耳聋左慈丸对老年性耳聋小鼠耳蜗组织cAMP含量及PKA表达的影响

Effect of Erlong Zuoci Pill on Expression Levels of cAMP and PKA in Tissue of Mice with Presbycusis

目的:通过研究耳聋左慈丸对老年性耳聋小鼠耳蜗组织cAMP含量及PKA表达的影响,探讨老年性耳聋的治疗机制。方法:选取75只耳廓反应灵敏的雄性清洁级昆明小鼠,随机抽取15只作为空白对照组,腹腔注射生理盐水。剩余60只进行老年性耳聋小鼠造模,腹腔注射D-半乳糖(150 mg/kg),每日1次,共60 d。造模成功后将老年性耳聋模型小鼠随机分为模型组、耳聋左慈丸低、中、高剂量组。空白对照组和模型组给予生理盐水灌服,耳聋左慈丸低、中、高剂量组给予耳聋左慈丸(0.3、0.6、1.2 g/kg)灌服,每日1次,共14 d。运用耳蜗基底膜铺片技术观察小鼠耳蜗毛细胞形态学变化;采用ELISA法检测耳蜗组织中cAMP含量;用免疫组织化学技术检测PKA的蛋白表达水平;用RT-PCR技术检测PKA mRNA表达水平。结果:与空白对照组相比,模型组小鼠耳蜗基底膜毛细胞明显缺失或散乱,cAMP含量及PKA蛋白和mRNA表达明显下降;与模型组相比,经过耳聋左慈丸治疗的小鼠耳蜗毛细胞排列较整齐,cAMP含量及PKA蛋白和mRNA表达升高,其中高剂量组变化最为明显。结论:耳聋左慈丸可以通过调节耳蜗组织中cAMP含量及PKA的表达水平发挥其对老年性耳聋的治疗作用。

Objective： To discuss the therapeutic mechanism of presbycusis by researching the effect of Erlong Zuoci pill on cAMP content and PKA in tissue of mice with presbycusis. Methods： Seventy-five auricle responsive cleaning degree male Kunming mice were randomly selected and fifteen as the blank control group,which were treated with NS in empty stomach. The other sixty were the presbycusis modeling group,which were treated with D-galactose（ 150 mg/kg） in empty stomach,once a day,with sixty days. After duplicating the model,the presbycusis modeling mice were randomly divided into the modeling group,Erlong Zuoci Pill low-dose group,middle-dose group and high-dose group,with fifteen mice in each group. The blank control group and the modeling group were drenched with NS and the Erlong Zuoci Pill low-dose group,mid-dose group and high-dose group were drenched with concentrated solutions of Erlong Zuoci Pill（ 0. 3 g/kg,0. 6 g/kg,1. 2 g/kg）,once a day,with fourteen days. It observed morphological change of cochlear hair cells in mice to use cochlear basilar membrane whole-mount technique. It tested the cAMP content by using ELISA,the protein expression level of PKA by IHC and the expression level of PKA and mRNA by using RT-PCR technique. Results： Compared with CG,basement membrane cells of mice cochlea in modeling group were obviously lacked and scattered,and cAMP content,protein expression of PKA and mRNA expression were clearly declined. Compared with the modeling group,cochlear hair cells in mice treated with Erlong Zuoci Pill tidily arranged. The cAMP content,protein expression of PKA and mRNA expression clearly increased,with significant changes in high-dose group,with high increasing in high-dose. Conclusion： Erlong Zuoci Pill treats presbycusis through adjusting the cAMP content and expression level of PKA in cochlear tissue.