Identification of psoriatic arthritis mediators in synovial fluid by quantitative mass spectrometry

Daniela Cretu, Ioannis Prassas, Punit Saraon, Ihor Batruch, Rajiv Gandhi, Eleftherios P Diamandis, Vinod Chandran, Daniela Cretu, Ioannis Prassas, Punit Saraon, Ihor Batruch, Rajiv Gandhi, Eleftherios P Diamandis, Vinod Chandran

Abstract

Background: Synovial fluid (SF) is a dynamic reservoir for proteins originating from the synovial membrane, cartilage, and plasma, and may therefore reflect the pathophysiological conditions that give rise to arthritis. Our goal was to identify and quantify protein mediators of psoriatic arthritis (PsA) in SF.

Methods: Age and gender-matched pooled SF samples from 10 PsA and 10 controls [early osteoarthritis (OA)], were subjected to label-free quantitative proteomics using liquid chromatography coupled to mass spectrometry (LC-MS/MS), to identify differentially expressed proteins based on the ratios of the extracted ion current of each protein between the two groups. Pathway analysis and public database searches were conducted to ensure these proteins held relevance to PsA. Multiplexed selected reaction monitoring (SRM) assays were then utilized to confirm the elevated proteins in the discovery samples and in an independent set of samples from patients with PsA and controls.

Results: We determined that 137 proteins were differentially expressed between PsA and control SF, and 44 were upregulated. The pathways associated with these proteins were acute-phase response signalling, granulocyte adhesion and diapedesis, and production of nitric oxide and reactive oxygen species in macrophages. The expression of 12 proteins was subsequently quantified using SRM assays.

Conclusions: Our in-depth proteomic analysis of the PSA SF proteome identified 12 proteins which were significantly elevated in PsA SF compared to early OA SF. These proteins may be linked to the pathogenesis of PsA, as well serve as putative biomarkers and/or therapeutic targets for this disease.

Keywords: Early osteoarthritis; Mass spectrometry; Mediators; Proteins; Proteomics; Psoriatic arthritis; Selected reaction monitoring assays; Synovial fluid.

Figures

Figure 1
Figure 1
Verification of elevated proteins in PsA synovial fluid (Set I) by selected reaction monitoring assays, normalized against housekeeping proteins. Symbols represent SF samples from individual subjects; horizontal lines depict the mean, and vertical lines the standard deviation. ****Indicates P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05; ns: non-significant.
Figure 2
Figure 2
Verification and concentration of elevated proteins in PsA synovial fluid (Set II) by selected reaction monitoring assays, normalized against heavy-labeled peptides. Symbols represent SF samples from individual subjects; horizontal lines depict the mean, and vertical lines the standard deviation. ****Indicates P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05; ns: non-significant.
Figure 3
Figure 3
Summary of the experimental design.

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