水通道蛋白在内淋巴囊和肾脏的表达及加压素对水通道蛋白表达的影响

Expression of aquaporin in rats′endolymphatic sac and kidney and the effect of anti-diuretic hormone on the expression of aquaporin

目的 从形态学上直接验证水通道蛋白 (aquaporin ,AQP) 2、3、4在内淋巴囊上皮的表达 ;观察加压素 (anti diuretichormone,AVP)及血管加压素受体 2 (V2receptor,V2 R)受体促效剂去精氨酸加压素 [(deamino Cys1,D Arg8) vasopressin ,dDAVP]对AQP2在内淋巴囊上表达的变化 ,并与肾脏比较 ,为耳肾相关提供参考。方法 成年大鼠 5只 ,经活体心脏灌注 ,取双侧颞骨 ,按石蜡包埋技术处理和切片。使用免疫组织化学方法标记确认AQP2、3、4在内淋巴囊的表达 ;使用成年大鼠 10只观察AVP及DDAVP对AQP2在内淋巴囊和肾脏上表达的变化。结果 荧光显微镜下 ,AQP 2、3、4一抗在内淋巴囊上皮胞膜胞浆显示荧光染色 ,肾脏集合管主细胞胞膜胞浆有稳定清晰染色反应。AVP组 (B组 )和DDAVP组 (C组 )的内淋巴囊上皮胞膜胞浆AQP 2的表达明显较阴性对照组 (A组 )降低 ,表现为荧光减弱 ,图像分析示灰度明显减弱 (P <0 0 1) ;B组和C组AQP 2的表达则差异无显著性。B组和C组肾脏集合管主细胞胞膜胞浆AQP 2的表达明显较A组增强 ,表现为棕黄色颗粒染色明显加深 ,图像分析示灰度明显减弱 (P <0 0 5 ) ,密度明显增加 (P <0 0 5 ) ,总积分密度 (totalintegrateddensity,IOD)明显提高 (P <0 0 5 ) ,面积明显增加 (P <0 0 5 ) ;

Objective To confirm the expression of aquaporin2,3,4(AQP2,3,4, water channel protein) in rats′endolymphatic sac (ES)and kidney, and to investigate and compare the effects of anti-diuretic hormone (AVP) and DDAVP [(deamino-Cys1, D-Arg8)-Vasopressin,V2-receptor agonist] on the expression of AQP2 in rats′ ES and kidney. Methods Thirty healthy Swards white rats were divided into the negative control, AVP group and dDAVP group, respectively, and were cardiacally perfused. The temporal bones and kidneys were taken out, then processed and sectioned by paraffin-embedded technique. The sections of ES were labeled with fluorescent antibody by immunohistochemical method, and kidney′s with avidin-biotin-peroxidase complex method (ABC). The expression of AQP-2,3,4 were confirmed in the ES of rats, and the different effects of the AVP and DDAVP onto the ES and kidney were observed. The slides used were analyzed by image-analyzer and the subsequent data were dealt with statistically. Results In the cytomembrane and cytoplasm of ES′ epithelia, the constant and clear fluorescent reaction could be observed in normal control group with the first antibody of AQP2,3,4. Significant feeble fluorescent reaction of the first antibody of AQP-2 was revealed in AVP group and DDAVP group and showed much lower of gray (P<0.01), less intensive of fluorescent (P<0.01) under the fluorescence microscope. In the principal cell of renal colleting duct, it was on the contrary. Compared with the control group, significant stain was revealed in AVP group and DDAVP group and showed lower of gray (P<0.05), greater density(P<0.05), higher IOD(P<0.05) and more stained area (P<0.05). There is no different between the AVP group and DDAVP group in expression of AQP-2 in two sites. Conclusions AQP-2,3,4 were expressed both in rats′epithelia of endolymphatic sac (ES)and principal cell of renal collecting duct. AVP promotes the expression of AQP2 in kidney but inhibits in ES. AVP maybe play important role to control the expression of AQP-2 in ES and kidney probably by the role of AVP-V2R-cAMP-AQP2.