低频强声暴露后巴马香猪耳蜗caspase-3表达及毛细胞死亡观察

The Observation of Cochlea Hair Cell Death and Expression of Caspase-3 after Intense Low Frequency Noise Exposure in Bama Pig

目的了解高强度低频噪声(下称:低频强声)暴露对巴马香猪耳蜗凋亡因子半胱氨酸天冬氨酸蛋白酶-3(caspase-3)表达的影响,同时观察耳蜗毛细胞死亡情况。方法将8只巴马香猪随机分为对照组2只及实验组6只;所有动物均于实验前行听性脑干反应(ABR)检测,然后将实验组随机分为噪声暴露后即刻组、36h组及84h组,每组各2只,分别暴露于50Hz、142dB SPL的噪声中5min,于噪声暴露后相应时间点行ABR检测,用免疫组织化学染色和免疫荧光DAPI标记细胞核两种方法观察耳蜗组织中caspase-3的表达,同时通过细胞计数方法计算毛细胞缺失率判定细胞死亡情况。对照组不给予噪声暴露,其他条件与实验组相同。结果噪声暴露前8只巴马香猪的ABR平均阈值为61.25±10.72dB nHL;低频强声暴露后实验组中的即刻组、36h组和84h组的动物双耳ABR均未引出;实验各组动物耳蜗各部位均可见caspase-3阳性表达,且随着暴露后时间延长,caspase-3阳性表达逐渐减弱;噪声暴露后即刻组、36h组与正常对照组相比较,耳蜗三排外毛细胞排列及层次明显错乱;噪声暴露后84h组耳蜗三排外毛细胞消失,仅可见一层内毛细胞;正常对照组及噪声暴露后即刻组、36h组、48h组耳蜗外毛细胞缺失率分别为0%、0%、14.06%、100%。结论低频强声暴露使巴马香猪听性脑干反应阈值较暴露前明显提高,随暴露后时间延长,耳蜗组织中caspase-3阳性表达减弱,毛细胞死亡数量明显增加,螺旋神经节区域也出现了细胞凋亡现象。

Objective To investigate the effects of intense low frequency noise on expression of caspase 3 in bama pigs, and observe the death of cochlea hair cell. Methods 8 bama pigs were randomly divided into a normal control group（2 pigs） and experimental groups（6 pigs）. Auditory brainstem responses（ABR） were tested before the experiment. The control group was the same as the experimental groups except noise exposure, The experimental group was randomly divided into immediate group,36 h group, and 84 h group（2 pigs per group）. They were ex posed to intense low frequency noise at 142 dB of 50 Hz for 5 min according to the three time points. ABR were test ed again before the cochlea were collected at different time points. The expression of caspase--3 was studied through immunohistochemistry and immunofluorescence, hair cell loss rate by counting to determine cell death. Results The average ABR threshold of 8 bama pigs（16 ears） before the exposure was 61.25±10.72 dB nHL. ABR thresh-olds were not elicited after exposure to the noise. Different parts of bama pigs showed positive expression of caspase-3 in two ways. As time prolonged after exposure, the positive expression of easpase-3 gradually weakened. The immediate group and 36 h group, compared with the control group, showed the apparent misplace of three out hair ceils in the arrangement and levels. The 84 h group through immunofluorescence lost out hair cells, and only inner hair ceils were visible. Conclusion ABR thresholds were elevated after noise exposure. The procedure of hair ceil nucleus damage and caspase--3 expression is different, and the noise can induce opening apoptotic program of spiral ganglion.