听力损失中表达上调的miR-196a-5p靶向抑制Neuritn的表达  

Up-regulated miR-196a-5p can target the expression of Neuritin in hearing loss

作　　者：宋银 张雪[1] 孙嘉伟 黄娟 胡金池 杨磊[2] 黄瑾[1] SONG Yin;ZHANG Xue;SUN Jiawei;HUANG Juan;HU Jinchi;YANG Lei;HUANG Jin(Medical College of Shihezi University,Shihezi,Xinjiang 832000,China;Department of Medicine,Hangzhou Normal University,Hangzhou,Zhejiang 310036,China)

机构地区：[1]石河子大学医学院,新疆石河子832003 [2]杭州师范大学医学部,浙江杭州310036

出　　处：《石河子大学学报（自然科学版）》2021年第3期352-358,共7页Journal of Shihezi University(Natural Science)

基　　金：国家科技重大专项(民口)重大新药创制(2019ZX09301-161);杭州市重大科技创新项目(20152013A01)。

摘　　要：目的筛选鉴定听力损失中上调且靶向调控Neuritin的miRNA。方法构建CBA小鼠药物性听力损失模型(硫酸卡那霉素+呋塞米),采用高通量测序检测听力损失中差异表达的miRNA。利用生物信息学技术,分析预测靶向Neuritin的miRNA,取测序结果中高表达miRNA与靶向Neuritin的miRNA交集得到候选miRNA。使用实时荧光定量PCR(RT-qPCR)明确听力损失中候选miRNA高表达,使用miRNA mimics转染293T细胞,观察目标miRNA对Neuritin表达的靶向抑制作用。结果通过高通量测序与生物信息学分析获得3个在听力损失中表达升高且可能靶向调控Neuritin的候选miRNA,即miR-196a-5p,miR-211-5p和miR-6395。RT-qPCR实验发现miR-196a-5p在小鼠听力损失12h与24h后在Corti器中的均表达升高(P<0.05);miR-6395在小鼠听力损失后12h表达降低,24h后表达升高(P<0.05);miR-211-5p在小鼠听力损失后12h与24h表达均降低(P<0.05)。Western Blot实验发现转染miR-196a-5p后293T细胞的Neuritin表达下降(P<0.05)。结论听力损失中表达上调的miR-196a-5p可靶向抑制Neuritin的表达。Objective Screen and identify miRNAs up-regulated in hearing loss and target Neuritin.Method First,a CBA mouse model of drug-induced hearing loss(kanamycin sulfate+furosemide)was constructed,and then high-throughput sequencing was used to detect the differentially expressed miRNAs in hearing loss.Candidate miRNAs were obtained by intersecting miRNAs between targetingNeuritin miRNAs and high-expressing miRNAs in the sequencing result.Real-time fluorescent quantitative PCR(RT-qPCR)was used to clarify the authenticity of the high expression of candidate miRNAs in hearing loss,and miRNA mimics were transfected into 293 T cells to observe the targeted inhibition of the target miRNA on Neuritin expression.Results Through high-throughput sequencing and bioinformatics analysis,three candidate miRNAs with elevated-expression in hearing loss and with possibly targeted-regulation of Neuritin,miR-196a-5p,miR-211-5p and miR-6395,were obtained.RT-qPCR showed that the expression of miR-196a-5p ncreased after 12 h and 24 h in the organ of Corti after hearing loss in mice(P<0.05);the expression of miR-6395 decreased at 12 h and 24 h after hearing loss in mice.The expression increased afterwards(P<0.05);the expression of miR-211-5p decreased at 12 h and 24 h after hearing loss in mice(P<0.05).The expression of Neuritin was decreased in 293 T cells transfected with miR-196a-5p(P<0.05).Conclusion Increased miR-196a-5p target to inhibit Neuritin’s expression in hearing loss.

关 键 词：NEURITIN miRNA 听力损失 生物信息学 高通量测序 

分 类 号：R764[医药卫生—耳鼻咽喉科]