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Background: Macrophages are believed to be local and systemic master actors in disease, critically involved in shaping inflammation, and in disease resolution via effective efferocytosis. Impaired efferocytosis, a hallmark of failed inflammation resolution, has been reported in an increasing number of immune-mediated inflammatory diseases, like Rheumatoid Arthritis (RA). Macrophage plasticity and capability for reprogramming, make them an attractive target for novel disease-modifying therapies. While cell-based therapies currently being scrutinized for their unique potential to fulfil a lost function, including macrophage-based therapies, the use of resolution-type macrophage secretome, i.e. the molecules they release, might provide a simpler and yet potentially safer approach to restore homeostasis in RA. Objectives: The present study attempted to understand the inflammatory profile and functionality of RA patient's monocyte-derived macrophages and evaluate their plasticity and capability for reprogramming to resolution-type macrophages as a source of RA patient-derived secretome with therapeutic potential. Methods: Patients being followed up at CHU Besançon between 2015 and 2021 were included in this prospective single center study. Inclusion criteria were 18 to 80 year-old patients with RA according to ACR/EULAR 2010 criteria, with a DAS28 ≥ 2.6, with or without csDMARDS (methotrexate, leflunomide or sulfasalazine), and naïve of biological agents or systemic corticosteroids for 6 months. Blood inflammatory cytokines and lipid mediators were quantified using CBA multiplex and MS analysis, respectively. Blood monocytes from RA patients or healthy donors (HD) were isolated by density gradient, and examined after differentiation into macrophages using M-CSF during 7 days, or not. Inflammatory responses and efferocytosis capacities were analyzed using flow cytometry. Results: A total of 28 patients and 31 HD were included. Evaluating inflammatory mediators, we found pro-inflammatory cytokines TNF-α, IL-6, IL-8, IL-1β and IL-12 increased in RA patient's plasma compared to HD. We also detected a global increase of the pro-inflammatory lipid mediators derived from omega-3 fatty acid in patient plasma, confirming the ongoing systemic inflammatory nature of disease in RA patients. Furthermore, we observed that monocytes and dendritic cells (DC) of RA patients presented higher levels of the co-stimulatory marker CD40, as compared to HD, which was not true on plasmacytoid DC. Thus, we confirm that circulating myeloid cells exhibit an inflammatory profile in RA patients. To evaluate how inflammatory pattern affects the efferocytosis capacities of macrophages, monocytes were differentiated into macrophages and evaluated for their phagocytosis of apoptotic cells. Interestingly, we observed that the efferocytic capacities of patient monocytes-derived macrophages were not impaired ex vivo. This was confirmed by similar expression of membrane engulfment receptors at the macrophage stage. Culturing macrophages with apoptotic cells generate resolution-type macrophages secreting pro-resolutive mediators. Interestingly, when exposed to HD resolution mediators, the efferocytic capacity of RA macrophages could be further increased by 67% 5-303%, min-max. Our data show that in RA patients, monocytes remained plastic and capable to reprogram into resolution-type macrophages outside of patient inflammatory environment, and with that, could be considered not only a potential target but also a cellular source to generate autologous patient-specific resolution mediator secretome. Conclusion: Our study revealed that despite their inflammatory profile, RA patient monocytes remained plastic and conserved ex vivo their efferocytic capacities, and used to produce patient-specific autologous resolution-type secretome. Further investigations are ongoing to confirm the therapeutic properties of RA patient own macrophage-derived secretomes to be proposed as next generation disease modifying therapeutic modality/bDMARD for RA patients. REFERENCES: NIL. Acknowledgements: NIL. Disclosure of Interests: Mélanie COUTURIER MED'INN'Pharma, F-25000 Besançon, France, MED'INN'Pharma, F-25000 Besançon, France, Emilie GAIFFE: None declared, Charline VAUCHY: None declared, Susanne BEHLKE MED'INN'Pharma, F-25000 Besançon, France, Eric Toussirot: None declared, Sylvain PERRUCHE MED'INN'Pharma, F-25000 Besançon, France, MED'INN'Pharma, F-25000 Besançon, France.
Couturier et al. (Sat,) studied this question.