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OBJECTIVES: Osteophytes are highly prevalent in osteoarthritis (OA) and are associated with pain and functional disability. These pathological outgrowths of cartilage and bone typically form at the junction of articular cartilage, periosteum and synovium. The aim of this study was to identify the cells forming osteophytes in OA. METHODS: Fluorescent genetic cell-labelling and tracing mouse models were induced with tamoxifen to switch on reporter expression, as appropriate, followed by surgery to induce destabilisation of the medial meniscus. Contributions of fluorescently labelled cells to osteophytes after 2 or 8 weeks, and their molecular identity, were analysed by histology, immunofluorescence staining and RNA in situ hybridisation. Pdgfrα-H2BGFP mice and Pdgfrα-CreER mice crossed with multicolour Confetti reporter mice were used for identification and clonal tracing of mesenchymal progenitors. Mice carrying Col2-CreER, Nes-CreER, LepR-Cre, Grem1-CreER, Gdf5-Cre, Sox9-CreER or Prg4-CreER were crossed with tdTomato reporter mice to lineage-trace chondrocytes and stem/progenitor cell subpopulations. RESULTS: Articular chondrocytes, or skeletal stem cells identified by Nes, LepR or Grem1 expression, did not give rise to osteophytes. Instead, osteophytes derived from Pdgfrα-expressing stem/progenitor cells in periosteum and synovium that are descendants from the Gdf5-expressing embryonic joint interzone. Further, we show that Sox9-expressing progenitors in periosteum supplied hybrid skeletal cells to the early osteophyte, while Prg4-expressing progenitors from synovial lining contributed to cartilage capping the osteophyte, but not to bone. CONCLUSION: Our findings reveal distinct periosteal and synovial skeletal progenitors that cooperate to form osteophytes in OA. These cell populations could be targeted in disease modification for treatment of OA.

Funder: NHS Blood and Transplant; FundRef: http://dx.doi.org/10.13039/100009033, Funder: Victorian Government‘s Operational Infrastructure Support Program, Objectives: Osteophytes are highly prevalent in osteoarthritis (OA) and are associated with pain and functional disability. These pathological outgrowths of cartilage and bone typically form at the junction of articular cartilage, periosteum and synovium. The aim of this study was to identify the cells forming osteophytes in OA. Methods: Fluorescent genetic cell-labelling and tracing mouse models were induced with tamoxifen to switch on reporter expression, as appropriate, followed by surgery to induce destabilisation of the medial meniscus. Contributions of fluorescently labelled cells to osteophytes after 2 or 8 weeks, and their molecular identity, were analysed by histology, immunofluorescence staining and RNA in situ hybridisation. Pdgfrα-H2BGFP mice and Pdgfrα-CreER mice crossed with multicolour Confetti reporter mice were used for identification and clonal tracing of mesenchymal progenitors. Mice carrying Col2-CreER, Nes-CreER, LepR-Cre, Grem1-CreER, Gdf5-Cre, Sox9-CreER or Prg4-CreER were crossed with tdTomato reporter mice to lineage-trace chondrocytes and stem/progenitor cell subpopulations. Results: Articular chondrocytes, or skeletal stem cells identified by Nes, LepR or Grem1 expression, did not give rise to osteophytes. Instead, osteophytes derived from Pdgfrα-expressing stem/progenitor cells in periosteum and synovium that are descendants from the Gdf5-expressing embryonic joint interzone. Further, we show that Sox9-expressing progenitors in periosteum supplied hybrid skeletal cells to the early osteophyte, while Prg4-expressing progenitors from synovial lining contributed to cartilage capping the osteophyte, but not to bone. Conclusion: Our findings reveal distinct periosteal and synovial skeletal progenitors that cooperate to form osteophytes in OA. These cell populations could be targeted in disease modification for treatment of OA.

Funder: Biotechnology and Biological Sciences Research Council; Id: http://dx.doi.org/10.13039/501100000268, Funder: Versus Arthritis; Id: http://dx.doi.org/10.13039/501100012041, Lipids play a crucial role in signaling and metabolism, regulating the development and maintenance of the skeleton. Membrane lipids have been hypothesized to act as intermediates upstream of orphan phosphatase 1 (PHOSPHO1), a major contributor to phosphate generation required for bone mineralization. Here, we spatially resolve the lipid atlas of the healthy mouse knee and demonstrate the effects of PHOSPHO1 ablation on the growth plate lipidome. Lipids spanning 17 subclasses were mapped across the knee joints of healthy juvenile and adult mice using matrix-assisted laser desorption ionization imaging mass spectrometry (MALDI-IMS), with annotation supported by shotgun lipidomics. Multivariate analysis identified 96 and 80 lipid ions with differential abundances across joint tissues in juvenile and adult mice, respectively. In both ages, marrow was enriched in phospholipid platelet activating factors (PAFs) and related metabolites, cortical bone had a low lipid content, whereas lysophospholipids were strikingly enriched in the growth plate, an active site of mineralization and PHOSPHO1 activity. Spatially-resolved profiling of PHOSPHO1-knockout (KO) mice across the resting, proliferating, and hypertrophic growth plate zones revealed 272, 306, and 296 significantly upregulated, and 155, 220, and 190 significantly downregulated features, respectively, relative to wild-type (WT) controls. Of note, phosphatidylcholine, lysophosphatidylcholine, sphingomyelin, lysophosphatidylethanolamine, and phosphatidylethanolamine derived lipid ions were upregulated in PHOSPHO1-KO versus WT. Our imaging pipeline has established a spatially-resolved lipid signature of joint tissues and has demonstrated that PHOSPHO1 ablation significantly alters the growth plate lipidome, highlighting an essential role of the PHOSPHO1-mediated membrane phospholipid metabolism in lipid and bone homeostasis. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

ObjectivesOsteophytes are highly prevalent in osteoarthritis (OA) and are associated with pain and functional disability. These pathological outgrowths of cartilage and bone typically form at the junction of articular cartilage, periosteum and synovium. The aim of this study was to identify the cells forming osteophytes in OA.MethodsFluorescent genetic cell-labelling and tracing mouse models were induced with tamoxifen to switch on reporter expression, as appropriate, followed by surgery to induce destabilisation of the medial meniscus. Contributions of fluorescently labelled cells to osteophytes after 2 or 8 weeks, and their molecular identity, were analysed by histology, immunofluorescence staining and RNA in situ hybridisation. Pdgfrα-H2BGFP mice and Pdgfrα-CreER mice crossed with multicolour Confetti reporter mice were used for identification and clonal tracing of mesenchymal progenitors. Mice carrying Col2-CreER, Nes-CreER, LepR-Cre, Grem1-CreER, Gdf5-Cre, Sox9-CreER or Prg4-CreER were crossed with tdTomato reporter mice to lineage-trace chondrocytes and stem/progenitor cell subpopulations.ResultsArticular chondrocytes, or skeletal stem cells identified by Nes, LepR or Grem1 expression, did not give rise to osteophytes. Instead, osteophytes derived from Pdgfrα-expressing stem/progenitor cells in periosteum and synovium that are descendants from the Gdf5-expressing embryonic joint interzone. Further, we show that Sox9-expressing progenitors in periosteum supplied hybrid skeletal cells to the early osteophyte, while Prg4-expressing progenitors from synovial lining contributed to cartilage capping the osteophyte, but not to bone.ConclusionOur findings reveal distinct periosteal and synovial skeletal progenitors that cooperate to form osteophytes in OA. These cell populations could be targeted in disease modification for treatment of OA.