葛根素对庆大霉素所致小鼠耳蜗毛细胞损伤的拮抗效应

Antagonistic effects of puerarin against gentamycin-induced hair cells' injury in the cochlea of mouse

目的观察葛根素对庆大霉素所致小鼠耳蜗毛细胞损伤的拮抗效应。方法完整分离小鼠耳蜗基底膜,无血清培养液常规培养24小时,然后同时加入0.3ml庆大霉素和不同浓度葛根素(1mg/ml,2mg/ml,4mg/ml),共同培养24h,并设立平行对照。标本置荧光显微镜下观察,行全耳蜗毛细胞计数,应用全耳蜗毛细胞定量分析软件自动分析,根据耳蜗毛细胞损失率的变化确定葛根素对庆大霉素所致耳蜗毛细胞损伤效应的影响。结果不同浓度组葛根素均表现小鼠耳蜗毛细胞损失率显著降低(P<0.05～0.01),但以4mg/ml浓度效果最好,呈现剂量依赖性特点。结论葛根素对庆大霉素所致的小鼠耳蜗毛细胞损伤有明显拮抗效应。

Objective To investigate the antagonist effects of puerarin against gentamycin-induced hair cells injury in mouse cochlea based on an experimental study. Methods The experiments were carried out in vitro among newborn mice, with their intact basilar membranes being separated out from the cochlea of them. Then, basilar membranes were routinely cultured in serum free medium for 24 hrs, followed by the co-culturing program with gentamycin at a concentration of 0.3 mM mixed with various concentration levels of puerarin at 1 mg/ml, 2 mg/ml or 4 mg/ml for another 24 hrs respectively and with several paralleled controls. By the end of culturing procedures, all fixed specimens were stained with 1 ： 200 TRITC labeled phalloidin solution and then, observed under fluorescence microscope at 400× to do whole cochlear hair cells counting, supported by hair cells quantitative analysis software. As a result of the automatic analysis, the missing rates of hair cells were calculated for different series of experimentalsystem so as being able to evaluate the antagonist effects ofpuerarin against gentamycin-induced inner ear injury in mice＇s cochlea. Results It was shown from these experiments that various groups of puerarin at different concentrations could induce significantly reduced hair cells missing rates in a dose dependent way respectively （P〈0.05- 0.01）, while the most significant antagonist effect on gentamycin-induced inner ear damage was brought by this reagent at the concentration of 4 mg/ml. Conclusion It can be concluded from this study that puerarin holds very marked antagonistic effect against gentamycin-induced hair cells＇ damage in the inner ear of mice.