欣怡1, 赵洋2, 王俊杰2, 方会龙1()   

  1. 1.湘南学院，基础医学院，湖南 郴州 423000
    2.湘南学院，药学院，湖南 郴州 423000
  • 收稿日期:2024-04-22 出版日期:2025-06-20 发布日期:2025-06-27
  • 通讯作者: 方会龙 E-mail:3078770751@qq.com;huilongfang@163.com
  • 作者简介:管丽萍，在读本科生，E-mail: 3078770751@qq.com
  • 基金资助:
    湖南省卫生健康委科研课题(202101062143);国家级大学生创新创业训练计划项目(S202410545006);湖南省大学生创新创业训练计划项目(S202310545023S);湖南省普通高等学校教学改革研究项目(HNJG-2020-0932);湖南省高校思想政治工作精品项目(21JP031)

Compound Centella asiatica formula alleviates Schistosoma japonicum-induced liver fibrosis in mice by inhibiting the inflammation-fibrosis cascade via regulating the TLR4/MyD88 pathway

Liping GUAN1(), Yan YAN1, Xinyi LU1, Zhifeng LI1, Hui GAO2, Dong CAO1, Chenxi HOU1, Jingyu ZENG1, Xinyi LI1, Yang ZHAO2, Junjie WANG2, Huilong FANG1()   

  1. 1.School of Basic Medical Sciences, Xiangnan University, Chenzhou 423000, China
    2.School of Pharmacy, Xiangnan University, Chenzhou 423000, China
  • Received:2024-04-22 Online:2025-06-20 Published:2025-06-27
  • Contact: Huilong FANG E-mail:3078770751@qq.com;huilongfang@163.com

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摘要：

目的 基于网络药理学和分子对接，联合日本血吸虫（Sj）感染小鼠引起肝纤维化实验验证，探究复方积雪草（CCA）有效成分抗Sj病肝纤维化的作用机制。 方法 利用传统中药系统药理学数据库筛选CCA的活性成分及其作用靶点，与Sj病肝纤维化发病机制相关的靶点进行比对，获得CCA抗Sj病肝纤维化的潜在作用靶点；采用STRING数据库，构建药物疾病交集靶点的PPI网络，筛选核心靶点；通过Cytoscape 3.9.1构建“药物-成分-靶点-通路-疾病”网络图；运用David数据库进行GO和KEGG富集分析；利用PyMol和AutoDockVina1.1.2对CCA主要活性成分与核心靶点使用分子对接技术进行活性验证。将36只小鼠随机均分为正常对照组（control）、模型组（Sj）、复方积雪草组（Sj+CCA）；control组不做任何处理，其余经腹部皮肤感染血吸虫尾蚴建模成功8周后，分别每天灌胃生理盐水和CCA水煎剂，连续8周。采用HE、Masson染色法检测小鼠肝组织病理变化；免疫组化染色检测肝脏Collagen-Ⅰ、Collagen-Ⅲ的表达水平；ELISA法测定小鼠血清中IL-6和TNF-α水平变化；蛋白质印迹法检测肝组织TLR4（TOLL样受体4）、MyD88（髓分化因子88）蛋白表达。 结果 筛选得到107种CCA活性成分（OB≥30%，DL≥0.18），791个成分靶点，Sj病肝纤维化靶点集和CCA靶点集进行交集分析后共得到37个关键靶点。其中TNF、TP53、JUN、MMP9、CXCL为核心靶点，涉及的主要信号通路为 Lipid and atherosclerosis、IL-17 signaling pathway、Fluid shear stress and atherosclerosis、Chagas disease、Pathways in cancer等。分子对接结果提示，CCA107个活性成分中槲皮素与 TNF、TP53、JUN、MMP9有较好的结合活性。在小鼠Sj病肝纤维化模型中，CCA处理能够减轻炎性细胞浸润，减少Collagen-I、Collagen-III沉积（P<0.001，P<0.01），改善肝组织结构，降低其炎症因子IL-6、TNF-α的含量（P<0.05），下调TLR4、MyD88蛋白表达（P<0.01）。 结论 CCA可能作用于TNF、MMP9、JUN、TP53靶点，调控TLR4/MyD88信号通路，抑制IL-6、TNF-α等促炎因子释放，进而减少HSC活化及胶原纤维产生，降低ECM的沉积，阻断Sj虫卵肉芽肿的形成，发挥抗Sj感染引起的肝纤维化和保肝的作用。

关键词: 日本血吸虫病肝纤维化, 复方积雪草, 网络药理学, 分子对接, 实验验证

Abstract:

Objective To explore the therapeutic mechanism of compound Centella asiatica formula (CCA) for alleviating Schistosoma japonicum (Sj)-induced liver fibrosis in mice. Methods The active components and targets of CCA were identified using the TCMSP database with cross-analysis of Sj-related liver fibrosis targets. A "drug-component-target-pathway-disease" network was constructed using Cytoscape 3.9.1. Functional enrichment analysis (GO/KEGG) was performed using DAVID. Molecular docking study was carried out to validate interactions between the core targets and the key compounds. For experimental validation of the results, 36 mice were divided into control group, Sj-infected model group, and CCA-treated groups. In the latter two groups, liver fibrosis was induced via abdominal infection with Sj cercariae for 8 weeks, followed by 8 weeks of daily treatment with CCA decoction or saline. Hepatic pathology of the mice was assessedwith HE and Masson staining, and hepatic expressions of collagen-I and collagen-III were detected using immunohistochemistry; serum IL-6 and TNF-α levels were determined with ELISA. Hepatic expressions of TLR4 and MyD88 proteins were analyzed with Western blotting. Results We identified a total of 107 bioactive CCA components and 791 targets, including 37 intersection targets linked to Sj-induced fibrosis. The core targets included TNF, TP53, JUN, MMP9, and CXCL8, involving the IL-17 signaling, lipid metabolism, TLR4/MyD88 axis, and cancer pathways. Molecular docking study confirmed strong binding affinity between quercetin (a primary CCA component) and TNF/TP53/JUN/MMP9. In Sj-infected mouse models, CCA treatment significantly attenuated hepatic inflammatory cell infiltration, reduced collagen-I and collagen-III deposition, improved tissue architecture, reduced serum IL-6 and TNF-α levels, and downregulated TLR4 and MyD88 expressions in the liver. Conclusion CCA mitigates Sj-induced liver fibrosis by targeting TNF, TP53, JUN, and MMP9 to modulate the TLR4/MyD88 pathway, thereby suppressing pro-inflammatory cytokine release, inhibiting hepatic stellate cell activation, reducing collagen deposition, and preventing granuloma formation in the liver.

Key words: Schistosoma japonicum-induced hepatic fibrosis, compound Centella asiatica formula, network pharmacology, molecular docking, experimental validation