基于网络药理学与分子生物学实验探究L-紫苏醇延缓椎间盘退变的作用机制  

Mechanism of Perillyl Alcohol in Treating Intervertebral Disc Degeneration Explored Using Network Pharmacology and Molecular Biology Experiments

作　　者：杨江佳 张蒯祥 殷建林 刘曼 罗亚鸽 马言 罗漫丽 赵志强 李记天 YANG Jiangjia;ZHANG Kuaixiang;YIN Jianin;LIU Man;LUO Yage;MA Yan;LUO Mani;ZHAO Zhiqiang;LI Jitian(Hunan University of Traditional Chinese Medicine,Changsha 410000,China;Luoyang Orthopedic-Traumatological Hospital of Henan Province(Henan Province Orthopedic Hospital),Zhengzhou 450016,China;Henan University of Traditional Chinese Medicine,Zhengzhou 450046,China)

机构地区：[1]湖南中医药大学,长沙410000 [2]河南省洛阳正骨医院(河南省骨科医院),郑州450016 [3]河南中医药大学,郑州450046

出　　处：《中国现代应用药学》2025年第5期724-734,共11页Chinese Journal of Modern Applied Pharmacy

基　　金：河南省重点研发专项(231111310500);河南省省级科技研发计划联合基金项目(225200810084);河南省医学科技攻关计划联合共建项目(LHGJ20230490);河南省“科创中原”行动项目(2023HYTP040);河南省中医药拔尖人才培养项目(2022ZYBJ24);河南省中原医学科技创新发展基金会科学研究项目(ZYYC202303ZD)。

摘　　要：目的通过网络药理学探究L-紫苏醇(perillyl alcohol,PA)延缓椎间盘退变(intervertebral disc degeneration,IDD)的潜在分子作用机制,并运用分子对接和分子生物学实验进行验证。方法利用TCMSP、GeneCards、Swiss Target Prediction数据库获取PA的作用靶点;利用GeneCards、OMIM获得IDD相关靶点,并利用Venny 2.1.0在线网站对PA和IDD相关靶点取交集;使用String数据库和Cytoscape软件构建蛋白质-蛋白质相互作用(protein-protein interaction,PPI)图谱;DAVID及微生信进行GO和KEGG分析及结果可视化;借助AutoDock Vina及PyMOL软件将PA与IDD的核心靶点进行分子对接;应用实时荧光定量PCR(quantitative real-time PCR,qRT-PCR)、细胞免疫荧光及活性氧检测验证网络药理学预测结果。结果本研究共筛选PA靶点213个,IDD靶点2286个,获得61个PA与IDD交集靶点,经过PPI网络图谱分析获得PA延缓IDD的关键作用靶点为AKT1、EGF、TP53、EGFR等14个。GO富集分析涉及蛋白磷酸化的正向调控、细胞程序性死亡的正向调控、细胞周期蛋白、丝氨酸/苏氨酸激酶调节等,KEGG富集分析表明PA延缓IDD的作用机制可能涉及JAK-STAT、ErbB、HIF-1等多种信号通路。分子对接结果表明PA与ALB、EGFR、NOTCH1核心靶蛋白具有较强的结合活性。与IL-1β组相比,IL-1β+PA组MMP-1、MMP-13、ADAMTS4、ADAMTS5的mRNA表达水平降低,ACAN的mRNA表达上调,并且IL-1β+PA组HIF-1α与NOTCH1的mRNA表达上调,差异具有统计学意义。结论PA可能通过靶向NOTCH1、EGFR、ALB3等核心靶点激活HIF-1α/NOTCH1信号通路延缓IDD进程。OBJECTIVE To explore the mechanism of perillyl alcohol(PA)in delaying intervertebral disc degeneration(IDD)through network pharmacology,and validate its mechanism by molecular docking and molecular biology experiments.METHODS The targets of PA were obtained from TCMSP,GeneCards and Swiss Target Prediction database.GeneCards and OMIM were used to obtain IDD-related targets,and the common targets of PA and IDD were obtained from the Venny 2.1.0 website.String database and Cytoscape software were used to construct the protein-protein interaction(PPI)network.DAVID and bioinformatics website were used for GO and KEGG analysis.AutoDock Vina and PyMOL were used for molecular docking verification of the core targets of PA and IDD.The network pharmacological prediction results were verified by quantitative real-time PCR(qRT-PCR),cellular immunofluorescence and reactive oxygen species detection.RESULTS A total of 213 PA targets and 2286 IDD targets were screened in this study,and 61 intersection targets of PA and IDD were obtained.PPI network analysis showed that 14 key targets of PA to delay IDD were AKT1,EGF,TP53,EGFR,etc.GO enrichment analysis involved the positive Chin J Mod Appl Pharm,2025 March,Vol.42,No.5 regulation of protein phosphorylation,positive regulation of programmed cell death,cyclin,serine/threonine kinase regulation,etc.KEGG enrichment analysis showed that PA delayed IDD involving JAK-STAT,ErbB,HIF-1 and other signaling pathways.Molecular docking results showed that PA had strong affinity and relatively stable binding conformation with the core targets such as ALB,EGFR and NOTCH1.Compared with the IL-1βgroup,the expression levels of MMP-1,MMP-13,ADAMTS4 and ADAMTS5 mRNA in the IL-1β+PA treatment group were decreased,ACAN was up-regulated,HIF-1αand NOTCH1 in the IL-1β+PA group were up-regulated,too.The difference was statistically significant.CONCLUSION PA might act on NOTCH1,EGFR,ALB3 and other core targets to delay the progression of IDD through HIF-1α/NOTCH1 signaling pathway.

关 键 词：网络药理学 分子对接 椎间盘退变 L-紫苏醇 炎症 氧化应激 

分 类 号：R285.5[医药卫生—中药学]