靶向Notch信号通路与内耳发育相关microRNA的筛选及microRNA-384-5p的实验分析  

Screening of microRNAs targeting Notch signaling pathway implicated in inner ear development and the role of microRNA-384-5p

作　　者：陈智斌[1] 浦懋懋 姚俊[2] 曹新[2] 程雷[1] Chen Zhibin;Pu Maomao;Yao Jun;Cao Xin;Cheng Lei(Department of Otorhinolaryngology,the First Affiliated Hospital of Nanjing Medical University,Nanfing 210029,China;Department of Biotechnology,School of Basic Medical Science,Nanfing Medical University,Nanjing 211166,China)

机构地区：[1]南京医科大学第一附属医院耳鼻咽喉科,210029 [2]南京医科大学基础医学院生物技术系,211166

出　　处：《中华耳鼻咽喉头颈外科杂志》2018年第11期830-837,共8页Chinese Journal of Otorhinolaryngology Head and Neck Surgery

基　　金：江苏省重点研发计划(社会发展)项目(BE2016762)。

摘　　要：目的筛选内耳中靶向调控Notch信号通路的关键microRNA,验证其对Notch信号通路的调控作用。方法构建Notch信号通路相关基因与microRNA相互作用网络,筛选核心基因并提取Core-Notch子网络,再通过拓扑学分析与GO分析筛选对Notch信号通路具有调控作用的关键microRNA,并在体内、外水平进行验证。结果以筛选的microRNA-384-5p(miR-384-5p)为研究对象,预测其对Notch信号通路具有较强调控作用。实验结果显示,miR.384-5p在小鼠脑与耳蜗组织中特异性表达;miR-384-5p-mimic转染HeLa细胞后,Notch1的表达水平显著下调;双荧光素酶报告基因检测进一步验证了miR-384-5p对Notch信号通路中Notch1和Dll4的负调控作用。结论基于构建的Core-Notch网络筛选内耳发育相关的microRNA,筛选出来的miR-384-5p分子可靶向调控Notch信号通路,可作为潜在的毛细胞修复的干预靶点。Objective To screen the key microRNAs targeting Notch signaling pathway in inner ear and investigate its potential regulating function .Methods The interaction network and the Core-Notch network, involved with key genes in Notch signal pathway and differential-expressed microRNAs in inner ear, were constructed by bioinformatics methods .The important microRNAs in regulating Notch signaling pathway were screened via topological and GO analysis, followed by in vivo and in vitro investigation.Results MiRNA-384-5p was identified as a key regulator specifically expressed in mouse brain and inner ear , which could down-regulate Notch1.The Notch1 expression was found significantly down-regulated in miRNA-384-5p-mimic-transfected HeLa cells.The dual-luciferase reporter gene assay further confirmed the effect of miRNA-384-5p on the down-regulation of Notch1 and Dll4 in Notch signaling pathway.Conclusions The Core-Notch network is constructed to screen microRNAs implicated in inner ear development , and miRNA-384-5p is screened and verified to be target-regulating the Notch signaling pathway , which could be the potential target in the regeneration of impaired hair cells .

关 键 词：受体 Notch 耳 内 毛细胞 听觉 miR-384-5p 

分 类 号：R764[医药卫生—耳鼻咽喉科]