应用嗜热菌蛋白酶快速分离活性纯椭圆囊感觉上皮细胞及其意义

Isolation of Pure Living Utricular Sensory Epithelial Cell by Use of Thermolysin

刘俊;孔维佳;张亚民;张丹;

1:华中科技大学同济医学院附属协和医院耳鼻咽喉科

2:华中科技大学同济医学院附属协和医院耳鼻咽喉科

3:华中科技大学同济医学院附属协和医院耳鼻咽喉科

4:华中科技大学同济医学院附属协和医院耳鼻咽喉科 武汉430022

摘要
目的建立纯椭圆囊感觉上皮细胞(pure utricular sensory epithelial cell,PUSEC)的快速分离方法,为内耳细胞培养及毛细胞再生等机制研究提供大量的活性细胞。方法通过对出生5天大鼠PUSEC的形态、生长特征的观察,采用透射电镜,紧密联接蛋白(ZO-1)、细胞角蛋白、波形蛋白免疫荧光细胞化学、RT-PCR检测毛细胞的特征性标志物Calretinin、Brn3.a mRNA的表达等方法鉴定PUSEC来源。结果PUSEC呈扁平、多角形、核大而圆的上皮细胞形态,细胞之间连接紧密,形成单层时呈“铺路石样”外观。可见由成百个PUSEC包绕液体而成的dome。PUSEC表达ZO-1和细胞角蛋白,不表达波形蛋白,表达毛细胞的特征性标志物Calretinin、Brn3.a的mRNA。结论PUSEC来源于椭圆囊感觉上皮;纯椭圆囊感觉上皮细胞成功分离为内耳细胞培养及毛细胞再生机制的研究提供大量的活性细胞;成功分离的关键成份是嗜热菌蛋白酶。
关键词
嗜热菌蛋白酶;椭圆囊;感觉上皮;细胞培养;毛细胞;再生;支持细胞
基金项目(Foundation):
国家自然科学基金(编号39925035);; 国家杰出青年基金(编号39925035);; 卫生部临床学科重点项目基金(编号2001321)资助
作者
刘俊;孔维佳;张亚民;张丹;
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