An increase in mitochondrial damage has been associated with a decline in the ability to mitigate damage through mitophagy in age-related pathologies. The present study aimed to investigate the changes of mitophagy in a mouse model with age-related hearing loss. C57BL/6J mice were divided into two groups: young (1 month) and aged (12 months). Hearing tests were conducted through the measurement of auditory brainstem response (ABR). Mitochondrial DNA copy number, the level of mitochondrial DNA damage, mitochondrial biogenesis, and mitophagy-related genes and proteins were investigated using real-time PCR and western blot analysis. Coexpression of mitophagosomes and lysosomes in the cochlea was investigated through immunofluorescence imaging analysis. Major players of mitophagy, Parkin and BNIP3, were also investigated through immunohistochemical staining in the cochlea. Hearing thresholds were observed to have increased in the aged group. The mitochondrial DNA copy number, PGC-1α, and PGC-1β significantly decreased in the cochlea of mice in the aged group. The mRNA levels of PINK1, Parkin, MUL1, Atg5, Atg12, Atg13, NIX, and BNIP3 significantly decreased in the cochlea of the mice in the aged group. The level of mitochondrial DNA damage significantly increased in the cochlea of mice in the aged group. Protein levels of PINK1, Parkin, BNIP3, COX4, LC3B, and all OXPHOS subunits significantly decreased in the cochlea of the mice in the aged group. Immunofluorescence imaging analysis of mitophagosomes and lysosomes revealed a decrease in the colocalization in the cochlea of mice in the aged group. Immunohistochemical imaging analysis of Parkin and BNIP3 revealed their decreased expression in aged cochlea. Our results indicate that reduced mitophagy with aging might be attributed to the cellular changes that occur in aged cochlea in the development of age-related hearing loss.

中文翻译：

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降低了老年C57BL / 6J小鼠耳蜗的线粒体吞噬能力。

线粒体损伤的增加与年龄相关疾病中通过线粒体减轻损伤的能力下降有关。本研究旨在调查与年龄相关的听力损失的小鼠模型中线粒体的变化。C57BL / 6J小鼠分为两组：年轻（1个月）和年龄（12个月）。通过测量听觉脑干反应（ABR）进行听力测试。使用实时荧光定量PCR和Western blot分析了线粒体DNA的拷贝数，线粒体DNA损伤的水平，线粒体的生物发生以及与线粒体相关的基因和蛋白质。通过免疫荧光成像分析研究了耳蜗中的线粒体和溶酶体的共表达。线粒体，Parkin和BNIP3的主要参与者，还通过耳蜗中的免疫组织化学染色对它们进行了研究。观察到老年人的听力阈值增加。老年小鼠的耳蜗中线粒体DNA拷贝数，PGC-1α和PGC-1β明显降低。在老年组小鼠的耳蜗中，PINK1，Parkin，MUL1，Atg5，Atg12，Atg13，NIX和BNIP3的mRNA水平显着降低。老年组小鼠的耳蜗中线粒体DNA损伤水平显着增加。老年组小鼠的耳蜗中PINK1，Parkin，BNIP3，COX4，LC3B和所有OXPHOS亚基的蛋白水平显着降低。线粒体和溶酶体的免疫荧光成像分析显示，老年组小鼠耳蜗的共定位降低。Parkin和BNIP3的免疫组织化学成像分析显示，它们在老年耳蜗中表达降低。我们的结果表明，随着年龄的增长，线粒体细胞减少可能归因于与年龄有关的听力损失的发展，老年耳蜗中发生了细胞变化。