年龄相关性核性白内障晶状体核中不可逆聚集体的定性和定量分析  

Qualitative and quantitative analysis of irreversible aggregates in lens nuclei of age-related nuclear cataract

作　　者：苏胜[1] 刘平[1] 冷非[1] 关立南[1] 田霈[1] 王朝[1] 

机构地区：[1]哈尔滨医科大学附属第一医院眼科医院,黑龙江省哈尔滨市150001

出　　处：《眼科新进展》2013年第1期9-12,共4页Recent Advances in Ophthalmology

基　　金：国家自然科学基金资助(编号:30973275);"十一五"国家科技支撑计划重点项目资助(编号:2007BAI07A05)~~

摘　　要：目的鉴定分析在年龄相关性核性白内障(age-related nuclear cataract,ARNC)晶状体核中发现的不可逆聚集体的蛋白质组成。方法用强解聚试剂和促溶剂溶解ARNC晶状体核(8例)和年龄匹配的正常晶状体核(8例),提取总蛋白,经十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)分离,在ARNC晶状体样品中发现的相对分子质量高的聚集体进行液相色谱串联质谱(LC-MS/MS)鉴定分析,确定聚集体的组成成分,并根据Mascot搜索数据估算各成分的相对含量。结果 SDS-PAGE结果显示,与正常晶状体核相比,ARNC样品在相对分子质量≤20000处蛋白含量显著减少(均为P<0.05),但在相对分子质量高(>200000)处蛋白质含量显著增加(均为P<0.01)。在ARNC晶状体内存在不可逆的相对分子质量高的聚集体。LC-MS/MS分析显示,这些聚集体主要来源于各种晶状体蛋白,同时还包括Filensin、Phakinin和碳酰还原酶1。在所有组分中αA-晶状体蛋白含量最高(19.1%)。αA-、αB-、γD-晶状体蛋白和DKFZp434A0627比其他晶状体蛋白更容易聚集。结论晶状体蛋白,尤其是αA-晶状体蛋白在白内障产生和发展过程中发生不可逆的聚集,细胞骨架蛋白1可能参与和加速这一过程。Objeetive To determine the protein compositions of irreversible ag- gregates found in lens nuclei of age-related nuclear cataract （ARNC）. Methods ARNC （8 cases） and age-matched normal lens nuclei （8 cases） were respectively ana- lyzed by powerfttl deaggregating reagents and chaotropes to extract total proteins. After sodium dodecyl sulfate polyacrylamide gel electrophoresis （ SDS-PAGE）, the high mo- lecular weight （HMW） aggregates found in ARNC lens nuclei were identified by liquid chromatography tandem mass spectrometry （LC-MS/MS）. The compositions of the irre- versible aggregates and their relative abundance were investigated based on the data of Mascot search. Results SDS-PAGE showed decreased protein levels at Mr〈.20 000 in ARNC lenses （ all P 〈 0.05 ）, but significantly increased levels at HMW （Mr 〉 200 000, all P 〈 0.01 ）. Irreversible HMW aggregates were present in ARNC lens. LC-MS/MS analysis showed that the HMW aggregates derived largely from crystallins also contained filen- sin,phakinin,and carbonyl reductase 1. In all the components, oua,-crystallin accounted for the highest fraction （19.1%）. aA-,aB-,and y/D-crystallin and DKFZp434A0627 were more prone to aggregate than other crystallins. Conelusion The crystallins, especial- ly aA-crystallin, aggregate irreversibly during ARNC development. Filensin, phakinin, and carbonyl reductase 1 may also be involved in and/or accelerate this process.

关 键 词：晶状体 年龄相关性核性白内障 不可逆聚集体 蛋白质组 液相色谱串联质谱 

分 类 号：R779.6[医药卫生—眼科]