Synovial fluid CD69+CD8+ T cells with tissue-resident phenotype mediate perforin-dependent citrullination in rheumatoid arthritis

Abstract

Objectives: Although the importance of tissue-resident memory T (TRM) cells in organ-specific chronic inflammation has been recognised, little is known about their role in rheumatoid arthritis (RA). Here, we examined the characteristics of synovial fluid CD8+ T cells that express canonical TRM markers CD69 and CD103, and their role in the pathogenesis of RA. Methods: Synovial fluid mononuclear cells (SFMCs) were obtained from patients with RA. Flow cytometric analysis of surface markers and cytotoxic molecules of CD8+ T cells was performed. TCR repertoire of CD8+ T cells was analysed by TCRVβ CDR3 sequencing. Citrullination with the formation of neutrophil extracellular trap (NET) was evaluated by immunofluorescence staining. Results: The frequency of CD8+ T cells was increased in SFMCs, and these CD8+ T cells were primarily comprised of CD45RA- memory T cells expressing CD69 and/or CD103. CD69+CD8+ T cells exhibited TRM phenotypes, including upregulation of CXCR6, CD49a and CD101, and downregulation of S1PR1 and KLF2. TCR repertoire analysis showed that these cells were an oligoclonally expanded population with increased expression of cytotoxic molecules. The treatment of neutrophils with supernatant from IL-15-stimulated CD69+CD8+ T cells induced perforin-mediated histone citrullination and NET formation irrespective of their CD103 expression. The frequency of perforin-expressing cells among CD69+CD8+ T cells in SFMCs was significantly higher in patients with anti-citrullinated protein antibody (ACPA) than in those without ACPA. Conclusion: CD69+CD8+ T cells in the SFMCs of RA patients exhibit TRM-like features. These cells may participate in the pathogenesis of RA via perforin-mediated citrullination.