氯沙坦对自发性2型糖尿病KKAy小鼠肾小球蛋白表达谱的影响  

Effect of losartan on the glomerular protein expression profile of type 2 diabetic KKAy mice

作　　者：范秋灵[1] 杨刚[1] 刘晓丹[1] 马健飞[1] 冯江敏[1] 姜奕[1] 王力宁[1] 

机构地区：[1]中国医科大学附属第一医院肾内科,沈阳110001

出　　处：《中华肾脏病杂志》2012年第6期476-483,共8页Chinese Journal of Nephrology

基　　金：国家自然科学基金（30700369）;辽宁省教育厅科学技术研究项目（L2010658）;沈阳市科技计划项目（F11-264-1-38）

摘　　要：目的利用比较蛋白质组学双向电泳技术研究体系来观察血管紧张素受体拮抗剂（ARB）氯沙坦对自发性2型糖尿病KKAy小鼠肾小球蛋白表达谱的影响。方法8周龄自发性2型糖尿病KKAy小鼠随机分为氯沙坦治疗组（饮用水中喂入氯沙坦粉末10mg·kg^-1·d^-1）和非治疗组；8周龄C57BL／6小鼠作为正常对照组。饲养12周后，经胸主动脉磁珠灌流分离肾小球，提取肾小球蛋白。DIGE最小荧光标记法标记，行二维荧光差异凝胶电泳。应用Typhoon多功能成像系统扫描凝胶，DeCyder2D差异分析软件进行图像分析。采用基质辅助激光解析电离飞行时间质谱（MALDI-TOF-MS）鉴定差异表达蛋白质。结果KKAy小鼠的体质量、血糖和尿白蛋白排泄率较正常对照C57BL／6小鼠显著升高（均P〈0．05），氯沙坦治疗显著改善2型糖尿病KKAy小鼠尿白蛋白肌酐比[（539．71±100．23）mg／g比（728±177．19）mg／g]、肾小球基底膜增厚和系膜基质增生等肾脏病理损害，而对血糖无影响。通过DeCyder2D差异分析软件，发现KKAy氯沙坦治疗组与KKAy非治疗组肾小球内表达差异大于1．1倍的蛋白质斑点62个。经肽质量指纹图分析，鉴定出41种蛋白质。其中表达上调的蛋白28种，包括甘油激酶、亚硫酸盐氧化酶、甘氨酸脒基转移酶、腺苷高半胱氨酸酶等；表达下调的蛋白质13种，包括3．巯基丙酮酸硫基转移酶、ATP合酶亚单位d、60000热休克蛋白、线粒体应激蛋白70（又名75000葡萄糖调节蛋白，GRP75）等。有6种蛋白在20周龄KKAy小鼠和C57BL／6小鼠肾小球内存在差异表达，氯沙坦治疗抑制了其在糖尿病状态下的表达变化，包括丙酮酸脱氢酶复合物的二氢硫辛酰赖氨酸残基乙酰基转移酶组分、琥珀酰辅酶A连接酶[GDP-形成1亚单位β、ATP合酶亚单位d、GRP75、核苷二磷酸盐结合部分X模体19和硒结合蛋白1。结论氯沙坦可明显减轻KKAyObjective To investigate the effects of angiotensin receptor blocker （ARB） losartan on the glomerular protein expression profile of spontaneous type 2 diabetic KKAy mice by two-dimensional differential gel electrophoresis and MALDI-TOF mass spectrometry. Methods 8-week-old spontaneous type 2 diabetic KKAy mice were randomly divided into losartan （10 mg· kg-1· d-1 given in drinking water） treatment group and non-treatment group. Eight-week-old C57BL/ 6 mice were used as normal control. The glomeruli were separated by magnetic bead perfusion through thoracic aorta at age of 20 weeks, then glomerular protein was extracted. The glomemlar protein expression profile was investigated by CyDyes minimal fluorescence labelling, two- dimensional differential gel electrophoresis and MALDI-TOF mass spectrometry. Results KKAy mice developed higher body weight and blood glucose, higher urinary microalbumin creatinine ratio at age of 20 weeks than C57BL/6 mice at the same age （all P〈0.05）. Losartan treatment markedly reduced urinary microalbumin creatinine ratio [（539.71 ±100.23）mg/g vs （728±177.19） mg/g], attenuated mesangial expansion and the thickening of glomerular basement membrane, but had no effect on the blood glucose. By DeCyder 2-D differential analysis software, 62 protein spots of differential expression were found in glomeruli between losartan treatment and non-treatment KKAy mice at age of 20 weeks. Among them, 41 proteins were identified by peptide mass fingerprinting. The expressions of 28 proteins were up-regulated by losartan treatment, including glycerokinase, sulfite oxidase, glycine amidinotransferase, adenosylhomocysteinase, etc. The expressions of 13 proteins were down-regulated by losartan treatment, including 3-mercaptopyruvate suffurtransferase, ATP synthase subunit d, 60 000 heat shock protein, stress-70 protein （alternative name 75 000 glucose-regulated protein, GRP75）, etc. Six differentially expressed proteins were found in glomeruli between non-treatment KKAy mice and

关 键 词：糖尿病肾病 肾小球 蛋白质组学 氯沙坦 KKAY小鼠 

分 类 号：R285.5[医药卫生—中药学]