糖尿病性白内障和年龄相关性白内障晶状体蛋白质组学的差异分析

摘要

Background With the changes of diet and living style,the diabetes has become the major diseases affecting human health.Diabetic cataract is a common complication of diabetes. Objective The present study was to investigate the difference of lens proteomics between diabetic cataract and age related cataract using two dimensional electrophoresis (2-DE) and mass spectrometry in order to postpone happening of diabetic cataract and offer the effective approach to the prevention and therapy of diabetic cataract. Methods The lenses were obtained from 8 diabetic patients and 12 age-related cataract patients during the surgery to extract the protein by lysis and centrifugation.The lens proteins were separated using immobilized pH gradients 2-DE.Image analysis was carried out using PDQuest Advanced-8.0.1 software package.Significant difference of the crystallines was identified by matrixassisted laser adsorption/ionization time of-flight-mass spectrometry (MALDI-TOF-MS) and peptide mass fingerprint combined with protein database. Results The maps of 2-DE showed that lens proteins of diabetic cataract and age related cataract were in the section of pH 5-9 with the relative molecular weight 14000-97000;while relative molecular weight of more abundant crystalline was localized at 20000-31000.About 3 differential protein spots were detected by image analysis software.Two crystallines,αB and βB1 crystallin,were identified using MALDI-TOF-MS.Conclusions Proteomic analysis of lens can be accomplished and the proteins can be well separated,moreover,differential proteins can be analyzed using 2-DE and mass spectrometry between diabetic cataract and age related cataract.These results indicate that αB and βB1 crystallin proteins accelerate the development of diabetic cataract.This technique offers a new avenue for clarity of lens proteins of diabetic cataract other than age related cataract.%背景 目前糖尿病已成为影响人类健康的主要疾病之一,糖尿病性白内障为糖尿病患者的常见并发症之一,进行糖尿病性白内障晶状体中的蛋白质组学研究对该病的防治具有重要意义. 目的 研究糖尿病性白内障和年龄相关性白内障患者晶状体蛋白质组学的差异,探讨糖尿病性白内障的发病机制.方法 手术采集诊断明确的糖尿病性白内障患者8例8眼及年龄相关性白内障患者12例12眼的晶状体,裂解离心提取蛋白,采用固相pH梯度(IPG)等电聚焦双向凝胶电泳法对糖尿病性白内障和年龄相关性白内障患者的晶状体蛋白质进行分离,分析电泳图像的差异,使用基质辅助激光解析离子飞行时间质谱仪( MALDI-TOF-MS)测定蛋白质斑点的肽质量指纹谱,结合蛋白质数据库检索,确定蛋白质种类. 结果 IPG双向凝胶电泳显示,糖尿病性白内障和年龄相关性白内障的晶状体蛋白质相似处在于大部分都分布在等电点( PI) pH值5～9、相对分子质量14000 ～97000的区域内；而高丰度晶状体蛋白质的相对分子质量为20000 ～ 31000.分析软件检测出3个差异的蛋白质点,经质谱鉴定得到其中2个高丰度晶状体蛋白质的种类为αB晶状体蛋白和βB1晶状体蛋白.结论 双向电泳联合质谱鉴定可有效分离晶状体蛋白质,糖尿病性白内障与年龄相关性白内障晶状体蛋白质组学之间的差异表明,αB晶状体蛋白和βB1晶状体蛋白加速了糖尿病性白内障的发展.