Identification of diabetic nephropathy-selective proteins in human plasma by multi-lectin affinity chromatography and LC-MS/MS
Identification of diabetic nephropathy-selective proteins in human plasma by multi-lectin affinity chromatography and LC-MS/MS
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DOI：
10.1002/prca.200900196
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发表时间：
2010-07-01
影响因子：
2
通讯作者：
Cho, Je-Yoel
中科院分区：
文献类型：
--
作者：
Ahn, Jung-Mo;Kim, Byung-Gyu;Cho, Je-Yoel
目的糖尿病肾病（diabetic nephropathy，DN）是糖尿病的严重并发症.微量白蛋白尿已被确定为糖尿病肾病发展的危险因素。近年来，微量白蛋白尿在糖尿病肾病的危险性评估和预测方面有一定的局限性，因此，需要寻找更特异性的生物标志物来预测糖尿病肾病。当肾损伤开始时，可能发生糖蛋白渗漏到血液中，从而改变血液的糖蛋白质组谱。在这里，我们使用了血浆糖蛋白富集与蛋白质组学分析相结合的方法来发现潜在的DN生物标志物。我们从6名2型糖尿病对照（DC）和6名2型DN患者的血浆中分离糖蛋白，采用多凝集素亲和色谱法。捕获的糖蛋白通过1-D PAGE和分离的蛋白质的胰蛋白酶解物通过LC-MS/MS进行分析。结果：从比较和半定量蛋白质组学分析，我们确定了13个上调和14个下调的糖蛋白在DN血浆中。在上调的糖蛋白中，Lumican、vasorin和视黄醇结合蛋白-4的水平通过个体血浆样品的Western印迹分析来验证。总的来说，我们的研究结果表明，生物标志物的发现有相当大的潜力预测糖尿病患者的糖尿病肾病。
Purpose Diabetic nephropathy (DN) is a serious complication of diabetes mellitus. Microalbuminuria has been established as a risk factor for the development of diabetic renal disease. Recently, microalbuminuria has been reported to have limitations in determining disease risk and predicting DN Therefore, identification of more specific biomarkers for prediction of DN is needed.Experimental design. When kidney damage is initiated, glycoprotein leakage into the blood may occur, thus altering the glycoproteome profile of the blood. Here, we have used a combined approach of glycoprotein enrichment of plasma with a proteomic analysis to discover potential DN biomarkers. We isolated glycoproteins from plasma provided by six type 2 diabetes control (DC) and six type 2 DN patients using multi-lectin affinity chromatography. Captured glycoproteins were resolved by 1-D PAGE and tryptic digests of isolated proteins were analyzed by LC-MS/MS.Results: From the comparative and semi-quantitative proteome analysis, we identified 13 up- and 14 down-regulated glycoproteins in DN plasma. Among the up-regulated glycoproteins, the levels of lumican, vasorin and retinol binding protein-4 were verified by Western blot analysis of individual plasma samples.Conclusion and clinical relevance. Collectively, our findings show that biomarker discovery has considerable potential for predicting diabetic nephropathy in diabetic patients.