老年性耳聋发病机制及药物筛查研究

Presbycusis or age-related hearing loss (AHL) affects over 40% of the aging population. Many factors can cause AHL, including ototoxicity drugs and noise exposure. Due to increases in the aging population of our country, AHL significantly affects quality of life. The mechanism of AHL is poorly understood; there is no effective treatment. Study of the human temporal bone may provide limited information, but can only be performed in post-mortem cases. For this reason, we have developed a deaf (erlong, erl) mouse model. Our preliminary study has demonstrated that erl mice start to lose hearing at one month, and by three months of age become completely deaf. This short time window is ideal to test drug effects on hearing protection. We have identified erl as a new allele of the Cdh23 gene with a 208?T>C transition causing an amino-acid substitution (70S-P). Caspase expression was upregulated in mutant inner ears. Hearing was preserved (up to 35-dB improvement) in pan-caspase inhibitor Z-VAD-FMK-treated mutants compared with untreated mutants (P<0.05). Outer hair cell (OHC) loss in the cochleae of Z-VAD-FMK-treated mutants was significantly reduced compared with untreated mice. This establishes the basis for testing more hearing protection drugs as proposed in this application.

老年性聋（AHL）严重影响大量老年人的生活质量。目前 AHL的发生机理尚不明确，且无有效药物。研究人类颞骨能提供有用信息，但只能在死后尸检才能做到。为此，我们培育了耳聋小鼠模型，前期研究结果表明：其特征是出生后1个月出现听力减退，随年龄增加逐渐加重，至3个月听力近全聋。期间毛细胞缺失从耳蜗基底部向顶部蔓延。遗传测序表明 erl 是 Cdh23 基因的一个新的等位基因，突变发生在(208T＞C）, 引起 CDH23 蛋白中氨基酸的替换(70S－P)。与对照鼠比较，耳聋小鼠内耳的凋亡信号Caspases显著升高；给予caspase 抑制剂后，小鼠听阈值得到近30 dB的保护。钙信号是Caspases的上游信号，在前期研究的基础上，我们提出假说：阻止钙离子通道，可有效改善耳聋小鼠听力。本课题拟观察经腹腔给耳聋小鼠两个阻止钙离子通道的药物观察听力保护作用，探讨其机制，为AHL的防治提供新途径。

钙粘蛋白23（Cdh23）作为内耳毛细胞尖端连接的组成部分与年龄相关的听力损失相关。本项目研究发现了一种新的小鼠 Cdh23 基因突变形式（erl 突变）， 即Cdh23基因 3 号外显子的单个点突变（208T>C）。作为一种新的 DFNB12 动物模型，纯合型 Cdh23erl/erl小鼠是年龄相关性听力损失的新型动物模型，呈现渐进性耳聋的表型特点，本课题对erl 小鼠表型进行了全面研究，采用多种听功能检测方法对小鼠听力损失程度进行评估，利用多种形态学方法对内耳毛细胞、螺旋神经节细胞进行观察。利用免疫荧光等技术对erl 小鼠听力损失发生机制进行研究，证明内质网应激参与erl小鼠模型内耳毛细胞凋亡从而导致听力损失的假说，阐明内质网应激在毛细胞凋亡通路中的发生机制。另外利用erl小鼠和另一渐进性耳聋小鼠模型 NOD/LtJ- Cdh23ahl/ahl小鼠，完成了多种听力保护药物的筛选工作，筛选药物包括乙琥胺（ethosuximide）、促红细胞生成素（EPO）、牛磺熊去氧胆酸（Tauroursodeoxycholic acid，TUDCA）等。评价不同药物的听力保护作用效果，阐明分子作用机制，并证明Cdh23erl/erl是试验听力保护药物的理想模型，为临床治疗老年性耳聋提供了新的研究方向。共发表SCI 文章6 篇。