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Arthritis Rheumatol. 2019 Nov 8. doi: 10.1002/art.41161. [Epub ahead of print]

Multiparameter Analysis Identifies Heterogeneity in Knee Osteoarthritis Synovial Responses.

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Division of Rheumatology, University of Washington, Seattle, WA, 98019, USA.
Department of Pharmacology, University Medical Center of Johannes Gutenberg University, Mainz, 55101, Germany.
Benaroya Research Institute, Virginia Mason Medical Center, Seattle, WA, 98101, USA.
Department of Orthopedics and Sports Medicine, Virginia Mason Medical Center, Seattle, WA, 90101, USA.



Synovial membrane inflammation is common in osteoarthritis (OA) and increases cartilage injury. However, synovial fluid and histology studies suggest that OA inflammatory responses are not homogeneous. Greater understanding of these responses may provide new insights into OA disease mechanisms. Our objective was to develop a novel, multi-parameter approach to phenotype synovial responses in knee OA.


Cell composition and soluble protein production was measured by flow cytometry and multiplex ELISA in synovium collected from OA patients undergoing knee replacement surgery (n=35).


Testing disaggregation conditions showed that aggressive digestion improved synovial cell yield and mesenchymal staining by flow cytometry, but negatively impacted CD4+ T cell and CD56+ natural killer (NK) cell staining. Less aggressive digestion preserved these markers and showed highly variable T cell infiltration (range 0-43%, n=32). Correlation analysis identified mesenchymal subpopulations associated with different non-mesenchymal populations, including macrophages and T cells (CD45+CD11b+HLA-DR+ myeloid cells with podoplanin (PDPN)+CD73+CD90-CD34-mesenchymal cells, r=0.65 p<0.0001; CD45+CD3+T cells withPDPN+CD73+CD90+CD34+ mesenchymal cells, r=0.50 p=0.003). IL-6 measured by flow cytometry correlated strongly with IL-6 released by ex vivo culture of synovial tissue (r=0.59 p=0.0012) and was highest in mesenchymal cells co-expressing CD90 and CD34. IL-6, IL-8, complement factor D (CFD), and IL-10 release positively correlated with tissue cellularity (p=0.0042, 0.018, 0.0012, and 0.038, respectively). Additionally, increased CD8+ T cell numbers also correlated with retinol binding protein 4 (RBP4) (p=0.033). Finally, combining flow cytometry and multiplex data identified patient clusters with different types of inflammatory responses.


We used a novel approach to analyze OA synovium, identifying patient-specific inflammatory clusters. This study argues that phenotyping synovial inflammation may provide new insights into OA patient heterogeneity and biomarker development.


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