Department of Nephrology, The First Hospital, China Medical University, Shenyang, China.
J Nephrol. 2013 May-Jun;26(3):517-26. doi: 10.5301/jn.5000176. Epub 2012 Jun 7.
To investigate the pathogenesis of diabetic nephropathy (DN) and to search for novel therapeutic targets, the glomerular protein expression profile of KKAy mice treated by losartan was analyzed by two-dimensional differential gel electrophoresis (2D-DIGE).
The eight-week-old KKAy mice were divided into the losartan treatment group and the non-treatment group, and C57BL/6 mice were used as the control group. After 12 weeks treatment, glomeruli were isolated by abdominal perfusion with magnetic beads, and the glomerular proteins were extracted. The glomerular protein expression profiles were investigated using 2D-DIGE and MALDI-TOF mass spectrometry. Western blot analysis was used to confirm the results of proteomics.
Losartan treatment improved albuminuria and renal pathologic lesion of KKAy mice. A total of 62 glomerular proteins were differentially expressed between the KKAy losartan treatment mice and KKAy non-treatment mice. Among them, the expression of 28 proteins were up-regulated, including glycerokinase, sulfite oxidase, glycine amidinotransferase, and adenosylhomocysteinase. The expression of 13 proteins were down-regulated, including 3-mercaptopyruvate sulfurtransferase, ATP synthase subunit d, 60 kDa heat shock protein, and 75 kDa glucose-regulated protein(GRP75). A total of six proteins were found to be differentially expressed between the KKAy non-treatment mice and C57BL/6 mice glomeruli, and their differential expression was suppressed by losartan treatment, including mitochondrial ATP synthase subunit d, GRP75, and selenium-binding protein 1 et al.
Treatment with losartan suppresses the differential expression of mitochondrial ATP synthase subunit d, GRP75, selenium-binding protein 1 etc. In diabetic KKAy mice glomeruli, may play a renoprotective role by reducing glomerular mitochondrial ROS genesis and inhibiting oxidative stress.
为了研究糖尿病肾病（DN）的发病机制并寻找新的治疗靶点，本研究采用二维差异凝胶电泳（2D-DIGE）分析了氯沙坦治疗的 KKAy 小鼠的肾小球蛋白表达谱。
将 8 周龄 KKAy 小鼠分为氯沙坦治疗组和未治疗组，并用 C57BL/6 小鼠作为对照组。用磁珠腹腔灌注分离肾小球，提取肾小球蛋白。采用 2D-DIGE 和 MALDI-TOF 质谱技术研究肾小球蛋白表达谱。Western blot 分析用于验证蛋白质组学结果。
氯沙坦治疗改善了 KKAy 小鼠的蛋白尿和肾脏病理病变。KKAy 氯沙坦治疗组与 KKAy 未治疗组小鼠的肾小球蛋白表达谱存在 62 种差异表达蛋白。其中，28 种蛋白表达上调，包括甘油激酶、亚硫酸氧化酶、甘氨酸酰胺转移酶和腺苷同型半胱氨酸酶。13 种蛋白表达下调，包括 3-巯基丙酮酸硫转移酶、ATP 合酶亚基 d、60 kDa 热休克蛋白和 75 kDa 葡萄糖调节蛋白（GRP75）。KKAy 未治疗组与 C57BL/6 小鼠肾小球之间存在 6 种差异表达蛋白，氯沙坦治疗可抑制其差异表达，包括线粒体 ATP 合酶亚基 d、GRP75 和硒结合蛋白 1 等。
氯沙坦抑制了线粒体 ATP 合酶亚基 d、GRP75、硒结合蛋白 1 等在糖尿病 KKAy 小鼠肾小球中的差异表达，通过减少肾小球线粒体 ROS 生成和抑制氧化应激，可能发挥肾脏保护作用。
