基于网络药理学探析黄精抗骨质疏松的机制及验证

Mechanism of Polygonati Rhizoma on Treating Osteoporosis Based on Network Pharmacology and Preliminary Verification

目的:基于网络药理学及分子对接方法分析黄精抗骨质疏松(OP)的潜在机制,并通过实验进行验证。方法:利用中药系统药理学数据库(TCMSP)2.3进行条件检索,获取黄精的活性成分及其对应作用靶点,并在疾病相关的基因与突变位点数据库(DisGeNET)7.0获取骨质疏松的治疗靶标,两者交集即为黄精抗骨质疏松的潜在靶基因。并用Cytoscape 3.7.1构建“黄精-活性成分-潜在靶点”网络,借助STRING数据库11.0进行蛋白质-蛋白质相互作用(PPI)分析,构建PPI网络。此外,采用Metascape数据库3.5分析其参与的基因本体(GO)富集分析及京都基因与基因组百科全书(KEGG)富集分析。选取黄精重要活性成分与靶点通过AutoDock Vina 1.1.2进行分子对接验证,最后借助小鼠胚胎成骨细胞前体细胞(MC3T3-E1)观察β-谷甾醇对成骨分化的影响。结果:黄精活性成分有12个,其作用靶点与骨质疏松治疗靶点相映射得到潜在靶点32个。Cytoscape 3.7.1拓扑分析得到黄芩素、β-谷甾醇等6个主要成分,蛋白激酶B1(Akt1)、肿瘤蛋白p53(TP53)、血管内皮生长因子A(VEGFA)、JUN原癌基因(JUN)、基质金属蛋白酶-9(MMP-9)、原癌基因c-FOS(FOS)等关键靶点。GO和KEGG富集分析中,共获得995个GO条目,涉及活性氧调节,生长调节等181条信号通路。分子对接显示黄精中核心成分与各自的靶点表现出良好的对接活性。细胞实验结果显示β-谷甾醇在2.5,5μmol·L^(-1)浓度时具有显著促进成骨分化的作用。结论:黄精可通过调节炎症、氧化应激、凋亡、代谢等作用,发挥治疗骨质疏松的效果,其中β-谷甾醇可促进MC3T3-E1细胞成骨分化。

Objective: To explore the potential mechanism of Polygonati Rhizoma on the treatment of osteoporosis(OP)based on network pharmacology and molecular docking method and to verify the mechanism by experiments. Method: The main active ingredients and corresponding targets of Polygonati Rhizoma were screened out from Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP)2.3 by conditional searching. The treatment targets were obtained from the genes related to OP and DisGeNET 7.0. The potential target genes of Polygonati Rhizoma for treating OP were obtained by the crossing of the corresponding targets and the treatment targets. Cytoscape 3.7.1 was used to construct the“Polygonati Rhizoma-active ingredient-potential target”network. The protein-protein interaction(PPI)analysis was carried out by STRING 11.0,and the PPI network was constructed. Metascape 3.5 was used to conduct Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses of the key targets. The core ingredients and key targets of Polygonati Rhizoma were selected for molecular docking by AutoDock Vina1.1.2. Finally,the effect of β-sitosterol on osteogenic differentiation of MC3T3-E1 cells in rats was observed.Result: Twelve active ingredients and 32 potential targets of Polygonati Rhizoma for OP treatment were screened out. Six active ingredients including baicalein and β-sitosterol and key targets including protein kinase 1(Akt1),tumor suppressor p53(TP53),vascular endothelial growth factorA(VEGFA),proto-oncogene Jun(JUN),matrix metalloproteinase-9(MMP-9),and proto-oncogene c-Fos(FOS)were obtained by Cytoscape3.7.1 topological analysis. A total of 995 GO entries and 181 signaling pathways involving the response to reactive oxygen species and regulations of growth were obtained from GO and KEGG enrichment analyses. The results of molecular docking showed that the core active ingredients possessed good binding activities with the respective key targets. The results of cell experiments showed that β-sitosterol promoted the osteogenic differentiation at the concentration of 2.5 μmol·L^(-1)and 5 μmol·L^(-1). Conclusion: Polygonati Rhizoma had the therapeutic effect on treating OP by regulating inflammation,oxidative stress,apoptosis,and metabolism. The β-sitosterol significantly promoted the osteogenic differentiation of MC3T3-E1 cells.