摘要
目的观察强噪声暴露后耳蜗基底膜组织巨噬细胞形态的变化,探讨噪声性耳蜗损伤的免疫机制。方法将C57BL/6J小鼠接触持续噪声暴露(1-7k Hz,120d B SPL)1小时。应用电位反应测听仪,检测噪声暴露前和噪声暴露后10天不同频率短纯音(4、8、16和32 k Hz)诱发的动物双耳听性脑干反应阈值(ABR)。噪声暴露后1、4和10天处死动物,解剖取双侧耳蜗。采用鬼笔环肽(phalloidin)染色噪声暴露后10天毛细胞纤毛、表皮板的丝状肌动蛋白。白细胞共同抗原(CD45)荧光免疫抗体标记噪声暴露后1、4和10天耳蜗基底膜免疫细胞。以未接触噪声暴露动物耳蜗为对照,荧光显微镜下观察噪声暴露后小鼠耳蜗基底膜毛细胞和巨噬细胞形态变化,自耳蜗顶回到底回计数全耳蜗基底膜缺失的毛细胞和CD45荧光染色阳性细胞。结果噪声暴露后10天,不同频率短纯音诱发的ABR阈值均升高(F=1622.754,df=1,104,P<0.001;Tukey test,P<0.001)。耳蜗基底膜外毛细胞缺失数目明显多于正常对照组,底回缺失的外毛细胞数目多于顶回(F=92.484,df=1,40,P<0.001;Tukey test,P<0.001)。荧光显微镜下观察,生理条件下CD45阳性细胞主要为巨噬细胞,巨噬细胞分布于全耳蜗基底膜底面(鼓阶面)。细胞呈现多种形态,不同形态与其在耳蜗的不同部位相关。噪声暴露后1天,单核细胞渗入耳蜗基底膜,主要分布于耳蜗基底膜底回的上部。噪声暴露后4天,侵润的单核细胞转化为巨噬细胞,耳蜗基底膜CD45阳性细胞数目明显增加(F=15.205,df=3,46,P<0.001;Tukey test,P<0.001),耳蜗底回CD45阳性细胞数目多于顶回(P<0.05)。噪声暴露后10天,耳蜗基底膜CD45阳性细胞数目减少至噪声暴露前水平。结论免疫细胞参与了噪声性耳蜗损伤的反应,单核细胞的移入和转化可能在耳蜗细胞损伤和修复中起到重要的作用。
Objective To investigate the immune mechanism of cochlear damage induced by noise exposure,we observed changes in morphology of tissue macrophages beneath the cochlear basilar membrane following intense noise exposure.Methods C57BL/6J mice(4-8 weeks)were randomly divided into a control and a noise exposure group.Mice in the noise exposure group were exposed to a broadband noise(1-7 kHz)at 120 dB SPL for 1 hour.The auditory brain-stem response(ABR)thresholds elicited by tone bursts at 4,8,16 and 32 kHz were measured in both ears,and missing outer hair cells were quantified before and 10 days after the noise exposure.Leukocytes were visualized using immunolabeling of CD45 and counted across the apical and basal sections of the basilar membrane before and 1,4 and 10 days after the noise exposure under a fluorescence microscope.Results The noise exposure group exhibited a significant elevation of ABR thresholds at all tested frequencies after noise exposure(P<0.001).At 10 days post-noise exposure,the average number of missing cells in noise-injured ears was significantly higher than in control ears(P<0.001),with more cells missing in the basal section than in the apical section of the cochlea(P<0.001).CD45 staining revealed that tissue macrophages were present on the surface of the scala tympani side of the basilar membrane under physiological conditions and demonstrated different morphological features in the apical and basal sections.Monocytes infiltrated into the basilar membrane region on day 1 and transformed into macrophages 4 days after noise exposure across the whole basilar membrane,although primarily in the upper basal section(P<0.05),where greater sensory cell damage was present.Compared with the normal cochlea,the number of CD45 positive cells was increased at 4 days post-noise exposure(P<0.001),falling down to pre-noise levels on day 10.Conclusion Macrophages are involved in noise-induced cochlear injury responses,and monocyte infiltration and transformation may play an important role in the damage and repair of cochlear cells.
作者
杨卫平
许阳
胡博华
董有毅
杨仕明
YANG Weiping;XU Yang;HU Bohua;DONG Youyi;YANG Shiming(Department of Otolaryngology and Head,Neck Surgery,Institute of Otolaryngology,Chinese PLA General Hospital,Beijing,100853 China;Department of Respiratory Diseases,Chinese PLA General Hospital,Beijing 100853,China;Center for Hearing and Deafness,State University of New York at Buffalo,Buffalo,NY 14214,USA)
出处
《中华耳科学杂志》
CSCD
北大核心
2018年第1期73-79,共7页
Chinese Journal of Otology
基金
国家自然基金资助项目(No:30973304),美国NIH资助项目(NIDCDRO1DC010154)
关键词
噪声
耳蜗
巨噬细胞
Noise
Cochlea
Macrophages
作者简介
通讯作者：杨卫平:医学硕士，研究员，研究方向：主要从事聋病的发生机制和防治研究.Email:yangwp301@126.com;第一作者:许阳
