室间隔缺损生物标志物血清蛋白质组学分析

Proteomics measures as potential serum protein biomarkers for ventricular septal defect

目的应用同位素标记相对和绝对定量(iTRAQ)技术结合液相色谱串联基质辅助激光解析电离飞行时间质谱(LC-MALDI-TOF/TOF MS)方法筛选潜在的室间隔缺损血清标志蛋白,探索室间隔缺损的发病机制。方法收集室间隔缺损患者和与之相匹配的健康对照组、房间隔缺损患者血清各20例,组内等量混合后利用安捷伦公司生产的多重亲和去除系统(MARS)去除14种高丰度蛋白,iTRAQ试剂标记后应用二维液相色谱分离,经质谱鉴定进行相对定量和生物信息学分析。结果质谱鉴定出置信度>95%的蛋白质共329种,其中表达量有明显差异的蛋白质36种包括富含半胱氨酸的酸性分泌蛋白、脂联素等。室间隔缺损组较健康对照组和房间隔缺损组表达量均上调>1.2倍的蛋白有21种,下调<0.83倍的蛋白有16种。结论筛选出一系列室间隔缺损相关的差异性蛋白,为进一步探索室间隔缺损发病机制奠定基础。

Objective To detect differentially expressed proteins related with ventricular septal defect（VSD） by utilizing isobaric tags for relative and absolute quantitation（iTRAQ） labeling coupled with liquid chromatography and matrix-assisted laser desorption ionization tandem time of flight mass spectrometry（LC-MALDI-TOF/TOF MS） technology for the screening of potential serum protein biomarkers of VSD and to explore the pathogenesis of VSD.Methods The serum samples were collected from 20 children with VSD,20 healthy children and 20 children with atrial septal defect（ASD） matched to the VSD cases in age,gender and race.Multiple affinity removal system（MARS） was adopted to exclude 14 kinds of high abundant proteins in the serum.Proteins differentially expressed in the serum of patients with VSD were identified with iTRAQ labeling coupled with LC-MALDI-TOF/TOF MS technology and analysed with bioinformatics tools and methods.Results A total of 329 proteins with the confidence coefficient above 95% were recognised,in which 36 differentially expressed proteins related with VSD,including secreted protein acidic rich in cysteine（SPARC） and adiponection,were identified.For the 38 proteins,21 proteins were over-expressed（1.2 fold） and 15 proteins were under-expressed（0.83 fold） in VSD group compared with those in both healthy group and ASD group.Conclusion A serial of differentially expressed proteins associated with VSD were identified in this pilot study.The results suggest that iTRAQ labeling combined with LC-MALDI-TOF/TOF MS technology is a feasible strategy for the screening of potential serum protein biomarkers of VSD and the study of VSD pathogenesis.