The objective of this study was to explore the molecular mechanisms of acute noise-induced hearing loss and recovery of steady-state noise-induced hearing loss using miniature pigs. We used miniature pigs exposed to white noise at 120 dB (A) as a model. Auditory brainstem response (ABR) measurements were made before noise exposure, 1 day and 7 days after noise exposure. Proteomic Isobaric Tags for Relative and Absolute Quantification (iTRAQ) was used to observe changes in proteins of the miniature pig inner ear following noise exposure. Western blot and immunofluorescence were performed for further quantitative and qualitative analysis of proteomic changes. The average ABR-click threshold of miniature pigs before noise exposure, 1 day and 7 days after noise exposure, were 39.4 dB SPL, 67.1 dB SPL, and 50.8 dB SPL, respectively. In total, 2,158 proteins were identified using iTRAQ. Both gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) database analyses showed that immune and metabolic pathways were prominently involved during the impairment stage of acute hearing loss. During the recovery stage of acute hearing loss, most differentially expressed proteins were related to cholesterol metabolism. Western blot and immunofluorescence showed accumulation of reactive oxygen species and nuclear translocation of NF-κB (p65) in the hair cells of miniature pig inner ears during the acute hearing loss stage after noise exposure. Nuclear translocation of NF-κB (p65) may be associated with overexpression of downstream inflammatory factors. Apolipoprotein (Apo) A1 and Apo E were significantly upregulated during the recovery stage of hearing loss and may be related to activation of cholesterol metabolic pathways. This is the first study to use proteomics analysis to analyze the molecular mechanisms of acute noise-induced hearing loss and its recovery in a large animal model (miniature pigs). Our results showed that activation of metabolic, inflammatory, and innate immunity pathways may be involved in acute noise-induced hearing loss, while cholesterol metabolic pathways may play an important role in recovery of hearing ability following noise-induced hearing loss.

中文翻译：

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胆固醇代谢途径参与噪音诱发的听力损失的恢复。

这项研究的目的是探讨使用小型猪的急性噪声诱发的听力损失和稳态噪声诱发的听力损失恢复的分子机制。我们使用暴露在120 dB（A）白噪声下的小型猪作为模型。噪声暴露前，噪声暴露后1天和7天进行听觉脑干反应（ABR）测量。相对定量和绝对定量的蛋白质组同量异位标签（iTRAQ）用于观察噪声暴露后小型猪内耳蛋白质的变化。进行蛋白质印迹和免疫荧光分析以进一步定量和定性分析蛋白质组学变化。小型猪在噪声暴露前，噪声暴露后1天和7天的平均ABR点击阈值分别为39.4 dB SPL，67.1 dB SPL和50.8 dB SPL。总共2 使用iTRAQ鉴定出158种蛋白质。基因本体论和《京都基因与基因组百科全书》（KEGG）数据库分析均显示，急性听力损失的损害阶段主要涉及免疫和代谢途径。在急性听力损失的恢复阶段，大多数差异表达的蛋白质与胆固醇代谢有关。Western印迹和免疫荧光显示活性氧的积累和NF-暴露于噪音后的急性听力丧失阶段，小型猪内耳毛细胞中的κB（p65）。NF- κ的核易位B（p65）可能与下游炎症因子的过度表达有关。在听力丧失的恢复阶段，载脂蛋白（Apo）A1和Apo E显着上调，可能与胆固醇代谢途径的激活有关。这是首次使用蛋白质组学分析来分析大型动物模型（小型猪）中急性噪声引起的听力损失及其恢复的分子机制。我们的结果表明，代谢，炎症和先天免疫途径的激活可能与急性噪声诱发的听力损失有关，而胆固醇代谢途径可能在噪声诱发的听力损失后恢复听力中发挥重要作用。