College of Otolaryngology Head and Neck Surgery, Chinese PLA General Hospital, Beijing 100853, China.
National Clinical Research Center for Otolaryngologic Diseases, Beijing, China.
Neural Plast. 2020 Jun 12;2020:6235948. doi: 10.1155/2020/6235948. eCollection 2020.
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-B (p65) in the hair cells of miniature pig inner ears during the acute hearing loss stage after noise exposure. Nuclear translocation of NF-B (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.
本研究旨在通过小型猪模型探索急性噪声性听力损失和稳态噪声性听力损失恢复的分子机制。我们使用暴露于 120dB(A) 白噪声的小型猪作为模型。在噪声暴露前、噪声暴露后 1 天和 7 天进行听觉脑干反应（ABR）测量。采用同位素相对标记与绝对定量（iTRAQ）蛋白质组学方法观察噪声暴露后小型猪内耳蛋白质的变化。通过 Western blot 和免疫荧光进一步定量和定性分析蛋白质组变化。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 39.4dB SPL、67.1dB SPL 和 50.8dB SPL。使用 iTRAQ 共鉴定出 2158 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，免疫和代谢途径在急性听力损失损伤阶段明显参与。在急性听力损失恢复阶段，大多数差异表达蛋白与胆固醇代谢有关。Western blot 和免疫荧光显示，噪声暴露后急性听力损失阶段小型猪内耳毛细胞中活性氧的积累和 NF-B（p65）的核转位。NF-B（p65）的核转位可能与下游炎症因子的过度表达有关。载脂蛋白（Apo）A1 和 Apo E 在听力损失恢复阶段显著上调，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方法在大型动物模型（小型猪）中分析急性噪声性听力损失及其恢复的分子机制。我们的研究结果表明，代谢、炎症和固有免疫途径的激活可能与急性噪声性听力损失有关，而胆固醇代谢途径可能在噪声性听力损失后的听力恢复中发挥重要作用。
