ANALYSIS OF GENES AND PROTEINS RELATED TO COCHLEAR DAMAGE USING DNA MICROARRAY AND PROTEOMICS

我们产生了PolgD 257 A/D257 A（D257 A）突变小鼠（在DNA聚合酶γ的外切核酸酶结构域（Proof阅读活性结构域）中引入点突变的转基因小鼠）作为老年性耳聋动物模型。D257 A小鼠在9月龄时表现出平均41-52 dBSPL的ABR阈值偏移，而在该年龄的野生型小鼠中没有观察到ABR阈值偏移，这表明D257 A小鼠的听力损失是由于早期衰老而出现的。组织学上，该突变小鼠表现出耳蜗毛细胞和螺旋神经节细胞的严重变性。对耳蜗的DNA微阵列分析显示，与声音感知、神经传递、离子代谢、能量代谢、DNA合成和DNA修复相关的基因减少，与炎症和凋亡相关的基因增加。我们还通过给CBA小鼠喂饲二氧化锗建立了另一种线粒体功能障碍的动物模型。锗铁开始后3个月内 ...更多信息 在D257 A小鼠中，该小鼠发生了严重的听力损失和血管纹和Corti器官的严重变性，DNA微阵列分析揭示了与在D257 A小鼠中观察到的那些相似的基因表达变化。在另一个实验中，我们研究了热量限制（CR）是否可以预防老年性耳聋模型小鼠C57 BL/6小鼠的老年性耳聋。未接受CR的C57 BL/6小鼠在17月龄时出现听力损失和耳蜗组织变性，CR防止听力损失和耳蜗变性的表现直到该年龄。C57 BL/6小鼠耳蜗的DNA微阵列分析显示，CR防止了与年龄相关的基因表达变化，如声音感知，神经传递和能量代谢相关基因的减少，以及炎症和凋亡相关基因的增加。我们通过真实的实时RT-PCR分析证实了几个重要基因的表达变化，并通过免疫组化证实了其相关蛋白的表达变化。最后，我们对暴露于强烈声音的小鼠进行了DNA微阵列分析，目前正在对几个被认为是噪音引起的听力损失表现的关键基因进行详细分析。少

We generated a PolgD257A/D257A (D257A) mutant mouse (a transgenic mouse in which point mutations were introduced into exonuclease domain (Proof Reading activity domain) of DNA polymerase gamma) as a presbycusis animal model. The D257A mouse exhibited ABR threshold shifts of an average of 41-52 dBSPL at the age of 9 month, whereas no ABR threshold shifts were observed in a wild type mouse at this age, indicating that hearing loss appeared by aging early in D257A mouse. Histologically, this mutant mouse exhibited severe degeneration of the cochlear hair cells and spiral ganglion cells. DNA microarray analysis of their cochleae showed a decrease in genes related to sound perception, neurotransmission, ion metabolism, energy metabolism, DNA synthesis, and DNA repair and an increase in genes related to inflammation and apoptosis. We also generated another animal model showing mitochondria dysfunction by feeding CBA mouse with germanium dioxide. Within 3 months from starting the germanium fe … More eding, this mouse developed profound hearing loss and severe degeneration of the stria vascularis and an organ of Corti, and DNA microarray analysis revealed gene expression changes similar to those observed in D257A mouse. In another experiment, we examined whether caloric restriction (CR) could prevent presbycusis in C57BL/6 mouse, a presbycusis model mouse. The C57BL/6 mouse not subjected to CR developed hearing loss and degeneration of the cochlear tissues by the age of 17 months, and CR prevented the manifestation of hearing loss and cochlear degeneration until this age. DNA microarray analysis of the cochlea in C57BL/6 mouse both subjected and not subjected to CR revealed that CR prevented age-related changes of gene expression, such as a decrease of genes related to sound perception, neurotransmission, and energy metabolism, and an increase of genes related to inflammation and apoptosis. We confirmed the expression changes of several important genes by means of real time RT-PCR analysis and their related proteins by performing immunohistochemistry. Finally, we did DNA microarray analysis for mouse exposed to intense sound and are currently doing detailed analysis for several genes that are considered keys in the manifestation of noise0-induced hearing loss. Less