咖啡酸苯乙酯治疗脊髓损伤的网络药理学研究及实验验证

王柯文; 陈建敏; 李筱璐; 桂裕昌; 唐云; 许建文

doi:10.16190/j.cnki.45-1211/r.2023.05.013

咖啡酸苯乙酯治疗脊髓损伤的网络药理学研究及实验验证

Network pharmacological study and experimental verification of caffeic acid phenethyl ester in the treatment of spinal cord injury

摘要

摘要: 目的：运用网络药理学方法探究咖啡酸苯乙酯（CAPE）治疗脊髓损伤（SCI）的主要机制并行实验验证。方法：通过SwissTargetPrediction、SEA、STITCH和TargetNet数据库获取CAPE的作用靶点，通过GeneCards、Malacards、DrugBank、CTD及TTD 数据库获取SCI 相关基因。使用STRING 在线数据库获取交集基因的蛋白互作（PPI）网络并通过Cytoscape 软件筛选核心靶点，R软件对交集基因进行GO分析及KEGG通路富集分析，并对富集结果进行可视化，AutoDock软件进行分子对接。将24 只SD 大鼠随机分为假手术组、模型组、CAPE 低剂量组、CAPE 高剂量组，每组6 只。采用改良Allen 法建立SCI 大鼠模型，Basso Beattie Bresnahan（BBB）评分检测大鼠后肢运动功能，苏木精—伊红（HE）染色及尼氏染色观察脊髓组织结构及神经元，RT-qPCR 法验证CAPE 干预后大鼠脊髓组织核心靶点的表达。结果：共筛选出48 个CAPE 治疗SCI 的预测作用靶点，EGFR、JUN、ESR1、MMP9、PTGS2 为核心靶点。GO 分析和KEGG 分析显示，CAPE 治疗SCI 可能与雌激素、HIF-1、VEGF 和IL-17 等信号通路有关，分子对接表明CAPE 与EGFR、JUN、ESR1、MMP9、PTGS2蛋白均有较好的结合性。动物实验结果表明，CAPE干预明显提高SCI 大鼠BBB 评分，减轻SCI 大鼠脊髓组织的结构破坏以及神经元损伤。与模型组相比，CAPE 低剂量组及高剂量组JUN、MMP9、PTGS2、EGFR mRNA 表达水平降低，ESR1 mRNA 表达水平升高（均P< 0.05）。结论：CAPE 可能通过下调EGFR、JUN、MMP9和PTGS2基因表达及上调ESR1基因表达，调控雌激素、HIF-1、VEGF和IL-17等信号通路发挥SCI后的神经保护作用。

Abstract: Objective:To explore the main mechanism of caffeic acid phenethyl ester(CAPE)in the treatment of spinal cord injury(SCI)using the method of network pharmacology and test it experimentally.Methods:The ac-tion targets of CAPE were obtained from SwissTargetPrediction, SEA, STITCH and TargetNet databases, and SCI related genes were obtained from GeneCards, Malacards, DrugBank, CTD and TTD databases.The protein interaction (PPI) network of overlapping genes was obtained from STRING online database and the core genes were screened by Cytoscape software.GO analysis and KEGG pathway enrichment analysis of overlapping genes were carried out by R software, the enrichment results were visualized and AutoDock software was used for molecular docking.Twenty-four SD rats were randomly divided into sham operation group, model group, lowdose CAPE group and high-dose CAPE group, with 6 rats in each group.The SCI rat model was established by modified Allen method.The motor function of hind limbs in rats was evaluated by Basso Beattie Bresnahan(BBB) score.Hematoxylin-eosin (HE) staining and Nissl staining were used to observe the structural in-tegrity of spinal cord tissue and neuronal survival.Re-al-time fluorescence quantitative polymerase chain re-action (RT-qPCR) was used to verify the expression levels of core targets in spinal cord tissue of rats after CAPE intervention.Results:A total of 48 predictive targets for CAPE treatment of SCI were screened and EG-FR, JUN, ESR1, MMP9 and PTGS2 were the core targets.GO analysis and KEGG analysis showed that CAPE treatment of SCI might be related to estrogen, HIF-1, VEGF and IL-17 signaling pathways.Molecular docking showed that CAPE had good binding to EGFR, JUN, ESR1, MMP9 and PTGS2 proteins.The results of animal experiments showed that the intervention of CAPE significantly increased the BBB score of SCI rats, and re-duced the structural damage of spinal cord tissue and neuronal injury in SCI rats.Compared with the model group, the expression levels of JUN, MMP9, PTGS2 and EGFR mRNA in low-dose CAPE group and high-dose CAPE group decreased, while the expression level of ESR1 mRNA increased (all P< 0.05).Conclusion:CAPE may regulate the signaling pathways of estrogen, HIF-1, VEGF and IL-17 by down-regulating the EGFR, JUN, MMP9 and PTGS2 gene expressions and up-regulating the ESR1 gene expression, and then play a neuroprotec-tive role after SCI.

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