ISSN 1006-298X      CN 32-1425/R

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肾脏病与透析肾移植杂志 ›› 2014, Vol. 23 ›› Issue (2): 140-145.

• 论文 • 上一篇    下一篇

肾小球疾病患者尿液比较蛋白质组学研究方法的建立

  

  • 出版日期:2014-04-28 发布日期:2014-04-28

Establishment of comparative urine proteomics research strategy in patients with glomerulonephritis by iTRAQ technique.

  • Online:2014-04-28 Published:2014-04-28

摘要:

摘要 目的:采用蛋白质组学技术建立蛋白尿患者尿液蛋白质图谱分析方法,并通过对膜性肾病(MN)和局灶节段肾小球硬化(FSGS)患者尿液蛋白质图谱的比较分析以期寻找到具有鉴别诊断意义的尿液蛋白质标志物。方法:表现为肾病综合征的MN和FSGS患者各15例,采用白蛋白/IgG抗体清除联合ITRAQ(isobaric tags for relative and absolute quantitation)标记2D-LC MS/MS( Two dimensional liquid chromatography-tandem mass spectrometry)蛋白质组学技术对患者尿液蛋白质组进行全面分析并获取差异表达蛋白。对鉴定蛋白和差异蛋白进行GO(Gene ontology)分析。对鉴定蛋白丰度排序前10位蛋白与血蛋白质组丰度排序前10位蛋白进行比较。结果:共鉴定尿液蛋白809个, GO分析显示根据分子功能,鉴定蛋白主要属于催化活性、结合、受体活性三个类别;根据生物过程,鉴定蛋白主要属于代谢过程、细胞过程、发育过程三个类别;根据细胞定位,鉴定蛋白主要属于细胞外区域,细胞成分,细胞外基质三个类别。符合条件的差异蛋白16个,生物学功能包括参与纤溶调节、免疫反应、代谢过程、细胞增殖等。对表现为肾病综合征MN和FSGS患者尿液鉴定蛋白丰度排序前10位蛋白与血蛋白质组丰度排序前10位蛋白进行比较显示有较大差异,提示即使在病理情况下,肾脏可能仍对不同种类的蛋白保持不同的处理功能,蛋白尿可能有着不同于血蛋白质组的组成架构。结论:从患者尿液中获得的蛋白质组学图谱为进一步开展基于蛋白质组学生物标志物的研究提供了基础,从MN和FSGS患者尿液中获得的差异蛋白有待进一步验证其与疾病诊断,治疗及预后的关系。

关键词: :蛋白组学,  膜性肾病,   局灶节段性肾小球硬化, 同位素相对标记与绝对定量计数,  标志物

Abstract:

ABSTRACT Objective: To profile comprehensively urine proteomics and screen differentially expressed urine protein in membranous nephropathy (MN) and focal segmental glomerulosclerosis (FSGS) patients with nephrotic syndrome. Methodology: Fifteen MN patients and fifteen FSGS patients with nephrotic syndrome were enrolled into this study. Their urine samples were analyzed by Albumin/IgG antibody depletion combined with iTRAQ (isobaric tags for relative and absolute quantitation) labeled 2D-LC MS/MS(Two dimensional liquid chromatography-tandem mass spectrometry) for profiling urine proteomics and screening differentially expressed urine proteins. GO(Gene ontology)analysis was used to categorize identified protein and differentially expressed proteins according to molecular function、biological process and cellular component. The top 10 proteins in identified protein were compared with the top 10 proteins in plasma proteomics. Results: A total of 809 urine proteins were identified in MN and FSGS patients with nephrotic syndrome, of which 16 were differentially expressed between the two groups of subjects. GO analysis revealed that the molecular function of identified protein mainly included catalytic activity、binding and receptor activity, the biological process mainly involved metabolic process、cellular process and development process, the cellular component mainly contained extracellular region、cell part and extracellular matrix. The biological process of differentially expressed proteins involved fibrinolysis regulation、immune reaction、metabolic process、cell proliferation and so on. The top 10 proteins in urine proteomics from patients with nephrotic syndrome of MN and FSGS were compared with the top 10 proteins in plasma proteomics, which suggested even in pathological condition, the kidney may keep handling function for different protein, and proteinuria proteomics may has a different arctecture with plasma proteomics. Conclusion: Our proteinuria proteomics profile was the largest for now and provided a basis for future study. Our differentially expressed protein need to be validated as biomarkers for diagnosis, therapy and prognosis.

Key words: proteomics, MN,  FSGS, iTRAQ, biomarker