摘要
目的:明确化湿败毒方对甲型H1N1流感病毒致小鼠肺炎的治疗作用,基于转录组学探讨其作用机制。方法:采用甲型H1N1流感病毒滴鼻感染建立小鼠流感病毒性肺炎模型,连续给药5 d,考察小鼠一般状况、肺指数、病毒载量、肺组织病理形态、生存时间、生存时间延长率等指标,明确化湿败毒方对流感病毒性肺炎的治疗作用。采用转录组学技术检测模型组与正常组、化湿败毒方组与模型组小鼠肺组织的差异表达基因,筛选化湿败毒方治疗流感病毒性肺炎的潜在核心靶点;采用实时荧光定量聚合酶链式反应(Real-time PCR)验证化湿败毒方对核心靶点基因mRNA表达水平的影响。结果:与正常组比较,模型组小鼠肺指数及肺组织内病毒载量均明显升高(P<0.05,P<0.01);与模型组比较,化湿败毒方高剂量组明显延长甲型流感病毒感染小鼠的生存时间(P<0.05),明显降低小鼠肺指数值(P<0.05),降低肺组织病毒载量,化湿败毒方高、中、低剂量组可明显减轻小鼠肺组织炎症、瘀血、肿胀等病变(P<0.05,P<0.01)。肺组织转录组学分析显示,化湿败毒方干预后核心基因主要富集在核转录因子-κB(NF-κB)信号通路、白细胞介素-17(IL-17)信号通路、细胞因子-细胞因子受体相互作用等通路,TNF受体相关因子6(TRAF6)、核转录因子-κB抑制因子α(NFKBIA)、趋化因子配体(CCL)2、CCL7、CXC趋化因子配体2(CXCL2)为combined score值前5的节点基因;Real-time PCR验证显示,化湿败毒方能明显下调NF-κB信号通路关键基因TRAF6、NFKBIA及趋化因子CCL2、CCL7、CXCL2 mRNA表达(P<0.05,P<0.01)。结论:化湿败毒方对流感病毒性肺炎具有治疗作用,可能通过抑制NF-κB信号通路关键节点TRAF6、NFKBIA表达,抑制趋化因子CCL2、CCL7、CXCL2的表达,减少炎症细胞的募集,降低病毒载量,发挥抗流感病毒肺炎作用。
Objective:To clarify the therapeutic effect of Huashi Baidu prescription on pneumonia in mice caused by influenza A(H1N1)virus and explore its mechanism based on the transcriptome.Method:A mouse influenza viral pneumonia model was built by intranasal infection with influenza A virus,and mice were continuously administered the drug for five days,so as to investigate the general condition,lung index,viral load,pathological morphology of lung tissue,survival time,and prolongation rate of survival time of mice and clarify the therapeutic effect of Huashi Baidu prescription on influenza viral pneumonia.Transcriptome technology was used to detect the differentially expressed genes in the lung tissue of mice in the model group and the normal group,as well as the Huashi Baidu prescription group and the model group,and the potential core target of the Huashi Baidu prescription for the treatment of influenza viral pneumonia was screened.Real-time fluorescence quantitative polymerase chain reaction(Real-time PCR)was used to verify the effect of Huashi Baidu prescription on the mRNA expression level of core target genes.Result:Compared with the normal group,the lung index and viral load in the lung tissue of the model group were significantly increased(P<0.05,P<0.01).Compared with the model group,the high-dose group of Huashi Baidu prescription significantly prolonged the survival time of mice infected with influenza A virus(P<0.05)and significantly reduced the lung index value of mice(P<0.05)and the viral load of lung tissue.The high-dose,medium-dose,and low-dose groups of Huashi Baidu prescription could significantly reduce lung tissue inflammation,blood stasis,swelling,and other pathological changes in mice(P<0.05,P<0.01).Transcriptome analysis of lung tissue showed that core genes were mainly enriched in the nuclear transcription factor-κB(NF-κB)signaling pathway,interleukin-17(IL-17)signaling pathway,cytokine-cytokine receptor interaction,and other pathways after the intervention of Huashi Baidu prescription.TRAF6,NFKBIA,CCL2,CCL7,and CXCL2 were the top five node genes with combined score values.Real-time PCR validation showed that Huashi Baidu prescription significantly downregulated the mRNA expression of key genes TRAF6 and NFKBIA in the NF-κB signaling pathway,as well as chemokines CCL2,CCL7,and CXCL2(P<0.05,P<0.01).Conclusion:Huashi Baidu prescription has a therapeutic effect on influenza viral pneumonia,possibly by inhibiting the expression of key nodes TRAF6 and NFKBIA in the NF-κB signaling pathway and that of chemokines CCL2,CCL7,and CXCL2,reducing the recruitment of inflammatory cells and viral load,and exerting anti-influenza viral pneumonia effects.
作者
单中超
孙建辉
李建良
于泽玥
郝莉雨
邓玉荣
霍海如
李洪梅
黄璐琦
SHAN Zhongchao;SUN Jianhui;LI Jianliang;YU Zeyue;HAO Liyu;DENG Yurong;HUO Hairu;LI Hongmei;HUANG Luqi(Academician Workstation of Jiangxi University of Traditional Chinese Medicine,Nanchang 330004,China;Institute of Chinese Materia Medica,China Academy of Chinese Medicine Sciences,Beijing 100700,China;National Resource Center for Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,China)
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2023年第18期54-61,共8页
Chinese Journal of Experimental Traditional Medical Formulae
基金
中国中医科学院科技创新工程项目(CI2021A04605)
中国中医科学院中央级公益性科研院所基本科研业务费项目(ZZ15-WT-04,ZZ15-WT-08)。
关键词
甲型流感病毒
化湿败毒方
病毒载量
转录组测序
差异表达基因
influenza A virus
Huashi Baidu prescription
viral load
transcriptome sequencing
differentially expressed genes
