Caspase12在强噪声诱导下豚鼠耳蜗细胞凋亡中的作用

Role of caspase 12 in apoptosis of cochlea induced by intense noise in guinea pigs

目的探讨caspase12在强噪声诱导的豚鼠耳蜗细胞凋亡中的作用。方法选用健康雄性白色豚鼠32只，随机数字表法分为4组，将实验组动物暴露于声压级120dB、4kHz窄带噪声环境4h。分别对健康对照组及噪声刺激后第1、4、14天组豚鼠测试听性脑干反应（ABR）。每组取4只豚鼠耳蜗作石蜡切片，另外4只豚鼠提取耳蜗总蛋白。通过透射电镜和脱氧核糖核甘酸末端转移酶介导的缺口末端标记技术（ternial-deoxynucleotidyl transferase mediated nick end labeing，TUNEL）检测耳蜗凋亡细胞，免疫组化及蛋白质印迹（Western blot）法检测Bio／GRP78、caspase12蛋白的表达。结果透射电镜观察到实验组第1天耳蜗外毛细胞、螺旋神经节细胞出现凋亡的特征性改变。实验组耳蜗Corti器、侧壁的血管纹和螺旋神经节存在TUNEL阳性细胞，第1天组最多；对照组未见阳性细胞。免疫组化、Western blot检测实验组Bjp／GRP78蛋白明显高于对照组，且在第1、4、14天都维持在比较高的水平；caspase12蛋白含量在噪声刺激后迅速增高，第1、4天达到高峰，第14天表达明显下降，但仍维持较高水平。结论强噪声可以诱导豚鼠耳蜗细胞凋亡，caspase12活化引起内质网应激反应性凋亡通道可能是此过程的信号途径之一。

Objective To investigate the relationship between caspase 12 activation and endoplasmic reticulum stress mediated apoptosis of guinea pig cochlea cells induced by intense noise. Methods Thirty-two guinea pigs were randomly divided into 4 groups. The guinea pigs in the experiment groups were exposed to 4 kHz narrow band noise at 120 dB SPL for 4 h. After the noise expose for 1,4, 14 days of the experiment guinea pigs, auditory brainstem response （ABR） of the guinea pigs on experiment and control groups were tested before decapitated. Four guinea pig＇s cochleae of every group were taken to paraffin section, and the rest was extracted the total protein. Apoptosis was tested by terminal deoxynucleotidyl transferase（TDT）-mediated deoxyuridine triphosphate （d-UTP） nick and labeling method （TUNEL） and transmission electron microscopy. Expression of caspase 12, Bip/GRP78 was tested by immunohistochemistry and Western blot methods. Results The observation by transmission electron microscopy showed the features characteristic of apoptotic cells in the Corti and SGC of ld after the noise expose, but no in the control. There were higher expressions of Tunel-Positve cells in the OHC, SGC and SV of experiment groups, and there was significant differences compared with the control group （ P 〈 0.01 ）. Protein levels of Bip/GRP78 and caspase 12 were risen up after noise exposed, and there all were significant differences compared with the control group （P 〈 0.01 ）. Conclusion Intense noise causes cochlea cell lesion by inducing apoptosis to result in and caspase 12 induced endoplasmic reticulum stress-related apoptosis plays an important role in the procedure of apoptosis.