NMF308~(nmf/nmf)小鼠耳蜗毛细胞凋亡方式及z-VAD-FMK对毛细胞的保护作用

Roles of Caspase Inhibitors in Cochlear Hair Cells Survival and Preventing Age-Related Hearing Loss

李胜利;王玉虎;张敏燕;李白芽;郑庆印;朱雯瑾;朱宏亮;

1:西安交通大学第二附属医院科研中心实验室药剂科

2:西安交通大学医学院神经科学研究中心

3:Case Western Reserve University School of Medicine

4:Department of Otolaryngology Head & Neck Surgery.Cleveland USA

摘要
目的了解年龄相关性听力损失小鼠耳蜗毛细胞的凋亡方式,探讨半胱氨酸蛋白酶(caspase)抑制剂z-VAD-FMK[z-Val-Ala-Asp(Ome)-fluoromethylketone]防治老年性聋的效果。方法选用新生NMF308nmf/nmf小鼠14只和成年同窝NMF308nmf/nmf小鼠32只,分为新生小鼠腹腔注射组(14只)、成年小鼠腹腔注射组(14只)和成年小鼠圆窗膜注射组(18只),各组又分为治疗组(注射z-VAD-FMK)和对照组[注射二甲基亚枫(dimethylsulfoxide,DMSO)],采用圆窗膜局部和腹腔注射两种方法给药,给药前后行ABR检测,用免疫荧光染色化学技术TUNEL、caspase-3和PI(碘化丙啶)染色标记耳蜗毛细胞,观察各组耳蜗毛细胞的凋亡和存活状况。结果NMF308nmf/nmf小鼠从1月龄开始发生听力减退和毛细胞功能改变,到2月龄时,caspase-3激活表达的毛细胞凋亡现象是毛细胞凋亡出现的最早表现;TUNEL阳性标记特征稍晚出现;PI标记可见毛细胞细胞核固缩和碎片出现的时间从2月龄开始;到3月龄时该种小鼠听力基本丧失,耳蜗毛细胞严重缺失;而应用z-VAD-FMK治疗后,各治疗组小鼠ABR反应阈明显好于对照组;耳蜗毛细胞凋亡数目较对照组减少,存活数明显较对照组多,尤以圆窗膜注射组明显。结论 NMF308nmf/nmf小鼠耳蜗毛细胞的死亡方式以caspase-3凋亡途径为主,应用caspase-3抑制剂zVAD-FMK定向内耳给药,可以阻断caspase-3信号途径激活所导致的毛细胞凋亡,从而有效防治老年性聋。
关键词
老年性聋;外毛细胞;凋亡;抗凋亡抑制剂;动物模型
基金项目(Foundation):
国家自然科学基金对外交流与合作项目(30210103151);国家自然科学基金项目(81170922);; 美国NIH项目(R01DC007392)联合资助
作者
李胜利;王玉虎;张敏燕;李白芽;郑庆印;朱雯瑾;朱宏亮;
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