硫酸链霉素对体外培养大鼠前庭毛细胞的损害作用

Ototoxic effects of streptomycin in vestibular organotypic cultures

目的探讨硫酸链霉素对离体培养条件下大鼠前庭毛细胞的损害特征。方法取出生后3—4d的大鼠球囊斑和椭圆囊斑进行离体前庭器官培养，经培养过夜后再用不同浓度的硫酸链霉素培养液继续培养24h。应用荧光标记的鬼笔环肽对前庭毛细胞的静纤毛和表皮板进行染色，同时应用Topro-3DNA探针显示细胞核的形态，在激光共聚焦显微镜下计数和摄像。结果在离体培养条件下，对照组前庭毛细胞生长良好，随着培养液中硫酸链霉素浓度的增加，毛细胞的密度逐渐降低。培养24h后，0．25mmol／L硫酸链霉素可造成大约10％的前庭毛细胞缺损，1mmol／L硫酸链霉素损害的毛细胞数量在50％左右，3mmol／L硫酸链霉素破坏的毛细胞数量超过75％。细胞核染色显示硫酸链霉素引起的前庭毛细胞损害伴随着细胞核浓缩或破碎现象。存活毛细胞的数量随着硫酸链霉素浓度的增加而减少，但前庭的支持细胞未发生明显病变。结论在体外培养条件下，硫酸链霉素可引起剂量依赖性的大鼠前庭毛细胞缺失，其损伤机制可能与凋亡有关。

Objective To investigate the ototoxic effects of streptomycin in vestibular organotypic cultures. Methods F344 rats with age at postnatal day three or four were used for this study. The maculae of saccule and utricle were routinely dissected out and cultured with serum-free medium containing various dose of streptomycin for 24 hours. The ciliary turf of vestibular hair cells was stained with fluorescent phalloidin, and its nucleus was stained with topro-3 DNA probe. The vestibular hair cells were quantitatively counted and photographed under eonfocal fluorescent microscope. Results Morphological feature of vestibular hair cells were good in normal control cultures. However, the density of hair cells was decreased in evidence with increase of streptomycin sulfate concentrations. Twenty-four hours after streptomycin cultures, 0. 25 mmol/L streptomycin caused a 10% hair cell missing, 50% hair cell loss from 1 mmol/L streptomycin treatment, and more than 75% hair cells gone post-3 mmol/L streptomycin cultures. After streptomycin treatment, the nuclear shrinkage and fragmentation were found in vestibular hair cells, whereas the vestibular supporting cells were normal. Conclusion Streptomycin induced-vestibular hair cells lesion was in a dose dependent manner with nuclear shrinkage and fragmentation. This may suggest that streptomycin leads vestibular hair cell die through apoptosis.