基于网络药理学预测和验证当归芪枣精“一方多效”的作用机制

Prediction and Validation of"Multi-efficacy of One Formula"Mechanism of Danggui Qi Zao Extracts Based on Network Pharmacology

目的:采用网络药理学方法,探索当归芪枣精“一方多效”的作用机制,并对该方改善气血双虚证候的相关靶点蛋白进行验证。方法:在数据库检索的基础上,构建了蛋白-蛋白相互作用网络和成分-靶点-疾病网络,并对靶点进行基因本体(GO)富集分析和京都基因组百科全书(KEGG)通路分析。同时,采用放血和环磷酰胺并用建立气血双虚小鼠模型,以当归芪枣精低剂量组(4.0 g·kg)和高剂量组(8.0 g·kg)连续干预7 d,采用蛋白质免疫印迹(Western blot)法,检测骨髓中表皮生长因子受体(EGFR)、信号转导和转录激活因子1(STAT1)、丝裂原活化蛋白激酶(MAPK)及其磷酸化蛋白的表达。结果:筛选出53种活性成分,作用于110个潜在靶点,包括RB1、EGFR、ESR1等中枢蛋白,参与细胞增殖与凋亡、转录翻译和应激反应等生物过程,调控信号转导、免疫炎症以及癌症等信号通路,与变应性鼻炎、阿尔茨海默病、帕金森病、哮喘等307种疾病相关,尤以各类癌症为多。实验结果显示,与正常组相比,模型组小鼠骨髓组织EGFR、STAT1表达和p-MAPK/MAPK比值均显著降低(P<0.05);与模型组相比,当归芪枣精低剂量组、高剂量组小鼠骨髓组织EGFR、STAT1表达和p-MAPK/MAPK比值均显著升高(P<0.05);但两者在升高pMAPK/MAPK比值上有显著差异(P<0.05)。结论:当归芪枣精具有治疗神经系统、呼吸系统疾病,癌症等“一方多效”的治疗价值,EGFR、MAPK、STAT1等关键靶点共同参与调节气血双虚证候,可能是当归芪枣精气血双补的主要作用机制。

Objective:To explore the mechanism of"Multi-efficacy of One Formula"of Danggui Qi Zao extracts by network pharmacology,and the related target proteins of this formula in improving the syndrome of qi-blood deficiency were verified.Method:Based on database retrieval,protein-protein interaction networks and component-target-disease networks were constructed.Meanwhile,targets were analyzed by gene ontology(GO)enrichment analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis.At the same time,mice were subjected to bloodletting and cyclophosphamide to form the qi-blood deficiency model,and treated with Danggui Qi Zao extracts at low(4.0 g·kg)or highdose(8.0 g·kg)for 7 days.The expression of epidermal growth factor receptor(EGFR),signal transductor and transcription activator 1(STAT1),mitogen-activated protein kinase(MAPK)and phosphorylated MAPK in bone marrow were detected by Western blot.Result:Totally 53 active components were screened that acted on 110 potential targets,including hub proteins such as RB1,EGFR and ESR1,which involved in biological processes such as cell proliferation and apoptosis,transcriptional translation and stress response,and regulate signal transduction,immune inflammation and cancer signaling pathways.Besides,it associated with 307 diseases such as allergic rhinitis,Alzheimer’s disease,Parkinson’s disease,asthma and so on,especially cancer.The test results showed that compared with the normal group,EGFR and STAT1expressions and p-MAPK/MAPK ratio in bone marrow tissues of the model group were significantly decreased(P<0.05).Compared with the model group,EGFR and STAT1 expression and p-MAPK/MAPK ratio were significantly increased in the low and high dose groups(P<0.05),in addition,there was significant difference in increasing P-MAPK/MAPK ratio between the low and high dose groups(P<0.05).Conclusion:Danggui Qi Zao extracts has therapeutic values of"Multi-efficacy of One Formula"in the treatment of nervous system,respiratory system,cancer and other diseases.EGFR,MAPK,STAT1 and other key targets jointly participate in the regulation of qi-blood deficiency syndrome,which may be the main mechanism of qi-blood double supplement of Danggui Qi Zao extracts.