Taxonomy of fibroblasts and progenitors in the synovial joint at single-cell resolution

Abstract

ObjectivesFibroblasts in synovium include fibroblast-like synoviocytes (FLS) in the lining andThy1+ connective-tissue fibroblasts in the sublining. We aimed to investigate their developmental origin and relationship with adult progenitors.MethodsTo discriminate betweenGdf5-lineage cells deriving from the embryonic joint interzone and otherPdgfrα-expressing fibroblasts and progenitors, adultGdf5-Cre;Tom;Pdgfrα-H2BGFPmice were used and cartilage injury was induced to activate progenitors. Cells were isolated from knees, fibroblasts and progenitors were sorted by fluorescence-activated cell-sorting based on developmental origin, and analysed by single-cell RNA-sequencing. Flow cytometry and immunohistochemistry were used for validation. Clonal-lineage mapping was performed usingGdf5-Cre;Confettimice.ResultsIn steady state,Thy1+ sublining fibroblasts were of mixed ontogeny. In contrast,Thy1-Prg4+ lining fibroblasts predominantly derived from the embryonic joint interzone and includedPrg4-expressing progenitors distinct from molecularly defined FLS. Clonal-lineage tracing revealed compartmentalisation ofGdf5-lineage fibroblasts between lining and sublining. Following injury, lining hyperplasia resulted from proliferation and differentiation ofPrg4-expressing progenitors, with additional recruitment of non-Gdf5-lineage cells, into FLS. Consistent with this, a second population of proliferating cells, enriched near blood vessels in the sublining, supplied activated multipotent cells predicted to give rise toThy1+ fibroblasts, and to feed into the FLS differentiation trajectory. Transcriptional programmes regulating fibroblast differentiation trajectories were uncovered, identifying Sox5 and Foxo1 as key FLS transcription factors in mice and humans.ConclusionsOur findings blueprint a cell atlas of mouse synovial fibroblasts and progenitors in healthy and injured knees, and provide novel insights into the cellular and molecular principles governing the organisation and maintenance of adult synovial joints.

Molecular reframing of fibroblasts during resolution of arthritis

“…To avoid analytical bias due to unrelated exclusion of certain fibroblast subtypes 7 , we integrated 33 scRNA-seq datasets (unpublished and public, Supplemental Fig 1 a-e) from different arthritis models and their controls, resulting in to a total of 52,131 mesenchymal cells from joints, and performed unbiased graph-based clustering. Consistent with previous reports [8][9][10][11][12][13][14] , the integrated dataset confirmed a high heterogeneity of mesenchymal cells in joints. Unsupervised clustering revealed seven distinct mesenchymal cell types (Fig.…”

Recent developments in the synovial fibroblast pathobiology field in rheumatoid arthritis

“…In another study, the transcriptional co-factor Yes-associated protein (Yap) and its binding partner Snail were found to be upregulated in hyperplastic human RA synovium and detectable in RASF [46]. Gdf5-lineage synovial fibroblasts, cells deriving from the embryonic joint interzone [47] showed increased expression of Yap and adopted an erosive phenotype (podoplanin+Thy-1-), invading cartilage and bone. IL-6, but not TNFα or IL-1β, Jak-dependently activated Yap and induced Yap–Snail interaction.…”

Site of invasion revisited: epigenetic drivers of joint destruction in RA

“…Continuous, temporal-spatially regulated influx of Gdf5 -linage cells gives also rise to all other joint structures, such as cartilage, menisci and ligaments 19. However, a fraction not derived from this linage was found particularly in the sublining synovial compartment 20. Interestingly, in antigen-induced arthritis in mice, the proportion of lining SF (CD90 − ) increased much more in the Gdf5 -lineage SF than in SF that did not evolve from the Gdf5 -linage, where expansion of sublining SF (CD90 + ) dominated 21.…”