质膜钙泵1和质膜钙泵3在发育期大鼠耳蜗毛细胞的动态表达  

Dynamic Expression of PMCA1 and PMCA3 on Hair Cells in Developing Cochlea in Rats

作　　者：陈请国 褚汉启[1] 陈金 刘云[1] 杜智会 周良强[1] CHEN Qingguo;CHU Hanqi;CHEN Jin;LIU Yun;DU Zhihui;ZHOU Liangqiang(Department of OtolaryngologyHead and Neck Surgery,Tongji Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430030,China)

机构地区：[1]华中科技大学同济医学院附属同济医院耳鼻咽喉头颈外科,武汉430030

出　　处：《中华耳科学杂志》2025年第2期261-266,共6页Chinese Journal of Otology

基　　金：国家自然科学基金青年基金项目(81300827);湖北省自然科学基金面上项目(2023AFB686,2022CFB282)。

摘　　要：目的探究质膜钙泵1(plasma membrane calcium pumps1,PMCA1)和PMCA3在发育期大鼠耳蜗毛细胞的动态表达。方法麻醉后取出生2(P2)、6(P6)、13(P13)、21(P21)d的大鼠耳蜗,固定后分离出基底膜做铺片,通过定量免疫荧光标记法检测PMCA1和PMCA3在基底膜底回、中回、顶回外毛细胞(outer hair cells,OHCs)和内毛细胞(inner hair cells,IHCs)的表达部位和表达水平。用共聚焦显微镜捕获免疫荧光图片,使用Adobe Photoshop软件测量PMCA1和PMCA3的荧光强度,绘制年龄相关性表达曲线。结果在整个发育过程中,PMCA1在OHCs纤毛(OHCs hair bundle,OHCs-HB)、IHCs纤毛(IHCs hair bundle,IHCs-HB)均无表达。在出生早期,PMCA1在OHCs胞体(OHCs cell body,OHCs-CB)、IHCs胞体(IHCs cell body,IHCs-CB)侧膜的表达逐渐增强,P6后在OHCs-CB侧膜的表达逐渐下降,到成年期维持在微弱的水平;PMCA1在IHCs-CB侧膜的表达在P13达到高峰,维持在此水平进入成年期。在整个发育阶段,PMCA3在OHCs-HB、IHCs-HB均无表达,在出生6 d内的OHCs-CB、IHCs-CB侧膜均有较强的表达。P13后,PMCA3在OHCs-CB侧膜表达几乎消失,在IHCs-CB侧膜和表皮板下方项圈的表达虽然下降,但仍维持在一定水平进入成年期。P2、P6,PMCA1、PMCA3在OHCs-CB、IHCs-CB侧膜表达水平在基底膜底回最高,其次为中回、顶回,P13后,差异性消失。结论在大鼠发育过程中,PMCA1、PMCA3在毛细胞不同部位呈现动态表达,PMCA1、PMCA3在OHCs-HB、IHCs-HB均无表达,在出生早期的OHCs-CB侧膜有短暂的表达,在整个时期的IHCs-CB侧膜持续表达。由于毛细胞主要依靠PMCA来调控胞内Ca^(2+)水平,因此推测PMCA1和PMCA3在毛细胞的差异性表达与PMCA泵在毛细胞不同区域发挥着特殊的功能有关。Objective To report dynamic expression of plasma membrane calcium pumps 1 and 3(PMCA1 and PMCA3)on hair cells in developing cochleae in rats.Methods Rat cochleae were collected on Days 2,6,13 and 21 after birth.After fixation,the basilar membrane was isolated and prepared.Expression sites and levels of PMCA1 and PMCA3 on outer(OHCs)and inner hair cells(IHCs)in all turns were detected by quantitative immunofluorescence labeling,and photographed with confocal microscopy.The Adobe Photoshop software was used to measure fluorescence intensities and age-related expression curves were plotted.Results No PMCA1 expression was found on OHC or IHC hair bundles.At early postnatal days,PMCA1 expression on OHC and IHC basolateral membrane gradually increased,followed by a gradual decrease on OHC basolateral membrane after day 6 of birth and remained at a weak level until adulthood.PMCA1 expression on IHC basolateral membrane peaked at day 13 and remained at this level into adulthood.While PMCA3 was not expressed on OHC and IHC hair bundles,it was strongly expressed on the basolateral membrane during the first 6 days of life.After P13,PMCA3 expression on OHC basolateral membrane had almost disappeared,while its expression on IHC basolateral membrane and pericuticular necklace remained into adulthood,albeit at a decreased level.PMCA1 and PMCA3 expression on OHC and IHC basolateral membranes were at the highest levels in the basal turn,followed by the middle and apical turn before day 6 after birth,although this difference disappeared after P13.Conclusion PMCA1 and PMCA3 expression demonstrate a dynamic pattern during postnatal development on different parts of cochlear hair cells.PMCA1 and PMCA3 are not expressed on OHC and IHC hair bundles,and only transiently expressed on OHC basolateral membrane in early postnatal days,but continuously expressed in IHC basolateral membrane throughout the whole period.Considering that hair cells rely mainly on PMCA to regulate intracellular Ca^(2+)levels,it is speculated that the differential

关 键 词：质膜钙泵1 质膜钙泵3 大鼠 耳蜗 毛细胞 动态表达 

分 类 号：R73[医药卫生—肿瘤]