该研究基于代谢组学、网络药理学和分子对接技术探讨西洋参三七丹参颗粒抗衰老的作用机制.采用腹腔注射D-半乳糖(D-gal)诱导衰老小鼠模型,将小鼠随机分为对照组(control组),模型组(model组),阳性药组(褪黑素组,MT组),西洋参三七丹参颗粒低、中、高剂量组(XSD-L、XSD-M、XSD-H组).进行旷场实验,免疫荧光检测细胞周期阻滞蛋白(p16)和磷酸化组蛋白H2A变异体(phosphorylated histone family 2A variant,γH2AX)在脑组织中的表达,酶联免疫吸附法(ELISA)测定小鼠脑组织中炎症因子白细胞介素-1β(interleukin-1β,IL-1β)和白细胞介素-6(interleukin-6,IL-6)表达水平.对control组、model组、XSD-H组的小鼠血清进行代谢组学分析得到代谢过程及代谢物;通过网络药理学预测西洋参三七丹参颗粒有效化学成分和潜在靶点,构建"药物-有效化学成分-关键靶点"网络图,进行基因本体论(Gene Ontology,GO)分析、京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)分析以及蛋白-蛋白互作(protein-protein interaction,PPI)网络构建,阐明西洋参三七丹参颗粒抗衰老的作用机制.结果表明,西洋参三七丹参颗粒能明显改善D-gal小鼠衰老程度,显著改善D-gal小鼠总运动距离和平均运动速度,减少休息时间;显著降低D-gal小鼠IL-6和IL-1β的蛋白水平;明显降低p16和γH2AX的表达;与model组相比,XSD-H组显著上调了 66个差异代谢物(differential metabolites,DMs),下调了 91个DMs;筛选出4条关键代谢途径(色氨酸代谢、甘油磷脂代谢、嘧啶代谢、赖氨酸降解)和16种潜在生物标志物(赖氨酸、色氨酸、吲哚乙醛、磷脂酰胆碱、溶血磷脂酰胆碱、3-羟基邻氨基苯甲酸、褪黑素等);网络药理学方法筛选出西洋参三七丹参颗粒的主要活性成分58个,关键靶点62个;GO功能富集分析发现与基因表达的正向调控、药物反应等;KEGG通路富集筛选到涉及糖尿病并发症相关的AGE-RAGE信号通路、缺氧诱导因子-1信号通路等;并通过PPI网络以及分子对接筛选出STAT3、MAPK1、MAPK14、EGFR、FOS、STAT1等6个潜在核心靶点.
Anti-aging effect and molecular mechanism of Xiyangshen Sanqi Danshen Granules based on metabolomics and bioinformatics
This study investigated the anti-aging mechanism of Xiyangshen Sanqi Danshen Granules based on metabonomics,network pharmacology,and molecular docking.The aging mice model was induced by intraperitoneal injection of D-galactose(D-gal).Mice were randomly divided into a control group,model group,melatonin group(MT group),and low,medium,and high dose groups of Xiyangshen Sanqi Danshen Granules(XSD-L,XSD-M,and XSD-H).An open-field experiment was conducted,and the expression of cell cycle arrest proteins(p16)and phosphorylated histone family 2A variant(yH2AX)in the brain tissue was detected by immunofluorescence.The expression of interleukin-1β(IL-1β)and interleukin-6(IL-6)in the brain tissue was detected by enzyme-linked immunosorbent assay(ELISA).Metabolomics analysis was performed on the serum of mice in control,model,and XSD-H groups to obtain metabolic processes and metabolites.The effective chemical components and potential targets of Xiyangshen Sanqi Danshen Granules were predicted through network pharmacology,and the network diagram of"drug-effective chemical components-key targets"was constructed.Gene Ontology(GO)analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis were carried out,and a protein-protein interaction(PPI)network was constructed to clarify the anti-aging mechanism of Xiyangshen Sanqi Danshen Granules.The results showed that the Xiyangshen Sanqi Danshen Granules could significantly improve the aging degree of D-gal mice,significantly improve the total motion distance and the mean motion speed of D-gal mice,and reduce the rest time.In addition,Xiyangshen Sanqi Danshen Granules could significantly reduce the protein levels of IL-6 and IL-1β and the expression of p 16 and γH2AX in D-gal mice.Compared with the model group,66 differential metabolites(DMs)were significantly up-regulated,and 91 DMs were down-regulated in the XSD-H group.Moreover,four key metabolic pathways(tryptophan metabolism,glycerophospholipid metabolism,pyrimidine metabolism,and lysine degradation)and 16 biomarkers(lysine,tryptophan,indoleacetaldehyde,PCs,LysoPCs,3-hydroxyanthranilic acid,melatonin,etc)were screened out.58 main active components and 62 key targets of Xiyangshen Sanqi Danshen Granules were screened by network pharmacology.The GO functional enrichment analysis found the positive regulation of gene expression,drug response,etc.KEGG pathway enrichment screening involved diabetic complications-related AGE-RAGE signaling pathway,hypoxia inducible factor-1 signaling pathway,etc.Through the PPI network and molecular docking,six potential core targets of STAT3,MAPK1,MAPK14,EGFR,FOS,and STAT1 were screened.
万方数据
维普
