miR-145/SMAD3轴在慢性阻塞性肺疾病中的作用和机制研究  

Role and mechanism of miR-145/SMAD3 axis in chronic obstructive pulmonary disease

作　　者：谢圆媛[1] 胡海峰[2] 赵莉[3] 薛鹏[4] XIE Yuan-yuan;HU Hai-feng;ZHAO Li;XUE Peng(Department of Geriatrics,Affiliated Hospital of Yan'an University,Yan'an 716000,China;Department of Oncology,Affiliated Hospital of Yan'an University,Yan'an 716000,China;Department of Cardiovascular Medicine,Affiliated Hospital of Yan'an University,Yan'an 716000,China;Department of Anesthesiology,Affiliated Hospital of Yan'an University,Yan'an 716000,China)

机构地区：[1]延安大学附属医院老年病科,陕西延安716000 [2]延安大学附属医院肿瘤科,陕西延安716000 [3]延安大学附属医院心血管内科,陕西延安716000 [4]延安大学附属医院麻醉科,陕西延安716000

出　　处：《实用药物与临床》2020年第12期1074-1081,共8页Practical Pharmacy and Clinical Remedies

基　　金：陕西省科技计划项目(2019ZP07258)。

摘　　要：目的探讨miR-145通过SMAD3调节慢性阻塞性肺疾病(COPD)的发生及机制。方法检测延安大学附属医院有COPD的吸烟者、无COPD的吸烟者、无COPD的未吸烟者肺组织中miR-145、SMAD3的表达水平。用20%CEM处理支气管上皮细胞后,构建COPD的细胞模型,将miR-145 mimics/inhibitor转染该细胞后,通过ELISA方法检测细胞上清中IL-6、IL-8、IL-10表达水平,RT-PCR、Western blot检测SMAD3、IL-6、IL-8、IL-10和ECM降解相关蛋白MMP-9、EMMPRIN以及P13K、Akt、mTOR的表达,CCK-8法检测细胞的增殖情况。通过双荧光素酶报告基因实验检测SMAD3与miR-145的靶向关系,同时,将pEGFP-N1-SMAD3转染COPD模型细胞后,RT-PCR、Western blot检测IL-6、IL-8、IL-10、MMP-9、EMMPRIN及P13K、Akt、mTOR的表达。结果有COPD的吸烟者肺组织中miR-145低表达、SMAD3高表达;miR-145能够抑制COPD模型细胞IL-6、IL-8、IL-10的表达,促进MMP-9、EMMPRIN的表达;miR-145通过抑制P13K/Akt/mTOR通路的发生,抑制细胞的增殖;最后,双荧光素酶报告基因实验检测miR-145靶向SMAD3;过表达SMAD3后,能够促进IL-6、IL-8、IL-10、P13K、Akt、mTOR的表达,抑制MMP-9、EMMPRIN的表达。结论miR-145抑制SMAD3的表达,从而抑制慢性阻塞性肺泡细胞炎症的发生,促进ECM的降解,通过抑制P13K/Akt/mTOR通路的发生抑制细胞的增殖,减轻COPD的发生。Objective To investigate the role of miR-145 in regulating chronic obstructive pulmonary disease(COPD)through SMAD3 and its mechanism.Methods The expression of miR-145 and SMAD3 in the lung tissues of smokers with COPD,smokers without COPD,and non-smokers without COPD were detected in our hospitals.Bronchial epithelial cells were treated with 20% CEM to construct COPD cell model.After miR-145 mimics/inhibitor were transfected into the cells,the expression levels of IL-6,IL-8,and IL-10 in the supernatant were detected by ELISA,and the expression of SMAD3,IL-6,IL-8,IL-10,MMP-9,EMMPRIN,P13K,Akt and mTOR were detected by RT-PCR and Western blot,and CCK-8 method was used to detect cell proliferation.Dual luciferase reporter gene assay was used to detect target relation between SMAD3 and miR-145.At the same time,pEGFP-N1-SMAD3 was transfected into model cells of COPD,and the expression of IL-6,IL-8,IL-10,MMP-9,EMMPRIN,P13K,Akt,mTOR were detected by RT-PCR and Western blot.Results The miR-145 was down-regulated and SMAD3 was overexpressed in the lung tissues of smoking patients with COPD;miR-145 could inhibit the expression of IL-6,IL-8 and IL-10,and promote the expression of MMP-9 and EMMPRIN.Moreover,miR-145 suppressed proliferation of bronchial epithelial cells by inhibiting P13K/Akt/mTOR pathway.Finally,dual luciferase reporter gene assay revealed that SMAD3 was the direct target of miR-145.Overexpression of SMAD3 promoted IL-6,IL-8,IL-10,P13K,Akt,mTOR expression,and inhibit MMP-9 and EMMPRIN expression.Conclusion The miR-145 inhibits the expression of SMAD3,inhibits the inflammation of chronic obstructive alveolar cells,promotes the degradation of ECM,and inhibits the cell proliferation by inhibiting the P13K/Akt/mTOR pathway,which ameliorates COPD.

关 键 词：MIR-145 SMAD3 支气管上皮细胞 P13K/Akt/mTOR 细胞炎症 ECM降解 

分 类 号：R563.9[医药卫生—呼吸系统]