尿道下裂仔鼠阴茎蛋白质组学分析及膜联蛋白A3的表达变化研究  

Proteomic analysis of the testis and differential expression of Annexin A3 in hypospadiac rats

作　　者：常俊锴[1] 张炜[1] 沈百欣[2] 魏云飞[1] 张力峰[1] 王勇[1] 冯宁翰[1] 

机构地区：[1]南京医科大学第一附属医院泌尿外科,江苏南京210029 [2]南京医科大学第二附属医院泌尿外科,江苏南京210011

出　　处：《中华男科学杂志》2010年第10期877-882,共6页National Journal of Andrology

基　　金：国家自然科学基金(30872596)~~

摘　　要：目的:探讨邻苯二甲酸二丁酯(DBP)胚胎期暴露对仔鼠阴茎组织蛋白表达谱的影响,分离并鉴定差异表达蛋白质。验证膜联蛋白A3在仔鼠阴茎中的表达变化,初步探讨其在尿道下裂发生中的作用。方法:SPF级SD孕鼠20只,妊娠14～18d(GD14～GD18),随机均分为2组,实验组给予DBP按800mg/kg染毒孕鼠,对照组按大豆油5ml/kg,2组孕鼠均每天灌胃1次连续5d。出生后3d取出实验组尿道下裂雄性仔鼠阴茎和对照组正常雄性仔鼠阴茎,提取总蛋白,进行二维凝胶电泳分离和图像分析,筛选出的差异蛋白质点利用质谱技术进行鉴定,并运用Western印迹和免疫组化法分析膜联蛋白A3在仔鼠阴茎中的表达变化。结果:共筛选出31个差异表达蛋白,其中17个通过质谱分析和SwissProt蛋白数据库检索得到鉴定,包括丙酮酸激酶M2、α烯醇化酶、膜联蛋白A3等。膜联蛋白A3相对表达量在尿道下裂组为1.851±0.014(n=10),正常对照组为2.603±0.012(n=10),两组的差异具有显著性(P<0.05)。膜联蛋白A3主要定位于仔鼠尿道上皮细胞。结论:运用蛋白质组学方法,建立了DBP孕期暴露致尿道下裂雄性仔鼠与正常仔鼠阴茎蛋白质差异表达谱系。膜联蛋白A3的表达变化可能在尿道下裂尿道沟融合障碍过程中发挥重要作用。Objective：To investigate the effect of in utero exposure to di-n-butyl phthalate （DBP） on the protein expression in the penile tissue of hypospadiac rats,isolate and identify differentially expressed proteins,and determine the role of the differential expression of Annexin A3 in the development of hypospadia in the rat offspring after maternal exposure to DBP. Methods：Twenty pregnant SD rats were randomly assigned to an experimental group,intragastrically administered DBP at 800 mg/kg,and a control group,given soybean oil at 5 ml/kg,both for 5 days. Three days after birth,the penises of the newborn rats were removed,and the total protein extracted for 2D-electrophoretic separation and image analysis. Differentially expressed protein spots were screened and identified by mass spectrometry,and the changes in the expression of Annexin A3 detected by Western blotting and immunohistochemistry. Results：Thirty-one differentially expressed protein spots were screened,of which 17 were identified by mass spectrometry and the SwissProt database,including pyruvate kinase M2,alpha-enolase,and Annexin A3. Western blot showed that Annexin A3 was mainly located in the urethral epithelia and had a lower expression in the hypospadiac rats （1.851±0.014,n = 10） than in the controls （2.603±0.012,n = 10） （P 0.05）. Conclusion：A pedigree of differentially expressed proteins in the penises of DBP-induced hypospadia and normal rats was established by the proteomic method. The differential expression of Annexin A3 may play an important role in the development of hypospadia.

关 键 词：邻苯二甲酸二丁酯 尿道下裂 蛋白质组学 膜联蛋白A3 

分 类 号：R691.1[医药卫生—泌尿科学]