听力阈值变化与卡那霉素耳慢性中毒豚鼠耳蜗细胞凋亡

Apoptosis of cochlear cells and hearing threshold in guinea pigs with kanamycin-induced chronic intoxication

目的:探讨卡那霉素耳慢性中毒后豚鼠耳蜗细胞凋亡现象及其在听力损伤中的作用。方法:实验于2003-01/12在第三军医大学大坪医院野战外科研究所完成。75只豚鼠随机分为正常对照组(n=15),卡那霉素耳慢性中毒组(n=60)。卡那霉素耳慢性中毒组进行硫酸卡那霉素200mg/(kg·d)连续肌肉注射14d,根据停药时间分为卡那霉素停药1,3,7,14d组,每组15只。分别处死正常对照组及卡那霉素停药1,3,7,14d组豚鼠,处死前检测其听脑干反应的变化,处死后以原位末端标记技术及耳蜗毛细胞核荧光染色计数等检测耳蜗细胞凋亡情况。结果:①豚鼠连续应用卡那霉素14d,停药1d耳蜗细胞即出现凋亡现象(原位缺口末端标记法阳性细胞数:柯蒂氏器0.69±0.57,螺旋神经节1.22±0.51,血管纹1.01±0.67),随停药时间延长,耳蜗细胞凋亡数增多(14d组原位缺口末端标记法阳性细胞数:柯蒂氏器4.91±0.94,螺旋神经节8.84±1.35,血管纹7.39±1.63),且愈近底回愈多,愈近顶回则愈少,对照组无凋亡现象。②豚鼠卡那霉素耳慢性中毒后,随停药时间延长,听脑干反应阈值14d组为43.20±6.73,正常对照组为5.60±1.78,14d组较正常对照组明显上升(P<0.01)。结论:①卡那霉素耳慢性中毒可引起耳蜗细胞发生凋亡,细胞凋亡是豚鼠耳蜗损伤的一条途径。

AIM:To investigate the apoptosis of cochlear cells induced by chronically ototoxic kanamycin and the effect of cochlear cell apoptosis on hearing loss in guinea pigs.METHODS:The experiment was completed in the Research Institute of Surgery,Daping Hospital,Third Military Medical University from January to December 2003.Seventy five guinea pigs were randomly divided into normal control group(n=15) and chronically ototoxic kanamycin treatment group.The guinea pigs in the latter group were intramuscularly injected with kanamycin 200 mg/kg per day for 14 consecutive days,then they were equally divided into 1 day,3 day,7 day and 14 day withdrawal groups according to their withdrawal time.Before scarification to the guinea pigs,the alteration of auditory brainstem response(ABR) was detected in all the groups.After scarification,the apoptosis of cochlear cells was determined by terminal deoxynucleotidyl transferase mediated dUTP nick and labeling(TUNEL) technology and fluorescence staining of cochlear hair cells at different time.RESULTS:① Apoptosis of cochlear cells was noted in 1 day withdrawal group after 14 day consecutive injection of kanamycin[TUNEL positive cells:Corti's organ:0.69± 0.57,spiral ganglion(SG):1.22± 0.51,stria vascularis(SV):1.01± 0.67].The apoptotic cochlear cells were increased over time of withdrawal(TUNEL positive cellsin in 14 day withdrawal group:Corti's organ:4.91± 0.94,SG:8.84± 1.35,SV:7.39± 1.63),and from basal turn to apical turn,the apoptotic cochlear cells were decreased gradually.No cell apoptosis was detected in the control animals.② As the withdrawal time prolonged after injection of chronically ototoxic kanamycin,the ABR threshold in 14 day withdrawal group(43.20± 6.73) was significantly higher than that in the control group(5.60± 1.78)(PCONCLUSION:① Chronically ototoxic kanamycin can induce the apoptosis of cochlear cells in guinea pigs.Cell apoptosis is a mode of cochlear cell injury after injection with kanamycin in guinea pigs.② The apoptosis of cochlear cells might be one of morphological bases in hearing loss induced by the chronic ototoxicity of kanamycin.