Background/Purpose: Synovial tissue macrophages (STMs) represent a mixed population of cells contributing to the pathogenesis of rheumatoid arthritis (RA). We identified an enrichment of Secreted phosphoprotein-1 (SPP1) STMs in RA compared to healthy control (HC) synovial tissue. We show SPP1+ STMs are also enriched in individuals at risk for RA (IAR), suggesting they represent an early disease-specific myeloid subset (1). The aim of this study was to uncover the specific contributions of SPP1+ STMs to RA pathogenesis, which may provide opportunities to reinstate joint homeostasis.

Methods: Peripheral blood was obtained from HC and RA patients. CD14+ monocytes were isolated and differentiated either in the presence of M-CSF (50ng/ml) or Ca2+ (2.5 mM) for 7 days to derive Macs or SPP1-expressing-Macs (S-Macs), respectively. Inflammatory cytokine production by these cells, and their defining metabolic parameters were measured by RT-PCR, ELISA and by the Seahorse-XFE technology. Paraffin embedded synovial tissue biopsies were obtained from HC, from RA patients undergoing key-hole arthroscopy, and from seropositive IAR who have no clinical signs of inflammation. Spatial transcriptomics was performed using a custom Xenium 5K+ panel. Only RA patients who fulfilled the revised ACR criteria were included in this study.

Results: RA SPP1+ STMs correlate significantly with clinical inflammation and display the highest expression of glycolytic genes compared to all other STMs. RA S-Macs have increased expression of SPP1 at both the gene and protein level compared to Macs. S-Macs display increased expression of IL-6, TNF, IL-1b and IL-8 and the glycolytic genes HIF1a and HK2 compared to Macs. Importantly, this is more pronounced in cells derived from RA vs HC indicating priming in the RA periphery. RA S-Macs display decreased baseline oxidative phosphorylation and maximal respiratory capacity compared to Macs derived from HC/RA blood, while concomitantly skewing the ECAR/OCR ratio in favor of glycolysis, indicating that RA S-Macs rely heavily on glycolysis. Finally, we conducted Xenium in situ spatial profiling technology to measure RNA expression levels of 5100 genes in unique paraffin-embedded synovial tissue from HC, IAR and RA. We annotated cell clusters (including macrophages, fibroblasts, T, B cells) and identified their localization to serve as a map of the synovial cellular organization across the evolution of disease.

Conclusion: We demonstrate that SPP1+ STMs correlate with clinical inflammation in RA and are metabolically re-programmed towards glycolysis, leading to sustained induction of pro-inflammatory responses. Xenium spatial analysis allows us to determine the cellular interactions and disease stage-specific factors that shape SPP1+ STMs in RA synovium. These cells may thus represent a potential pathway for therapeutic targeting.(1) Hanlon, Megan M et al. “Loss of synovial tissue macrophage homeostasis precedes rheumatoid arthritis clinical onset.” Sci Adv. 2024;10(39):eadj1252. doi:10.1126/sciadv.adj1252

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/pathogenic-role-of-spp1-macrophages-in-rheumatoid-arthritis/