噪声对大鼠耳蜗基底膜AIF表达的影响

发布时间：2018-11-18 22:00

【摘要】：实验一建立噪声性听力损伤动物模型 1目的通过建立噪声性听力损伤动物模型，观察噪声对大鼠听性脑干反应（auditorybrainstem response, ABR）阈值及耳蜗基底膜不同频率区域外毛细胞形态学的影响，探讨其对听觉系统的损伤作用。 2方法 40只SD大鼠随机分为正常对照组和噪声暴露组：噪声暴露组给予声强为115dBSPL白噪声暴露，每天2小时，连续3天，对照组不予噪声暴露。分别于噪声暴露前1日、暴露后1、3、7、14日对两组大鼠行ABR检测，并用在最后一次ABR检测后对两组大鼠耳蜗基底膜行鬼笔环肽—异硫氰酸荧光素(Phalloidin-FITC)染色。 3结果大鼠噪声暴露后与暴露前相比，ABR各频率反应阈值于暴露后1天达到最高，并随时间的推移逐渐恢复，14天时趋于稳定，听力低频阈移约10dB，高频阈移有30dB (P0.05)；基底膜铺片FITC染色示噪声暴露组底回基底膜毛细胞较顶回缺失严重，且有纤毛排列紊乱并出现融合，而对照组毛细胞排列整齐，纤毛呈V或W型，两组间外毛细胞计数比较差异有统计学意义(P0.05)。 4结论在噪声性听力损伤过程中，早期的听力损失存在暂时性阈移，在14天左右才趋于稳定，出现永久性阈移，ABR结果提示氧化还原反应亦在噪声暴露后1天时最为激烈，且耳蜗基底膜高频区域受到的氧化损伤较低频区域更为严重，不易恢复。实验二噪声对大鼠耳蜗不同频率区域基底膜AIF表达的影响 1目的探讨噪声暴露前后凋亡诱导因子（AIF）在大鼠不同回基底膜外毛细胞的表达差异以及与噪声性聋高频听力易损性的关系。 2方法 40只SD大鼠随机分为正常对照组和噪声暴露组：噪声暴露组给予声强为115dBSPL白噪声暴露，每天2小时，连续3天，对照组不予噪声暴露，其余条件与噪声暴露组相同。并于噪声暴露完成后次日，行耳蜗基底膜取材及铺片制备，分别采用RT-qPCR、Western blot和免疫荧光染色方法，从定性及定量角度观察两组大鼠耳蜗不同回基底膜处AIF的表达。 3结果免疫荧光染色显示，正常组中AIF在顶、底回基底膜外毛细胞胞浆中均有表达，细胞核中无表达，噪声暴露组AIF在顶、底回基底膜外毛细胞的荧光强度均较正常对照组强，且在细胞核中可见到少量AIF表达；RT-qPCR与Western blot结果相符，显示为，在正常情况下,顶回基底膜的AIF表达高于底回，噪声暴露后，AIF顶、底回基底膜表达均较正常对照组相应部位增高，且顶回较底回更为显著(P0.05)。 4结论正常情况下，AIF在耳蜗不同回基底膜外毛细胞中的表达存在明显差异，而噪声暴露后，AIF在耳蜗不同频率区域基底膜外毛细胞中的表达显著增高，并且在此过程中主要发挥了氧化还原酶的作用，凋亡诱导活性占次要地位，，说明耳蜗不同频率区域基底膜抵御氧化还原反应的能力存在明显差异，这可能是噪声性聋高频听力易损性的分子机制之一。
[Abstract]:Experiment 1: establish an animal model of noise-induced hearing impairment objective to observe the effects of noise on auditory brainstem response (auditorybrainstem response,) in rats by establishing an animal model of noise-induced hearing impairment. ABR) threshold and the morphological changes of outer hair cells in different frequency regions of the cochlear basement membrane. Methods 40 SD rats were randomly divided into normal control group and noise exposure group. The noise exposure group was exposed to 115dBSPL white noise for 2 hours a day for 3 days. The control group was not exposed to noise. ABR was detected on the 1st day before noise exposure, 1 day after exposure and 14 days after exposure, and the cochlea basement membrane of the two groups was stained with Phalloidin-FITC after the last ABR detection. Results compared with before and after noise exposure, the threshold of ABR frequency response reached the highest level on the first day after exposure, and gradually recovered with the passage of time, and tended to stabilize at 14 days, and the low frequency threshold of hearing shifted about 10 dB. High frequency threshold shifted to 30dB (P0.05); FITC staining showed that the basal membrane hair cells in the noise exposed group were more serious than those in the apical gyrus, and the cilia were disordered and fused, while in the control group, the hair cells were arranged neatly, and the cilia were V or W type. There was significant difference in the number of outer hair cells between the two groups (P0.05). Conclusion in the process of noise-induced hearing loss, there is a temporary threshold shift in the early stage of hearing loss, which tends to stabilize at about 14 days, and a permanent threshold shift occurs. The results of ABR showed that the redox reaction was the most intense one day after noise exposure and the oxidation damage in the high frequency region of the basal membrane of cochlea was more serious than that in the low frequency region and it was not easy to recover. Effects of noise on the expression of AIF in the basement membrane of rat cochlea at different frequencies 1 objective to investigate the expression of apoptosis-inducing factor (AIF) in rat hair cells with different return basal membranes before and after noise exposure. The relationship between high frequency hearing loss and noise deafness. Methods 40 SD rats were randomly divided into normal control group and noise exposure group. The noise exposure group was exposed to 115dBSPL white noise for 2 hours a day for 3 days. The control group was not exposed to noise, and the other conditions were the same as those in the noise exposure group. The next day after noise exposure, the basal membrane of cochlea was taken and prepared. The expression of AIF in different basal membrane of cochlea was observed qualitatively and quantitatively by RT-qPCR,Western blot and immunofluorescence staining. Results Immunofluorescence staining showed that AIF was expressed in the cytoplasm of basal membrane hair cells in the top and bottom gyrus, but not in the nucleus in the normal group, and AIF was at the top in the noise exposure group. The fluorescence intensity of basal membrane hair cells in the basal gyrus was stronger than that in the control group, and a small amount of AIF expression could be found in the nucleus. The results of RT-qPCR and Western blot showed that the expression of AIF in basal membrane of parietal gyrus was higher than that in basal gyrus under normal condition. After noise exposure, the expression of AIF in basal membrane of AIF top and bottom gyrus was higher than that in control group. The parietal gyrus was more significant than the bottom gyrus (P0.05). Conclusion under normal conditions, the expression of AIF in different basal membrane hair cells of cochlea is significantly different, but after noise exposure, the expression of AIF in different frequency regions of cochlea is significantly increased. In this process, the redox enzyme was mainly played, and the apoptosis-inducing activity was secondary, which indicated that the ability of the basement membrane to resist the redox reaction was significantly different in different regions of cochlea. This may be one of the molecular mechanisms of high frequency hearing vulnerability in noise deafness.
【学位授予单位】：第四军医大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R764.433

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