神经药理学报

神经药理学报 ›› 2024, Vol. 14 ›› Issue (6): 1-.DOI: 10.3969/j.issn.2095-1396.2024.06.001

• 研究论文 •    下一篇

基于网络药理学和分子对接探讨牛磺酸治疗年龄相关性黄斑病变的机制

陈晓阳，王子祎，谢亚昊，靳琪，刘思宁，安佳怡，刘亚萌，强孟泽，李家赓，陈茁茁，张元元，于嘉琛，杨志强   

  1. 1. 河北北方学院理学院，张家口，075000，中国 

    2. 河北北方学院药学院，张家口，075000，中国 

    3. 康泰医学检验服务河北有限公司，廊坊，065000，中国

  • 出版日期:2024-12-26 发布日期:2024-12-26
  • 通讯作者: 杨志强，硕士，研究方向：分子生物学；E-mail：916238625@qq.com
  • 作者简介:陈晓阳，硕士；研究方向：计算机辅助药物设计；E-mail：chenxiaoyang@hebeinu.edu.cn
  • 基金资助:
    河北省教育厅青年基金项目（No.JYT2024021），2023年大学生创新创业训练项目（No.202310092006），河北北方学院校级项 目（No.XJ2021028），河北省卫生厅项目（No.20200487），河北省自然科学基金资助项目（No.2023405003）

To Investigate the Mechanism of Taurine for Age-Related Macular Degeneration Based on Network Pharmacology and Molecular Docking

CHEN Xiao-yang, WANG Zi-yi, XIE Ya-hao, JIN Qi, LIU Si-ning, AN Jia-yi, LIU Ya-meng, QIANG Meng-ze, LI Jia-geng, CHEN Zhuo-zhuo, ZHANG Yuan-yuan, YU Jia-chen, YANG zhi-qiang   

  1. 1. College of Science, Hebei North University, Zhangjiakou, 075000, China 

    2. Department of Pharmacy, Hebei North University, Zhangjiahou, 075000, China 

    3. Kangtai Medical Laboratory Service Hebei Co., LTD., Langfang, 065000, China

  • Online:2024-12-26 Published:2024-12-26

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摘要：

目的：利用网络药理学及分子对接技术并通过体外实验探究牛磺酸（Taurine）对治疗年龄相关性黄斑病 变（age-related macular degeneration，AMD）的作用及机制。方法：利用不同数据库获取并筛出Taurine 和AMD 的交集靶点。通过蛋白互作用网络（protein-protein interaction networks，PPI）等探究Taurine 对AMD 作用机制， 并由分子对接技术初步验证。体外实验采用增殖检测试剂盒（CCK-8）、β 半乳糖苷酶试剂盒、蛋白质免疫印记、 ELISA 试剂盒、RT-qPCR 法验证Taurine 通过SRC-P53-MPO 轴抑制炎症对ARPE-19 细胞有抗衰老作用。结 果：通过网络药理学分析，Taurine 和AMD 的靶点交集过后包括了类固醇受体辅助活化因子（steroid receptor coactivating protein，SRC）、白细胞介素-6（interleukin-6，IL-6）、肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α） 等炎症相关因子以及肿瘤蛋白53（tumor protein 53，P53）、髓过氧化物酶（myeloperoxidase，MPO）等细胞衰老 的相关靶点；基因本体(gene ontology，GO) 生物功能以及京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes，KEGG) 通路富集分析的结果显示其包含了衰老炎症相关的信号通路。β- 半乳糖苷酶染色 结果与CCK-8 实验结果表明H2O2（100 μmol·L-1）作用于APRE-19 细胞培养24 h 较为合适，Taurine 10、20、40 mmol·L-1 为适合浓度。WB 结果显示，与模型组相比，Taurine 组SRC 明显上升，P53、MPO 明显下降。PCR 结 果显示，与模型组比较，Taurine 组SRC mRNA 水平明显上升，P53、MPO mRNA 水平明显下降。ELISA 结果显 示，与模型组相比，Taurine 组IL-6、TNF-α 明显下降。结论：通过网络药理学、分子对接及体外细胞实验证实了 Taurine 通过SRC-P53-MPO 轴抑制炎症对ARPE-19 细胞有抗衰老作用，为进一步阐释Taurine 治疗AMD 的作 用机制提供了科学依据。

关键词: 牛磺酸, 年龄相关性黄斑病变, 分子对接, 网络药理学, P53 靶点

Abstract:

Objective: To investigate the effect and mechanism of Taurine in the treatment of age-related macular degeneration (AMD) by using network pharmacology and molecular docking technology and in vitro experiments. Methods: The intersection targets of Taurine and AMD were obtained and screened from different databases. Through the Protein Interaction Network (PPI). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG Enrichment Analysis) signal pathway analysis, according to Cytoscape to establish a “drug-target-pathway” network to deeply explore the mechanism of Taurine action on AMD. Preliminary verification by molecular docking technology. The model of ARPE-19 cell senescence was established by H2O2 in vitro. Cell proliferation assay kit (CCK-8) was used to detect the optimal concentration and action time of H2O2 and Taurine. The senescence degree was detected by β-galactosidase kit staining. The levels of steroid receptor co-activating protein (SRC), tumor protein 53 (P53) ，and myeloperoxidase (MPO) were detected by Western blotting (WB) assay. The expression levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were detected by ELISA kit. Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) was used to analyze the expression levels of target genes SRC, P53 and MPO mRNA. Results: Network pharmacological analysis showed that the intersection of Taurine and AMD targets included inflammation-related factors such as SRC, IL-6, TNF-α, and cell senescence related targets such as P53 and MPO. The results of GO biological function and KEGG pathway enrichment analysis show that it contains signaling pathways related to aging inflammation. The results of β-galactosidase staining and CCK-8 experiment showed that the optimal concentration of H2O2 on APRE-19 cells was 100μmol·L-1 for 24 h, and Taurine 10 mmol·L-1, 20 mmol·L-1 and 40 mmol·L-1 were suitable concentrations. WB results showed that SRC was significantly increased and P53 and MPO were significantly decreased in Taurine group compared with model group. PCR results showed that compared with the model group, mRNA level of SRC was significantly increased, and P53 and MPO mRNA levels were significantly decreased in Taurine group. ELISA results showed that IL-6 and TNF-α were significantly decreased in Taurine group compared with model group. Conclusion: Network pharmacology, molecular docking and cell experiments were used to confirm that Taurine has an anti-aging effect on ARPE-19 cells by inhibiting inflammation through the SRCP53- MPO axis. It provides a scientific basis for further elucidation of the mechanism of action of taurine in the treatment of AMD.

Key words: Taurine, age-related macular degeneration, molecular docking, network pharmacology

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