一氧化氮对过氧化氢所致听力损失的保护作用

通过全耳蜗灌流法在体观察一氧化氮(N0)能否通过一氧化氮/环磷酸鸟苷(NO/cGMP)途径对抗过氧化氢这种氧自由基所致的听力损失。实验选用耳廓反射灵敏、无耳毒性药物使用史的健康杂色豚鼠(250-350 g)50只,雌雄不拘,随机分为5组,每组10只动物,分别行全耳蜗灌流人工外淋巴液;过氧化氢(H2O2);L-精氨酸(合成NO的底物);H2O2+L-精氨酸;H2O2+L-精氨酸+L-NNA(一氧化氮合成酶的抑制剂),均灌流2 h。通过圆窗龛电极,每隔30 min记录复合动作电位(compound action potential,CAP:由短声Click诱发)阈值,耳蜗微音器电位(cochlear microphonic,CM;由短纯音Tone Burst诱发)幅度,了解耳蜗功能的变化,并分离取出耳蜗基底膜并制备基底膜硬铺片,通过碘化毗啶(PI)和Hoecbst双染色方法,观察耳蜗组织各类细胞损伤情况。结果显示,灌流H2O2+L-精氨酸组的CAP阈移和CM下降幅度值明显低于单独灌流H2O2组,差异有显著性(氏P<0.05);前者形态学观察未见明显的细胞损伤,后者可见大量坏死红染的细胞。H2O1+L-精氨酸+L-NNA组CAP阈移和CM下降幅度与单独灌流H2O2组比较无统计学差异。实验结果提示NO可能通过NO/cGMP途径部分对抗过氧化氢所致的听力损失。

Protective effect of nitric oxide against hydrogen peroxide-induced hearing loss

Previous research showed that reactive oxygen species (ROS) play an important role in ototoxity. The present research was to investigate whether nitric oxide, an important neurotransmitter in the inner ear, could prevent hydrogen peroxide-induced hearing loss through the nitric oxide/cyclic GMP pathway in guinea pig cochlea. Fifty adult pigmented guinea pigs (250~350 g) of either sex with positive prier reflex were randomly divided into five groups. All of the animals underwent whole cochlear perfusion for two hours. The solution that was perfused into the cochlear of different group was artificial perilymph (AP) for group 1200 micromol/L H2O2 for group 2100 micromol/L L-Arg for group 3, H2O2+L-Arg for group 4 and H2O2+L-Arg+L-NNA for group 5 respectively. Compound action potential (CAP, evoked by click) and cochlear microphonic (CM, evoked by tone burst) were recorded every thirty minutes to show the effects of different reagents on cochlear function. In order to assess cell viability after perfusion, the fluorescent dyes Hoechst that stains all cell nuclei and propidium iodide (PI) that specifically stains nuclei of dead cells, were used. The CAP threshold shifts and CM amplitude decreased after perfusion with H2O2+L-Arg. They were significantly lower than those of H2O2 group. No obvious cell death was noticed after H2O2+L-Arg perfusion, while only 54% of hair cells were alive after H2O2 perfusion. There were no significant differences between the group of H2O2 and that of H2O2+L-Arg+L-NNA group. Our results suggest that nitric oxide may partly be able to protect guinea pigs from hydrogen peroxide-induced hearing loss.