| 周 智,姚 冰,王孟清,罗银河,胡 燕,丁 伊,邓羿駃,刘靖雷,马 啸.病毒诱发哮喘关键miRNA靶基因的预测与验证及潜在中药干预[J].,2024,(6):1019-1029 |
| 病毒诱发哮喘关键miRNA靶基因的预测与验证及潜在中药干预 |
| Prediction and Validation of Key MiRNA Target Genes for Virus-induced Asthma and Potential Traditional Chinese Medicine Interventions |
| 投稿时间：2023-10-14 修订日期：2023-11-12 |
| DOI：10.13241/j.cnki.pmb.2024.06.004 |
| 中文关键词: 儿童哮喘 呼吸道合胞病毒 靶基因 生物信息学 中医药 数据挖掘 |
| 英文关键词: Childhood asthma Respiratory syncytial virus Target gene Bioinformatics Traditional Chinese Medicine Data mining |
| 基金项目:国家自然科学基金项目（81774368；82174437）；全国名老中医药专家传承工作室(国中医药人教函〔2022〕75号)；湖南省自然科学基金项目（2022JJ30037；2023JJ60260）；湖南中医药大学中医儿科学湖南省"双一流"学科建设项目（教高厅函〔2022〕14号） |
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| 中文摘要: |
| 摘要 目的：探究病毒诱发哮喘关键微小RNA(miRNA)以及差异基因在哮喘发病中的调控作用，并寻找潜在的治疗靶点以及对应中药，为病毒诱发哮喘中医药辨证论治、早期截断哮喘的发生、传变提供诊疗思路。方法：首先通过查阅有关miRNA与哮喘关系的文献，根据文献的影响因子、报道次数及与呼吸道合胞病毒（Respiratory syncytial virus, RSV）、树突细胞、自噬等病毒诱发哮喘发病机制的密切程度进行积分统计，按得分高低筛选出6个候选miRNA：miRNA-155、miRNA-21、Let-7、miRNA-34a、miRNA-145、miRNA-146a。然后通过靶基因数据库分析其功能，并通过生物信息学方法筛选出与关键目标miRNA相关的信号通路。最后通过Coremine数据库查找核心基因对应中药，使用TCMSP、BATMAN-TCM数据库查找中药有效成分，利用Cytoscape软件进行可视化分析，并进行基因本体论（Gene ontology,GO）富集分析与基因组百科全书（Kyoto encyclopedia of genes and genomes，KEGG）通路富集分析。然后通过RSV联合鸡卵清蛋白诱导病毒诱发哮喘模型，并通过实时荧光定量PCR( qRT-PCR) 法检测生物信息学预测的核心miRNA表达水平。结果：体内实验验证结果显示：RSV诱导哮喘小鼠模型中气道上皮miRNA-21、Let-7、miRNA-155、miRNA-146a表达量显著降低，提示这四种miRNA可作为病毒诱发哮喘关键生物标志物；结果中共筛选出与病毒诱发哮喘最相关的10个核心靶基因，分别是：STK4、CTH、MRM3、EEF1A1、PDCD6IP、MARCKS、SLC25A5、GNAS、ALDOA、CFL1，也是治疗的关键靶点；一共包含对应中药共160多种，其中理气活血类药物、补虚药出现频次最高，具体的高频中药为：枳实、枳壳、香橼、油柑子、人参、人参叶、姜黄、郁金、丁香、水牛角。结论：MiRNA能够通过靶向调控多种靶基因的表达，参与病毒疾病的发生、发展过程。 RSV诱导哮喘气管组织中的miRNA-21、Let-7、miRNA-155、miRNA-146a均有显著下调，可作为病毒诱发哮喘关键生物标志物，也是治疗的关键靶点，同时能够从miRNA这一角度明确在病毒诱发哮喘中的中医药作用机制，可通过辨证论治适当使用理气活血类、补虚药等药物，以早期截断哮喘的病理进程。 |
| 英文摘要: |
| ABSTRACT Objective: To explore the regulatory role of key microRNAs (miRNAs) and differential genes in the pathogenesis of asthma induced by viruses, and to find potential therapeutic targets and corresponding traditional Chinese medicines, so as to provide diagnostic and therapeutic ideas for the differentiation and treatment of virus-induced asthma, the occurrence and transmission of early truncated asthma. Methods: First, by reviewing the literature on the relationship between miRNA and asthma, six candidate miRNAs were selected according to the impact factor, number of reports, and the closeness of the pathogenesis of asthma induced by respiratory syncytial virus (RSV), dendritic cells, autophagy and other viruses, and six candidate miRNAs were selected according to their scores: miRNA-155, miRNA-21, Let-7, miRNA-34a, miRNA-145, miRNA-146a. Its function was then analyzed by a database of target genes, and signaling pathways associated with key target miRNAs were screened out by bioinformatics methods. Finally, the core genes corresponding to traditional Chinese medicines were found through the Coremine database, the active ingredients of traditional Chinese medicines were found by TCMSP and BATMAN-TCM databases, and the Cytoscape software was used for visual analysis, and the enrichment analysis of gene ontology (GO) and the enrichment analysis of the Kyoto encyclopedia of genes and genomes (KEGG) pathway were carried out. Then, RSV combined with chicken egg albumin-induced virus-induced asthma model, and the expression level of core miRNA predicted by bioinformatics was detected by real-time real-time PCR (qRT-PCR). Results: In vivo experiments showed that the expression levels of miRNA-21, Let-7, miRNA-155 and miRNA-146a in the airway epithelium in RSV-induced asthma mouse models were significantly reduced, suggesting that these four miRNAs could be used as key biomarkers of virus-induced asthma. ResultsA total of 10 core target genes most related to virus-induced asthma were screened, namely: STK4, CTH, MRM3, EEF1A1, PDCD6IP, MARCKS, SLC25A5, GNAS, ALDOA, and CFL1, which were also the key targets for treatment. There are more than 160 kinds of corresponding traditional Chinese medicines, among which the frequency of qi regulating and activating blood drugs and deficiency tonic drugs is the highest, and the specific high-frequency Chinese medicines are: citrus aurantium, citrus aurantium, citron, citrus oil, ginseng, ginseng leaves, turmeric, tulip, cloves, and buffalo horn. Conclusion: MiRNAs can regulate the expression of a variety of target genes by targeting and participating in the occurrence and development of viral diseases. RSV-induced asthma-induced miRNA-21, Let-7, miRNA-155 and miRNA-146a were significantly down-regulated, which can be used as a key biomarker of virus-induced asthma and a key target for treatment. At the same time, the mechanism of action of traditional Chinese medicine in virus-induced asthma can be clarified from the perspective of miRNA, and the pathological process of asthma can be truncated at an early stage through syndrome differentiation and treatment. |
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