Profiling of endogenous peptides in human synovial fluid by NanoLC-MS: Method validation and peptide identification
Profiling of endogenous peptides in human synovial fluid by NanoLC-MS: Method validation and peptide identification
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DOI：
10.1021/pr0704534
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发表时间：
2007-11-01
影响因子：
4.4
通讯作者：
Hankemeier, Thomas
中科院分区：
文献类型：
--
作者：
Kamphorst, Jurre J.;van der Heijden, Rob;Hankemeier, Thomas
滑液可能含有退行性关节疾病（如骨关节炎）早期诊断和疾病进展的标志物。在这里，描述了一种方法，用于分析内源性肽在人类滑液中，使用超滤，固相萃取，纳米级液相色谱法，和高分辨率质谱。滑液的特征在于其高粘度，由润滑剂透明质酸的存在引起。该方法被证明能够消除高浓度的透明质酸，这似乎是获得满意的分析性能所必需的，即5- 15%的日内相对标准偏差，6- 16%的日间相对标准偏差，R-2 = 0.994的线性响应，飞摩尔范围内的检测限，回收率为14- 67%。利用开发的方法，在来自骨关节炎患者和健康对照的滑液样品中，总共鉴定了来自40种蛋白质的501种肽。在早期研究中与骨关节炎相关的六种蛋白质（胶原蛋白11、蛋白半胱氨酸蛋白聚糖4、血清淀粉样蛋白A、微管蛋白、波形蛋白和基质Gla）的肽裂解产物也可以用我们的分析方法鉴定。该方法的稳健性表明，它可以应用于系统生物学方法，以进一步研究退行性关节疾病，最终导致更好地了解疾病及其治疗，以及开发新的生物标志物来监测这些过程。
Synovial fluid potentially contains markers for early diagnosis and disease progression in degenerative joint diseases such as osteoarthritis. Here, a method is described for profiling endogenous peptides in human synovial fluid, using ultrafiltration, solid-phase extraction, nanoscale liquid chromatography, and high-resolution mass spectrometry. Synovial fluid is characterized by its high viscosity, caused by the presence of the lubricant hyaluronic acid. The method proved to be capable of eliminating the high concentrations of hyaluronic acid, which appeared to be necessary to obtain satisfactory analytical performance, that is, within-day relative standard deviations of 5-15%, between-day relative standard deviations of 6-16%, a linear response of R-2 = 0.994, a limit of detection in the femtomole range, and reproducible recoveries of 14-67%. With the developed method, in a synovial fluid sample from an osteoarthritis patient and a healthy control, in total, 501 peptides originating from 40 proteins were identified. Peptide cleavage products from six proteins that have been associated with osteoarthritis in earlier studies (collagen 11, proteoglcycan 4, serum amyloid A, tubulin, vimentin, and Matrix Gla) could also be identified with our profiling method. The robustness of the method indicates that it can be applied in systems biology approaches for further studies on degenerative joint disease, eventually leading to a better understanding of the disease and its therapy, as well as the development of novel biomarkers to monitor these processes.