燃煤污染型砷中毒患者血清差异蛋白的筛选

Identification of differential proteins of serum in the patients suffering from coal-burning arsenism

目的应用血清蛋白组学技术，对比分析燃煤污染型砷中毒患者与正常人血清中差异蛋白的表达，筛选与燃煤污染型砷中毒发生发展密切相关的蛋白质。方法2008年收集经临床诊断的贵州燃煤污染型砷中毒患者和健康人空腹血清标本各6例，用双向凝胶电泳（2-DE）分离砷中毒患者血清和健康人血清的总蛋白质，银染显色后经图像分析识别差异表达的蛋白质，再应用基质辅助激光解析电离飞行时间质谱（MALDI-TOF-MS）鉴定差异表达的蛋白质点。结果两组血清2-DE图谱平均点数分别为779±35和865±30，匹配率为90．1％。筛选出两组间的差异蛋白点60个，砷中毒组表达上调25个，表达下调35个。对其中表达量差异3倍以上的35个蛋白点进行MALDI-TOF-MS分析，鉴定出包括角蛋白10、载脂蛋白A-V、转铁蛋白、α1抗胰蛋白酶、锌α2糖蛋白、细胞外信号调节激酶3、空泡分选蛋白33、O-GlcNAc糖基转移酶等13个蛋白质，其中5个蛋白质在砷中毒组表达上调，8个蛋白质在砷中毒组表达下调。结论本研究建立了分辨率高、重复性较好的燃煤污染型砷中毒患者血清双向凝胶电泳图谱，识别并鉴定了一些与健康人血清差异表达的蛋白质，为进一步探讨地方性砷中毒发病机制，筛选燃煤污染型砷中毒早期特异性血清分子标志物奠定了初步基础。

Objective To compare and analyze the differential expression of proteins between coalburning amenism serum and normal human serum and identify the proteins related with arseniasis caused by coal-burning. Methods Serum samples were collected from 6 normal subjects and 6 patients suffering from coal-burning arsenism. 2-DE was performed to separate serum proteins. After silver staining, the differential expression of proteins was analyzed and then identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry （MALDI-TOF-MS）. Results There were an average of 779 ± 35 spots and 865± 30 spots on 2-DE matching of two groups and the matching rate was 90. 1% between two groups. From thees two groups, 60 different protein spots were identified. Up-regulated expression was observed in 25 proteins and down-regulated expression in 35 proteins in the patient serum group. Among which 35 with differential expression above three times were singled out and MALDI-TOF-MS analysis was carried out on them. Thirteen proteins were identified, including keratin 10, apolipoprotein A-V, transferrin, alpha-l-antitrypsin, human zine-alpha-2-glyeoprotein, mitogen-activated protein kinase 3, vacuolar protein sorting 33A, O-linked GlcNAc transferase and etc. Up-regulated expression was observed in 5 proteins and down-regulated expression in 8 proteins in the patient serum group. Conclusion The well-resolved and reproducible 2-DE serum patterns of patients suffering from coal-burning arsenism were established and some differentially expressed proteins characterized. These data will be used to screen the biomarker and to further study arseniasis caused by coal-burning.