title: Profiling of endogenous peptides in human synovial fluid by NanoLC-MS: method validation and peptide identification.
abstract: 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 R2=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 II, 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.
authors.affiliation: Division of Analytical Biosciences, LACDR, Leiden University, P.O. Box 9502, 2300 RA Leiden, the Netherlands.
authors.forename: Jurre J
authors.initials: JJ
authors.lastname: Kamphorst
authors.forename: Rob
authors.initials: R
authors.lastname: van der Heijden
authors.forename: Jeroen
authors.initials: J
authors.lastname: DeGroot
authors.forename: Floris P J G
authors.initials: FP
authors.lastname: Lafeber
authors.forename: Theo H
authors.initials: TH
authors.lastname: Reijmers
authors.forename: Benno
authors.initials: B
authors.lastname: van El
authors.forename: Ubbo R
authors.initials: UR
authors.lastname: Tjaden
authors.forename: Jan
authors.initials: J
authors.lastname: van der Greef
authors.forename: Thomas
authors.initials: T
authors.lastname: Hankemeier
mesh_terms: D000595:Amino Acid Sequence; D001172:Arthritis, Rheumatoid / Q000378:metabolism; D015415:Biomarkers / Q000737:chemistry; D002853:Chromatography, Liquid / Q000379:methods*; D020869:Gene Expression Profiling; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D010003:Osteoarthritis / Q000175:diagnosis / Q000378:metabolism; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D013582:Synovial Fluid / Q000378:metabolism*
chemical_list: D015415:Biomarkers; D010455:Peptides; D011506:Proteins