基于miR-34a/Notch信号通路研究补肺益肾方对慢性阻塞性肺疾病大鼠模型免疫失衡和炎症反应的影响  

Effect of Bufei Yishen Fang on immune imbalance and inflammatory response in COPD rats based on miR-34a/Notch signaling pathway

作　　者：栾英[1] 李敬蕊[1] 刘林林 高春燕[1] 张菊香[1] 王晶[1] 崔朝勃[1] LUAN Ying;LI Jingrui;LIU Linlin;GAO Chunyan;ZHANG Juxiang;WANG Jing;CUI Zhaobo(Hengshui Harrison International Peace Hospital,Hengshui 053000,China)

机构地区：[1]河北省衡水市哈励逊国际和平医院衡水市人民医院,衡水053000

出　　处：《中国免疫学杂志》2022年第9期1075-1081,共7页Chinese Journal of Immunology

基　　金：2018年度河北省医学科学研究重点课题计划(20181596)。

摘　　要：目的:观察补肺益肾方(BYF)对慢性阻塞性肺疾病(COPD)大鼠免疫功能及炎症反应的影响,并探讨微小RNA-34a(miR-34a)/果蝇双翅边缘缺刻同源基因(Notch)信号轴在其中的调控作用。方法:取SD大鼠72只,分为正常对照组、模型组、BYF组(3.7 g/kg)、miR-34a低表达组(miR-34a antagomir,4 nmol/kg)、miR-34a阴性对照组(antagomir-NC)、BYF^(+)miR-34a antagomir组(3.7 g/kg^(+)4 nmol/kg),每组12只;除正常对照组外,其余各组均用香烟暴露^(+)肺炎克雷伯杆菌感染法制备COPD模型。于造模成功后开始给药。观察大鼠一般行为;检测大鼠肺功能指标:用力肺活量(FVC)、第0.3秒用力呼气容积(FEV_(0.3))、肺阻力(R)、肺顺应性(Cdyn);血液分析系统检测肺泡灌洗液中炎症细胞数量;流式细胞仪检测外周血CD4^(+)/CD8^(+)、辅助性T细胞17(Th17)/调节性T细胞(Treg)水平;qRT-PCR检测肺组织miR-34a、Notch mRNA表达;HE染色检测肺组织病理变化;Western blot检测肺组织Notch、IL-17、IL-6、Th17转录因子RoRγt和Treg转录因子Foxp3水平。结果:与正常对照组相比,模型组大鼠死亡、喘促、肺泡结构破坏严重,肺功能指标R、肺泡灌洗液白细胞、淋巴细胞和巨噬细胞数量、血清CD8^(+)/CD4^(+)、Th17/Treg、肺组织Notch mRNA及蛋白、IL-17、IL-6、RoRγt、Foxp3、RoRγt/Foxp3蛋白表达升高(P<0.05),FVC、FEV_(0.3)、Cdyn水平和miR-34a表达降低(P<0.05)。与模型组相比,BYF组大鼠死亡、喘促、肺泡结构破坏缓解,免疫功能趋于平衡,肺功能及miR-34a表达升高,炎症细胞数量、Notch相关通路蛋白表达降低,且上述指标变化差异均有统计学意义(P<0.05);miR-34a antagomir组大鼠死亡、喘促、肺泡结构破坏进一步加重,肺功能及免疫功能损伤进一步加重,miR-34a/Notch轴及相关指标变化与模型组一致,且差异有统计学意义(P<0.05)。BYF^(+)miR-34a antagomir组大鼠上述指标与BYF组相比结果均相反(P<0.05)。AntagomirNC组与模型组相比�Objective:To observe the effects of Bufei Yishen Fang(BYF)on immune function and inflammatory response in rats with chronic obstructive pulmonary disease(COPD),and to investigate the regulation of microRNA-34a(miR-34a)/Drosophila double-wing margin nicked homologous gene(Notch)signal axis.Methods:Seventy-two SD rats were randomly divided into normal control group,model group,BYF group(3.7 g/kg),low-expressed miR-34a group(BYF+miR-34a antagomir,4 nmol/kg),BYF+miR-34a antagomir group(3.7 g/kg+4 nmol/kg)and miR-34a negative control(antagomir-NC)group,with 12 rats in each group;except the normal control group,COPD model was established by cigarette exposure and Klebsiella pneumoniae infection.The drug was administered after the successful modeling.The general behavior of rats was observed;the indexes of lung function in rats:the forced vital capacity(FVC),expiratory volume(FEV_(0.3)),pulmonary resistance(R)and culmonary dynamic compliance(Cdyn)were measured;the number of leukocytes(WBC),inflammatory cells in BALF were detected by blood analysis system;peripheral blood samples were collected and the levels of T lymphocyte subsets(CD4+/CD8+,helper T cell 17(Th17)/regulatory T cell(Treg)were detected by flow cytometry;the expression of miR-34a and Notch mRNA was detected by real-time quantitative fluorescence PCR(qRT-PCR);the pathological changes of lung tissue were detected by hematoxylin eosin(HE)staining;and the levels of Notch,IL-17,IL-6,RoRγt and Foxp3 were detected by Western blot.Results:Compared with normal control group,the death,dyspnea with rapid and short breath,and the destruction of alveolar structure were serious in the model group,the lung function index R,the numbers of leukocytes,lymphocytes and macrophages in BALF,expression of CD8+/CD4+,Th17/Treg in serum,Notch mRNA and protein,and IL-17,IL-6,RoRγt,Foxp3,RoRγt/Foxp3 protein in lung tissue were increased(P0.05).Conclusion:BYF may up-regulate the expression of miR-34a,inhibit the activation of Notch pathway,and improve the inflammatory response and

关 键 词：补肺益肾方 慢性阻塞性肺疾病 微小RNA-34a 果蝇双翅边缘缺刻同源基因 免疫失衡 炎症反应 

分 类 号：R563.1[医药卫生—呼吸系统]