Proteomic Analysis of Synovial Fluid From the Osteoarthritic Knee: Comparison With Transcriptome Analyses of Joint Tissues

Abstract

Objective The pathophysiology of the most common joint disease, osteoarthritis (OA), remains poorly understood. Since synovial fluid (SF) bathes joint cartilage and synovium, we reasoned that a comparative analysis of its protein constituents in health and OA could identify pathways involved in joint damage. A proteomic analysis of knee SF from OA patients and control subjects was performed and compared to microarray expression data from cartilage and synovium. Methods Age-matched knee SF samples from control subjects, and patients with early- and late-stage OA (n=10 per group) were compared using two-dimensional differential gel electrophoresis (2D-DIGE) and mass spectrometry (MS). A MS with multiplexed peptide selected reaction monitoring (SRM) assay was used to confirm differential expression of a subset of proteins in an independent OA patient cohort. Proteomic results were analyzed by Ingenuity pathway analysis and compared to published synovial tissue and cartilage mRNA profiles. Results 66 proteins were differentially present in healthy and OA SF. Three major pathways were identified among these proteins: the acute phase response, and the complement and coagulation pathways. Differential expression of 5 proteins was confirmed by SRM assay. A focused analysis of transcripts corresponding to the differentially present proteins indicates that both synovial and cartilage tissues may contribute to the OA SF proteome. Conclusion Proteins involved in the acute phase response, complement and coagulation pathways are differentially regulated in SF of OA patients suggesting they contribute to joint damage. Validation of these pathways and their utility as biomarkers or therapeutic targets in OA is warranted.

Recent advances in the understanding of molecular mechanisms of cartilage degeneration, synovitis and subchondral bone changes in osteoarthritis

“…The deposition and activation of increased formation of osteoclasts in joints (115) complement factors in OA cartilage have been documented in several studies in OA patients as well as in animal models of OA (94,95). Tarkowski et al observed expression of decay accelerating factor (DAF) in the synovial lining cell layer in OA along with C5b-9 terminal complement complex suggesting activation of a complement-mediated response (96,97).…”

Immune Modulation to Improve Tissue Engineering Outcomes for Cartilage Repair in the Osteoarthritic Joint

“…Proteomic studies identified differential expression of complement proteins in human OA synovial fluid compared to control, further suggesting a potential role of these proteins in catabolic processes associated with OA. 69 …”

Biomarkers and proteomic analysis of osteoarthritis

“…Ritter and Gobezie further improved the sensitivity of their method by using two-dimensional gel electrophoresis and a more sensitive LC-MS/MS system. They identified 66 proteins that were differentially present in healthy and OA synovial fluid (Ritter et al, 2013). Another study modified the immunodepletion step to yield additional insights into the synovial fluid proteome ; employing nano-LC-MALDI-TOF/TOF analysis, and after excluding the abundant proteins reported in the previous study, they identified a total of 136 proteins involved in the formation and remodeling of the extracellular matrix, including fibronectin, kininogen-1, cartilage acidic protein 1, and COMP.…”

“…Besides discovery proteomics, targeted proteomic analysis has been applied to the study of synovial fluid. Selected Reaction Monitoring (SRM) was applied in a study to validate differential protein expression among OA and control synovial fluids (Ritter et al, 2013). Balakrishnan et al employed Multiple Reaction Monitoring (MRM), the advanced format of SRM, to validate the expression of ANPEP, osteoglycin (OGN), and dickkopf WNT signaling pathway inhibitor 3 (DKK3).…”