OP0028 CD206+CD163+ PATHOGENIC MACROPHAGES ENRICHED IN RHEUMATOID ARTHRITIS SYNOVIAL TISSUE WITH DISTINCT TRANSCRIPTIONAL SIGNATURES
Authors: M. Hanlon, M. Canavan, Q. Song, N. Neto, P. Gallagher, R. Mullan, C. Hurson, M. Monaghan, S. Nagpal, D. Veale, U. Fearon
Journal: Annals of the Rheumatic Diseases (2021)

Background: Synovial tissue macrophages are an exquisitely plastic pool of innate cells that play a key role in RA disease progression. However, the precise nature, diversity, and function of macrophage subsets within the inflamed joint remains unexplored. Objectives: Therefore, the aims of this study are to phenotypically, transcriptionally and functionally characterise synovial tissue macrophages residing within the inflamed joint. Methods: Rheumatoid Arthritis, Psoriatic Arthritis, Osteoarthritis and healthy control synovial-tissue biopsies and synovial-fluid mononuclear cells were analysed using the following panel (CD40,-CD45,-CD64,-CD68,-CD163,-CD206,-CD253,-CCR4,-CCR7,-CXCR1,-CXCR3). CD206+CD163+ and CD206-CD163- macrophages were sorted from RA synovial-tissue by FACSAria sorter; RNAseq and FLIM analysis, autologous T-cell co-culture and heathy fibroblast experiments performed. Cytokine expression was measured by MSD immunoassay. Results: RA synovial tissue and fluid macrophages display markers typical of both M1 (CD40+CD253+) and M2 (CD206+CD163+) macrophages with a spectrum of macrophage activation states identified. Within this spectrum, significant enrichment of dominant CD206+CD163+ macrophage-subtype is present in synovial tissue versus fluid (p&lt;0.05). CD206+CD163+ synovial tissue macrophages express significantly more CD40 than synovial fluid (p&lt;0.0003), positively correlate with disease activity (r=0.6, p&lt;0.01), with baseline levels predicting response to therapy (p&lt;0.05). Moreover, CD206+CD163+CD40+ macrophages are enriched in RA synovial tissue compared to PsA and OA pathotypes (p&lt;0.05). While the CD206+CD163+ subset is present in healthy synovial tissue, expression of CD40 is completely absent in healthy synovium (p&lt;0.05) with dramatically decreased expression of CX3CR1 on RA macrophages. RNA-seq analysis indicates that CD206+CD163+ population is transcriptionally distinct from synovial tissue CD206-CD163-, synovial fluid CD206+CD163+, and RA monocyte-derived M1/M2 macrophages, with unique tissue-resident gene signatures. Moreover, differing metabolic demands between CD206+CD163+ and CD206-CD163- subsets was demonstrated by RNAseq and FLIM analysis. CD206+CD163+ macrophages enhance autologous T-cell responses, spontaneously secrete high levels of pro-inflammatory cytokines and activate healthy fibroblasts towards pro-inflammatory mechanisms thus further contributing to the local inflammatory response. Finally, inhibition of CD40 activity abrogates the expression of pro-inflammatory mediators (TNFa, IL-1B, IL-6, IFNy) and induces IL-10 expression in sorted CD206+CD163+ synovial tissue-macrophages suggesting a key role for CD40 in driving this pathogenic phenotype. Conclusion: This data identifies for the first-time enrichment of a previously undescribed dysfunctional dominant and transcriptionally distinct macrophage subtype in RA synovial tissue. Taken together, this data provides a greater understanding of the critical role tissue-resident macrophages play in perpetuating inflammation in RA. Further investigation of the molecular patterns and cues that shape specific synovial macrophage subsets may provide opportunities to reinstate RA joint homeostasis. Disclosure of Interests: Megan Hanlon: None declared, Mary Canavan: None declared, Qingxuan Song Employee of: Janssen Research &amp; Development, Nuno Neto: None declared, Phil Gallagher: None declared, Ronan Mullan: None declared, Conor Hurson: None declared, Michael Monaghan: None declared, Sunil Nagpal Employee of: Janssen Research &amp; Development, Douglas Veale Speakers bureau: Abbvie, Janssen, Novartis, MSD, Pfizer, UCB, Consultant of: Abbvie, Janssen, Novartis, MSD, Pfizer, UCB, Grant/research support from: Janssen, Abbvie, Pfizer, UCB, Ursula Fearon Speakers bureau: Abbvie, Grant/research support from: Janssen, Abbvie, Pfizer, UCB