内吞相关蛋白Myosin Ⅵ/Disabled-2在小鼠耳蜗毛细胞的表达及年龄相关性变化的研究  

The relation between age and expression of Myosin Ⅵ and Disabled-2 in the cochlea of mice

作　　者：陈知己 顾翔[1] 蔡婷[1] 宋锐[1] 周小清[1] 袁伟[1] 

机构地区：[1]第三军医大学附属西南医院耳鼻咽喉-头颈外科重庆,400038

出　　处：《临床耳鼻咽喉头颈外科杂志》2015年第7期645-649,共5页Journal of Clinical Otorhinolaryngology Head And Neck Surgery

基　　金：国家自然科学基金资助项目(No:81470694;81271080;30973301);重庆市自然基金资助项目(No:CSTC2009BB5170);第三军医大学基金项目(No:2010XLC06)

摘　　要：目的:研究不同年龄小鼠耳蜗毛细胞内吞相关蛋白MyosinⅥ和Disabled-2(Dab2)的表达及分布特点。方法:分别选取出生后2周、5周、9周及16月龄的昆明小鼠各10只,采用免疫荧光标记联合激光共聚焦显微镜扫描耳蜗基底膜铺片,观察MyosinⅥ及Dab2蛋白的表达定位,通过荧光定量逆转录PCR测定并比较不同年龄组小鼠耳蜗2种蛋白mRNA的表达水平。结果:MyosinⅥ及Dab2蛋白在内外毛细胞均有表达,支持细胞未见表达,且内毛细胞2种蛋白均存在局部高表达的亚细胞区域。比较不同年龄组MyosinⅥ荧光染色图像后发现,2周龄和16月龄小鼠荧光强度均低于5周龄和9周龄小鼠。比较不同年龄组小鼠2种蛋白mRNA表达后发现,2周龄小鼠mRNA相对表达水平均低于9周龄小鼠,差异有统计学意义(P<0.01),5周龄及16月龄组小鼠分别与9周龄小鼠相比,差异均无统计学意义(P>0.05)。结论:调控网格蛋白介导内吞过程的MyosinⅥ及Dab2蛋白表达于内外毛细胞,内毛细胞顶区表皮板及附近胞质可能是这一过程的主要部位。随着年龄的增长,MyosinⅥ及Dab2蛋白的表达可能会发生改变,网格蛋白介导内吞过程效率可能因此相继发生变化,内毛细胞功能损伤可能与此有关。Objective：To investigate the expression of MyosinⅥ and Disabled-2（Dab2）in the cochlea of mice at different ages.Method：Forty KM mice were divided into four groups according to age,named as postnatal 2week（P2w）,P5 w,P9w,P16 month.The localization of protein in the basilar membrane of mice cochlea was detected by immunofluorescence staining and laser scanning confocal microscope（LSCM）.The mRNA expression level of protein in cochlear at different ages was evaluated by real-time fluorescent quantitative reverse transcriptase-polymerase chain reaction（qRT-PCR）.Statistical analysis was performed by the SPSS18.0software.Result：MyosinⅥ and Disabled-2protein mainly expressed at the apical cytoplasm of hair cells.As for the inner hair cell,Dab2 labeling was abundant especially at the cuticular plate and nearby.Comparing four immunofluorescence staining images of MyosinⅥ,we found the fluorescence intensity of P2 wand P16m were weaker than that of P5 w and P9 w.After setting P9 was the control group,qRT-PCR revealed that the mRNA expression of MyosinⅥ and Dab2 in P2w was less than that in the control group（P〈0.01）,while no significant difference was found between P5 wand the control group,nor between P16 mand the control group（P〉0.05）.Conclusion：MyosinⅥand Dab2,two proteins which regulated the clathrin-mediated endocytosis,expressed at hair cells of mice cochlea.In the inner hair cell,this process of endocytosis may be more efficient at the cuticular plate and nearby.The expression level of protein may change in different ages,and this probably leads to a difference of CME,it also may cause a defect of inner hair cells function.

关 键 词：内毛细胞 网格蛋白介导内吞 囊泡 激光共聚焦显微镜扫描技术 

分 类 号：R945.7[医药卫生—微生物与生化药学]