Background/Purpose:

Osteoarthritis is the most common joint disease and involves whole joint, including degeneration of articular cartilage and surrounding tissues. Although widely prevalent, the exact mechanisms involved in pathogenesis of OA are not well understood. Increasing body of data suggest role of low-grade, subclinical, in pathogenesis of OA. However the role of immune cell subsets in OA is poorly investigated. In present study we have applied a combination of single cell mass cytometry for analysis of the peripheral immunome and next generation RNA-sequencing on cell pellet from knee joint fluid to characterise immune cells and pathways.

Methods:

Patients samples were taken from a randomized controlled trial on treatment of symptomatic knee OA using colchicine for 16 weeks (1)(2). Patients were grouped as having ≥30% improvement in WOMAC (improvement group) and improvement of WOMAC not reaching 30%(Worsening group) at study end regardless of treatment arms. For RNA sequencing analysis 16 paired patient samples, from baseline and study end, were used. For mass cytometry analysis, 37 marker panel was designed to aid in identification of major peripheral blood mononuclear cell (PBMCs) subsets. Unsupervised clustering of mass cytometry data was performed using in-house developed analysis software MARVIS. This software combines dimension reduction and clustering steps to identify all possible cellular subsets from high dimension data. Further, custom R scripts helped in identifying nodes that were differentially expressed between the study groups and also phenotype of these nodes.

Results:

Differential gene expression analysis comparing baseline vs week 16 from worsening group revealed changes in 696 genes and in improvement group 89 genes were identified. Estimation of cell type abundances, using Cibersort algorithm, showed increased frequency of M2 macrophages, follicular helper T cells and activated NK cells in worsening group and increased memory CD4 T cells in improvement group. Mass cytometry analysis of PBMCs showed increase in inflammatory Tbet+ ILC subsets in worsening group (n=6) and an increase in Th2 like GATA3+ CD4 T cells in improvement group (n=6) at baseline.

Conclusion:

Our systematic multi dimensional approach reveals differences in the alteration of immune cells in knee OA patients with/ without symptoms improvement, both at synovial fluid level and systemic blood level. Further analysis may provide insights into mechanistic pathways involved in pathogenesis of OA, the interaction between tissue and systemic circulation and for advent of novel OA therapies.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/microenvironmental-and-systematic-evaluation-of-the-immunome-in-osteoarthritis/