应用同位素标记相对和绝对定量蛋白质组学方法筛选骨关节炎患者关节液差异蛋白

Isobaric tag for relative and absolute quantitation-based proteomics analysis of synovial fluid from osteoarthritic knee

目的应用同位素标记相对和绝对定量（iTRAQ）联合液相串联质谱技术分析骨关节炎患者关节液中差异蛋白，寻找潜在的骨关节炎生物标记物。方法30例患者依据K-L评分分为对照组（K-L评分0级），早期OA患者（K-L评分Ⅱ级）、晚期OA患者（K-L评分Ⅳ级），每组10例。收集3组患者的膝关节液，经去除高丰度蛋白处理后行iTRAQ标记联合液相串联质谱技术检测，对检测结果进行Gene ontology、Pathway等分析，寻找3组之间明显差异蛋白。2组间比较采用t检验。结果本研究从关节液中共检测出1 283种蛋白，其中有268种蛋白为首次在关节液中发现。早期OA患者与对照组关节液中差异蛋白相比上调72种，下调249种；晚期OA患者与对照组关节液中差异蛋白相比上调128种，下调141种；晚期OA患者与早期OA患者关节液中差异蛋白相比上调128种，下调141种。3组之间明显差异且上调的蛋白共有8种。结论应用iTRAQ技术可以从关节液中检测出大量蛋白；通过对这些蛋白的深入分析，有助于了解OA的发病机制及发现新的OA生物标记物。

ObjectiveWe performed comprehensive proteomic analyses of synovial fluid by using the isobaric tags for relative and absolute quantitation （iTRAQ） method and LC-MS/MS, and searched for candidate biomarkers for osteoarthritis.MethodsSynovial fluid was collected from patients with late-stage OA, earlystage OA and the control group. Molecular variations were detected by the iTRAQ method. T test was used for statistical analysis.ResultsUsing the iTRAQ method, we identified 1 283 proteins from synovial fluid of patients with osteoarthritis, of which 268 proteins were not reported previously. There were 72 proteins upregulated and 249 proteins down-regulated between moderate OA group and controls. One hundred and twenty-eight proteins were up-regulated and 141 proteins were down-regulated between severe OA group and controls. One hundred and ninety-two proteins were up-regulated and 76 proteins were down-regulated between severe OA group and moderate OA group. Eight proteins were found to be up-regulated in the three groups. ConclusionThis is an in-depth analysis of the synovial fluid proteome from patients with osteoarthritis by iTRAQ method. The catalog of proteins generated in this study will further expand our knowledge regarding the pathophysiology of osteoarthritis and help us in identifying good biomarkers for early diagnosis.