Wang M, Wu F S, Cui B, Liang W, Zeng Q, Ma K F
School of Public Health, Tianjin Medical University, Tianjin 300070, China Institute for Occupational Health, Tianjin Centers for Disease Control and Prevention, Tianjin 300011, China.
Institute of Environmental and Operational Medicine, Academy of Military Medical Sciences, Academy of Military Sciences, Tianjin 300050, China.
Zhonghua Lao Dong Wei Sheng Zhi Ye Bing Za Zhi. 2024 Apr 20;42(4):241-247. doi: 10.3760/cma.j.cn121094-20230512-00171.
To explore the mechanism of noise-induced hidden hearing loss by proteomics. In October 2022, 64 SPF male C57BL/6J mice were divided into control group and noise exposure group with 32 mice in each group according to random sampling method. The noise exposure group was exposed to 100 dB sound pressure level, 2000-16000 Hz broadband noise for 2 h, and the mouse hidden hearing loss model was established. Auditory brainstem response (ABR) was used to test the change of hearing threshold of mice on the 7th day after noise exposure, the damage of basal membrane hair cells was observed by immunofluorescence, and the differentially expressed proteins in the inner ear of mice in each group were identified and analyzed by 4D-Label-free quantitative proteomics, and verified by Western blotting. The results were statistically analyzed by ANOVA and test. On the 7th day after noise exposure, there was no significant difference in hearing threshold between the control group and the noise exposure group at click and 8000 Hz acoustic stimulation (>0.05) . The hearing threshold in the noise exposure group was significantly higher than that in the control group under 16000 Hz acoustic stimulation (<0.05) . Confocal immunofluorescence showed that the basal membrane hair cells of cochlear tissue in noise exposure group were arranged neatly, but the relative expression of C-terminal binding protein 2 antibody of presynaptic membrane in middle gyrus and basal gyrus was significantly lower than that in control group (<0.05) . GO enrichment analysis showed that the functions of differentially expressed proteins were mainly concentrated in membrane potential regulation, ligand-gated channel activity, and ligand-gated ion channel activity. KEGG pathway enrichment analysis showed that differentially expressed proteins were significantly enriched in phosphatidylinositol 3 kinase-protein kinase B (PI3K-Akt) signaling pathway, NOD-like receptor signaling pathway, calcium signaling pathway, etc. Western blotting showed that the expression of inositol 1, 4, 5-trisphosphate receptor 3 (Itpr3) was increased and the expression of solute carrier family 38 member 2 (Slc38a2) was decreased in the noise exposure group (<0.05) . Through proteomic analysis, screening and verification of the differential expression proteins Itpr3 and Slc38a2 in the constructed mouse noise-induced hidden hearing loss model, the glutaminergic synaptic related pathways represented by Itpr3 and Slc38a2 may be involved in the occurrence of hidden hearing loss.
通过蛋白质组学探讨噪声性隐匿性听力损失的机制。2022年10月，将64只SPF级雄性C57BL/6J小鼠按随机抽样法分为对照组和噪声暴露组，每组32只。噪声暴露组给予100 dB声压级、2000 - 16000 Hz宽带噪声暴露2 h，建立小鼠隐匿性听力损失模型。采用听性脑干反应（ABR）检测噪声暴露后第7天小鼠听力阈值变化，通过免疫荧光观察基底膜毛细胞损伤情况，运用4D-Label-free定量蛋白质组学技术鉴定并分析各组小鼠内耳差异表达蛋白，并用蛋白质免疫印迹法进行验证。结果采用方差分析和检验进行统计学分析。噪声暴露后第7天，在短声和8000 Hz声刺激下，对照组与噪声暴露组听力阈值差异无统计学意义（>0.05）。在16000 Hz声刺激下，噪声暴露组听力阈值显著高于对照组（<0.05）。共聚焦免疫荧光显示，噪声暴露组耳蜗组织基底膜毛细胞排列整齐，但中回和基回突触前膜C末端结合蛋白2抗体相对表达量显著低于对照组（<0.05）。基因本体（GO）富集分析显示，差异表达蛋白的功能主要集中在膜电位调节、配体门控通道活性和配体门控离子通道活性。京都基因与基因组百科全书（KEGG）通路富集分析显示，差异表达蛋白显著富集于磷脂酰肌醇3激酶-蛋白激酶B（PI3K-Akt）信号通路、NOD样受体信号通路、钙信号通路等。蛋白质免疫印迹法显示，噪声暴露组肌醇1,4,5-三磷酸受体3（Itpr3）表达增加，溶质载体家族38成员2（Slc38a2）表达降低（<0.05）。通过蛋白质组学分析，筛选并验证构建的小鼠噪声性隐匿性听力损失模型中差异表达蛋白Itpr3和Slc38a2，以Itpr3和Slc38a2为代表的谷氨酸能突触相关通路可能参与隐匿性听力损失的发生。
