Yan Hai-Yan, Zou Chun-Cai
Pharmacy School of Wannan Medical College Wuhu 241002, China.
Zhongguo Zhong Yao Za Zhi. 2022 Mar;47(5):1370-1382. doi: 10.19540/j.cnki.cjcmm.20210923.401.
This study explored the anticoagulant material basis and mechanism of Trichosanthis Semen and its shell and kernel based on spectrum-effect relationship-integrated molecular docking. High performance liquid chromatography(HPLC) fingerprints of Trichosanthis Semen and its shell and kernel were established. Prothrombin time(PT) and activated partial thromboplastin time(APTT) in mice in the low-and high-dose(5, 30 g·kg(-1), respectively) Trichosanthis Semen, the shell, and kernel groups were determined as the coagulation markers. The spectrum-effect relationship and anticoagulant material basis of Trichosanthis Semen and its shell and kernel were analyzed with mean value calculation method of Deng's correlation degree(MATLAB) and the common effective component cluster was obtained. Then the common targets of the component cluster and coagulation were retrieved from TCMSP, Swiss-TargetPrediction, GenCLiP3, GeneCards, and DAVID, followed by Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment of the targets. The main anticoagulant molecular mechanism of the component cluster was verified by SYBYL-X 2.1.1. The spectrum-effect relationship of Trichosanthis Semen and its shell and kernel was in positive correlation with the dosage. The contribution of each component to anticoagulation was not the same, suggesting that the material basis for anticoagulation was different, but they have common effective components(i.e. common material basis), such as adenine(peak 3), uracil(peak 4), hypoxanthine(peak 6), xanthine(peak 9), and adenosine(peak 11). Network pharmacology showed that these components can act on multiple target proteins such as NOS3, KDR, and PTGS2, and exert anticoagulant effect through multiple pathways such as VEGF signaling pathway. They involved the biological functions such as proteolysis, cell component such as cytosol, and molecular functions. The results of molecular docking showed that the binding free energy of these components with NOS3(PDB ID: 1 D0 C), KDR(PDB ID: 5 AMN), and PTGS2(PDB ID: 4 COX) was ≤-5 kJ·mol(-1), and the docking conformations were stable. Spectrum-effect relationship-integrated molecular docking can be used for the optimization, virtual screening, and verification of complex chemical and biological information of Chinese medicine. Trichosanthis Semen and its shell and kernel have the common material basis for anticoagulation and they exert the anticoagulant through multiple targets and pathways.
本研究基于谱效关系结合分子对接技术，探讨了瓜蒌子及其壳、仁的抗凝物质基础及作用机制。建立了瓜蒌子及其壳、仁的高效液相色谱（HPLC）指纹图谱。将低、高剂量（分别为5、30 g·kg⁻¹）瓜蒌子组、壳组和仁组小鼠的凝血酶原时间（PT）和活化部分凝血活酶时间（APTT）作为凝血指标进行测定。采用邓氏关联度均值计算法（MATLAB）分析瓜蒌子及其壳、仁的谱效关系和抗凝物质基础，得到共同有效成分聚类。然后从中药系统药理学数据库与分析平台（TCMSP）、瑞士靶点预测（Swiss-TargetPrediction）、基因与蛋白质相互作用综合数据库（GenCLiP3）、基因卡片（GeneCards）和数据库注释、可视化与集成发现（DAVID）中检索该成分聚类与凝血相关的共同靶点，接着对这些靶点进行基因本体（GO）术语富集和京都基因与基因组百科全书（KEGG）通路富集分析。通过SYBYL-X 2.1.1软件验证该成分聚类的主要抗凝分子机制。瓜蒌子及其壳、仁的谱效关系与剂量呈正相关。各成分对抗凝作用的贡献不同，表明抗凝物质基础存在差异，但它们具有共同有效成分（即共同物质基础），如腺嘌呤（峰3）、尿嘧啶（峰4）、次黄嘌呤（峰6）、黄嘌呤（峰9）和腺苷（峰11）。网络药理学研究表明，这些成分可作用于一氧化氮合酶3（NOS3）、激酶插入域受体（KDR）和前列腺素内过氧化物合酶2（PTGS2）等多个靶蛋白，并通过血管内皮生长因子（VEGF）信号通路等多条途径发挥抗凝作用。它们涉及蛋白水解等生物学功能、胞质溶胶等细胞成分以及分子功能。分子对接结果表明，这些成分与NOS3（PDB编号：1D0C）、KDR（PDB编号：5AMN）和PTGS2（PDB编号：4COX）的结合自由能≤ -5 kJ·mol⁻¹，且对接构象稳定。谱效关系结合分子对接技术可用于优化、虚拟筛选和验证中药复杂的化学和生物学信息。瓜蒌子及其壳、仁具有共同的抗凝物质基础，并通过多个靶点和途径发挥抗凝作用。
