褪黑素对小鼠放疗后内耳Prestin蛋白表达的影响

Effect of melatonin on expression of Prestin protein in the inner ear of mice following radiotherapy

目的探讨褪黑素（melatonin）对一次性大剂量放射治疗（放疗）后小鼠内耳Prestin蛋白表达的影响，为研究放射性内耳损伤的机制并制定预防方案提供依据。方法4周大雄性Balb／c小鼠60只完全随机分为6组，包括对照组（A组）、50mg／kg褪黑素组（B组）、5mg／kg褪黑素组（C组）、50mg／kg褪黑素＋放疗组（D组）、5mg／kg褪黑素＋放疗组（E组）和放疗组（F组）。每个实验组再完全随机分成两组，分别于16Gy放疗后的第3、7天收集耳蜗标本行免疫组化染色及蛋白质印迹法（Westernblot）检测Prestin蛋白的表达情况。采用SPSS19．0软件进行统计分析。结果Prestin蛋白主要分布在耳蜗外毛细胞细胞核以上的侧膜，F组与A、B、C组相比，内耳Prestin蛋白表达上调，差异有统计学意义（P值均〈0．05）；D、E组能降低放疗后Prestin蛋白的异常表达，与F组相比，差异具有统计学意义（P值均〈0．05），且D组较E组效果显著，差异有统计学意义（P〈0．05）。结论耳蜗Prestin蛋白主要分布在外毛细胞细胞核以上的侧膜，接受大剂量放疗后Prestin表达上调；褪黑素可抑制放疔后Prestin蛋白的异常表达，作用与剂量相关。

Objective To investigate the effect of melatonin on the expression of prestin protein in the inner ear of mice following a single dose radiation therapy, so as to provide the basis for the mechanism study of radiation induced inner ear injury and its prevention. Methods Sixty 4-week-old male mice were randomly divided into six groups, including the control group （A group）, 50 mg/kg MLT group （B group）, 5 mg/kg MLT group （ C group ） , 50 mg/kg MLT ＋ radiotherapy group （ D group ） , 5 mg/kg MLT ＋ radiotherapy group （E group ）, and 16 Gy radiotherapy group （F group）. Each experimental group was randomly subdivided into two subgroups, which were killed to harvest the cochlea on the 3rd and 7th days following 16 Gy radiation. The specimens were used for immunostaining and Western blot to detect the expression of prestin protein. SPSS 19.0 software was used for statistical analysis. Results Prestin protein mainly distributed in the lateral membrane above the outer hair cell nucleus. When compared with A, B and C group, the expression of prestin protein in the inner ear was significantly up-regulated in F group （P 〈 0. 05 ）. However, D and E group reduced the abnormal expression of prestin following radiotherapy when compared with F group, the difference was statistically significant （ P 〈 0.05 ）, and the effect of D group was more significant than E group （ P 〈 0. 05 ）. Conclusions The prestin protein of cochlea is mainly distributed in the lateral membrane above the outer hair cell nucleus. Following the high-dose radiotherapy, the prestin expression is upregulated, and melatonin can control the abnormal expression of prestin protein induced by radiotherapy with dose dependent.