Apoptotic and Necrotic Death of OHCs in NIHL

描述（由申请人提供）：我们之前的研究发现，在高水平的噪音暴露后，外毛细胞（ohc）以凋亡和坏死的方式死亡。细胞凋亡或坏死的发生率与噪声水平和暴露后耳蜗病变的进展有关；然而，决定噪音暴露后OHC死亡途径的细胞机制尚不清楚。该应用主要关注于OHCs因凋亡或坏死而死亡的倾向是由死亡OHCs的能量水平和/或氧化状态调节的这一一般假设。为了验证这一假设，将龙猫暴露在以110 dB SPL为中心的4 kHz倍频噪声中1小时。在实验的第一部分，我们将检测噪声暴露后凋亡和坏死OHCs中琥珀酸脱氢酶（SDH）的活性，这是一种参与电子传递链中ATP合成的重要线粒体酶。在本研究的第二部分，ohc合成ATP将会被一种不可逆的SDH抑制剂3-硝基丙酸（3-NP）在耳蜗内阻断。我们将研究噪声暴露后细胞内ATP耗竭对细胞凋亡和坏死的影响。在研究的最后一部分，耳蜗抗氧化能力的改变对死亡途径的影响将被检查。耳蜗的抗氧化能力可以通过使用谷胱甘肽合成抑制剂l -丁硫氨酸-[S， R]-亚砜胺或谷胱甘肽单乙基酯（谷胱甘肽的类似物）改变耳蜗谷胱甘肽（GSH）的水平来控制。这些数据将有助于阐明耳蜗能量水平和抗氧化水平在噪声暴露后决定细胞凋亡或坏死发生率中的作用。我们的长期目标是探索有效的治疗策略，以减少噪音引起的听力损失。由于坏死的OHCs可能通过释放细胞内内容物对存活的OHCs产生更大程度的毒性应激，因此防止细胞凋亡转化为坏死可能改善整体耳蜗损伤。对调节细胞死亡途径的生物学机制的了解将为最终制定合理的保护策略提供基础。

DESCRIPTION (provided by applicant): Our previous studies have found that following a high level of noise exposure, outer hair cells (OHCs) die by both apoptosis and necrosis. The prevalence of apoptosis or necrosis is associated with the noise level and post-exposure progression of the cochlear lesion; however, the cellular mechanisms responsible for determining the pathways of OHC death following noise exposure are still not known. This application focuses on the general hypothesis that the propensity of OHCs to die by apoptosis or necrosis is regulated by the energy level and/or oxidative status of dying OHCs. To test this hypothesis, chinchillas will be exposed to an octave band noise centered at 4 kHz at 110 dB SPL for 1 hour. In the first part of the experiment, the activity of succinate dehydrogenase (SDH), an important mitochondrial enzyme that participates in ATP synthesis in the electron transport chain, will be examined in apoptotic and necrotic OHCs after the noise exposure. In the second part of the study, the synthesis of ATP by OHCs will be blocked by intracochlear application of 3-nitropropionic acid (3-NP), an irreversible inhibitor of SDH. The effect of intracellular ATP depletion on generation of apoptosis and necrosis following the noise exposure will be examined. In the last part of the study, the effect of the alteration of the cochlear antioxidant capacity on the death pathways will be examined. The cochlear antioxidant capacity will be manipulated by changing the level of cochlear glutathione (GSH) either with L-buthionine-[S, R]-sulfoximine, a GSH synthesis inhibitor, or with glutathione monoethyl ester, an analog of GSH. These data will help to elucidate the role of the cochlear energy level and the antioxidant level in determining the prevalence of either apoptosis or necrosis after noise exposure. Our long-term goal is to explore effective therapeutic strategies to reduce noise-induced hearing loss. Since necrotic OHCs potentially create a significantly greater level of toxic stress on surviving OHCs through release of intracellular contents, preventing the conversion of apoptosis to necrosis may ameliorate the overall cochlear damage. The knowledge of the biological mechanisms responsible for modulation of the cell death pathways will provide the basis for eventually developing a rational protective strategy.