狼蛛毒素在胆红素诱导的耳神经毒性中作用的实验研究  

Experimental study on the effect of psalmotoxin-1 in bilirubin-induced ototoxicity

作　　者：张玲[1] 叶海波[1] 邢雅智 丁大连[2] 时海波[1] ZHANG Ling;YE Haibo;XING Yazhi;DING Dalian;SHI Haibo(Department of Otolaryngology Head and Neck Surgery,Shanghai Sixth People's Hospital,Institute of Otolaryngology,Shanghai Jiaotong University,Shanghai Oriental Institute of Otolaryngology,Shanghai,200233,China;Center for Hearing and Deaf ness,the State University of New York College at Buffalo,New York,Buffalo,14214,USA)

机构地区：[1]上海交通大学附属第六人民医院耳鼻咽喉头颈外科,上海交通大学耳鼻咽喉科研究所,上海东方耳鼻咽喉科研究所,上海200233 [2]纽约州立大学布法罗分校听力与耳聋中心,美国纽约布法罗14214

出　　处：《中国耳鼻咽喉头颈外科》2021年第1期26-30,共5页Chinese Archives of Otolaryngology-Head and Neck Surgery

基　　金：国家自然科学基金资助项目(81800919)。

摘　　要：目的观察狼蛛毒素(psalmotoxin-1)对胆红素诱导的耳蜗组织毒性作用。方法新生3 d的SD大鼠,断头解剖获得耳蜗基底膜离体培养24 h,将耳蜗组织随机分5个组,对照组、50μmol/L胆红素组、100μmol/L胆红素组、100 ng/ml psalmotoxin-1组和100μmol/L胆红素+100 ng/ml psalmotoxin-1组。实验组培养基按各分组浓度,加入相应胆红素和(或)psalmotoxin-1溶液,对照组加入相应体积生理盐水。所有耳蜗样本继续培养24 h后终止,进行组织固定和免疫荧光染色,激光共聚焦显微镜观察,最后进行统计学分析。结果不同浓度胆红素作用后,50μmol/L胆红素组基底膜螺旋神经节神经元细胞(spiral ganglion neurons,SGNs)开始出现不同程度减少,但耳蜗毛细胞(hair cells,HCs)形态及数量完整;100μmol/L胆红素组耳蜗组织SGNs出现细胞体积明显缩小,伴随细胞核浓缩或碎裂,细胞数目明显减少,HCs出现不同程度损伤,SGNs数目与对照组比较,差异有统计学意义(t=18.89,P<0.0001)。100 ng/ml psalmotoxin-1组基底膜SGNs和HCs形态及数量较对照组均未出现明显改变(P均>0.05);100μmol/L胆红素+100 ng/ml psalmotoxin-1组SGNs形态变化及数量减少较单纯100μmol/L胆红素组明显减轻(t=21.63,P<0.0001)。结论在耳蜗器官培养中,胆红素以剂量依赖性方式损伤新生大鼠耳蜗SGNs和HCs;胆红素对耳蜗SGNs的损伤较HCs发生早,损伤程度重。在耳蜗器官培养中,psalmotoxin-1可减弱胆红素诱导的耳蜗毒性作用。OBJECTIVE To observe the effect of tarantula toxin(psalmotoxin-1)on bilirubin-induced cochlear tissue toxicity.METHODS The basal membrane of the cochlea was obtained by dissecting the severed head of 3-day-old SD rats and cultured in vitro for 24 hours.The cochlear tissues were randomly divided into 5 groups:control group,50μmol/L bilirubin group,100μmol/L bilirubin group,100 ng/ml psalmotoxin-1 group and 100μmol/L bilirubin+100 ng/ml psalmotoxin-1 group.According to the group concentration,bilirubin and(or)psalmotoxin-1 solution were added to the culture medium of the experimental group,and normal saline was added to the control group.All the cochlear samples were cultured for 24 hours and then the tissues were fixed and immunofluorescence stained.The laser confocal microscopy was used to observe the samples,and finally analyzed statistically.RESULTS In the 50μmol/L bilirubin group,SGNs began to decrease in varying degrees,but the morphology and quantity of hair cells were complete.In the 100μmol/L bilirubin group,the cell volume of SGNs was significantly reduced with nuclear concentration or fragmentation,the cell number was significantly reduced,HCs was damaged in varying degrees,and the number of SGNs was statistically different from that of the control group(t=18.89,P<0.0001).The morphology and quantity of SGNs and HCs in the basement membrane of 100 ng/mlpsalmotoxin-1 group were not significantly changed compared with that of the control group(P all>0.05).The morphological and the number changes of SGNs in 100μmol/L bilirubin+100 ng/ml psalmotoxin-1 group were significantly less than that in 100μmol/L bilirubin group(t=21.63,P<0.0001).CONCLUSION In cochlear organ culture,bilirubin damages SGNs and HCs of newborn rats in a dose-dependent manner.The damage of bilirubin to SGNs was earlier and more serious than HCs.In cochlear organ culture,psalmotoxin-1 can attenuate bilirubin induced cochlear toxicity.

关 键 词：动物实验 胆红素 毛细胞 听觉 螺旋神经节 狼蛛毒素 

分 类 号：R722.17[医药卫生—儿科]