Micheroli, R. | bioRxiv Science

bioRxiv2022

A comprehensive single-cell atlas of freshly dissociated human synovium in inflammatory arthritis with an optimized dissociation protocol for prospective fresh synovial biopsy collection

Single-cell RNA-sequencing is advancing our understanding of synovial pathobiology in inflammatory arthritis. Here, we optimized the protocol for the dissociation of fresh synovial biopsies and created a reference single-cell map of fresh human synovium in inflammatory arthritis. We utilized the published method for dissociating cryopreserved synovium and optimized it for dissociating small fresh synovial biopsies. The optimized protocol enabled the isolation of a good yield of consistently highly viable cells, minimizing the dropout rate of prospectively collected biopsies. Our reference synovium map comprised over 100000 unsorted single-cell profiles from 25 synovial tissues of patients with inflammatory arthritis. Synovial cells formed 11 lymphoid, 15 myeloid and 16 stromal cell clusters, including IFITM2+ synovial neutrophils. Using this reference map, we successfully annotated published synovial scRNA-seq datasets. Our dataset uncovered endothelial cell diversity and identified SOD2highSAA1+SAA2+ and SERPINE1+COL5A3+ fibroblast clusters, expressing genes linked to cartilage breakdown (SDC4) and extracellular matrix remodelling (LOXL2, TGFBI, TGFB1), respectively. We broadened the characterization of tissue resident FOLR2+COLEC12high and LYVE1+SLC40A1+ macrophages, inferring their extracellular matrix sensing and iron recycling activities. Our research brings an efficient synovium dissociation protocol and a reference annotation resource of fresh human synovium, while expanding the knowledge about synovial cell diversity in inflammatory arthritis.

genomics↗

bioRxiv2021

MAFB surrogates the glucocorticoid receptor ability to induce tolerogenesis in dendritic cells

Glucocorticoids (GCs) exert potent anti-inflammatory effects in immune cells through the glucocorticoid receptor (GR). Dendritic cells (DCs), central actors for coordinating immune responses, acquire tolerogenic properties in response to GCs. Tolerogenic DCs (tolDCs) have emerged as a potential treatment for various inflammatory diseases. To date, the underlying cell type-specific regulatory mechanisms orchestrating GC-mediated acquisition of immunosuppressive properties remain poorly understood. In this study, we investigated the transcriptomic and epigenomic remodeling associated with differentiation to DCs in the presence of GCs. Our analysis demonstrates a major role of MAFB in this process, in synergy with GR. GR and MAFB both interact with methylcytosine dioxygenase TET2 and bind to genomic loci that undergo specific demethylation in tolDCs. We also show that the role of MAFB is more extensive, binding to thousands of genomic loci in tolDCs. Finally, MAFB knockdown erases the tolerogenic properties of tolDCs and reverts the specific DNA demethylation and gene upregulation. The preeminent role of MAFB is also demonstrated in vivo for myeloid cells from synovium in rheumatoid arthritis following GC treatment. Our results imply that, once directly activated by GR, MAFB takes over the main roles to orchestrate the epigenomic and transcriptomic remodeling that define the tolerogenic phenotype.

immunology↗