Synovial macrophage diversity and activation of M-CSF signaling in post-traumatic osteoarthritis

Knights, Alexander J.; Farrell, Easton C.; Ellis, Olivia M.; Song, Michelle J.; Appleton, C. Thomas; Maerz, Tristan

doi:10.1101/2023.10.03.559514

Cited by 3 publications

(1 citation statement)

References 66 publications

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“…Csf1, also known as macrophage colony stimulating factor, is involved in the proliferation, survival, and differentiation of macrophages [42]. Similar to our results, Knights et al found SF-derived Csf1 signaling to function as a major crosstalk axis of macrophages activated by injury [43]. Collectively, synovial MSCs are considered to regulate their own activation by regulating their proliferation, polarity, and secretion via Ccl7, Spp1, and Csf1 signaling (Fig.…”

Section: Discussionsupporting

confidence: 89%

Macrophage depletion in inflamed rat knees prevents the activation of synovial mesenchymal stem cells by weakening Nampt and Spp1 signaling

Kodama,

Endo,

Sekiya

2024

Preprint

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Background Macrophages and mesenchymal stem cells (MSCs) engage in crucial interplay during inflammation and have significant roles in tissue regeneration. Synovial MSCs, as key players in joint regeneration, are known to proliferate together with macrophages in synovitis. However, the crosstalk between synovial MSCs and macrophages remains unclear. In this study, we investigated changes in the activation of synovial MSCs in inflamed rat knees following selective depletion of macrophages with clodronate liposomes. Methods Acute inflammation was induced in rat knee joints by injection of carrageenan (day 0). Clodronate liposomes were administered intra-articularly on days 1 and 4 to deplete macrophages, with empty liposomes as a control. Knee joints were collected on day 7 for evaluation by histology, flow cytometry, and colony-forming assays. Concurrently, synovial MSCs were cultured and subjected to proliferation assays, flow cytometry, and chondrogenesis assessments. We also analyzed their crosstalk using single-cell RNA sequencing (scRNA-seq). Results Clodronate liposome treatment significantly reduced CD68-positive macrophage numbers and suppressed synovitis. Immunohistochemistry and flow cytometry showed decreased expression of CD68 (a macrophage marker) and CD44 and CD271 (MSC markers) in the clodronate group, while CD73 expression remained unchanged. The number of colony-forming cells per 1,000 nucleated cells and per gram of synovium was significantly lower in the clodronate group than in the control group. Cultured synovial MSCs from both groups showed comparable proliferation, surface antigen expression, and chondrogenic capacity. scRNA-seq identified seven distinct synovial fibroblast (SF) subsets, with a notable decrease in the Mki67+ SF subset, corresponding to synovial MSCs, in the clodronate group. Clodronate treatment downregulated genes related to extracellular matrix organization and anabolic pathways in Mki67+ SF. Cell-cell communication analysis revealed diminished Nampt and Spp1 signaling interaction between macrophages and Mki67+ SF and diminished Ccl7, Spp1, and Csf1 signaling interaction between Mki67+ SF and macrophages in the clodronate group. Conclusions Macrophage depletion with clodronate liposomes suppressed synovitis and reduced the number and activity of synovial MSCs, highlighting the significance of macrophage-derived Nampt and Spp1 signals in synovial MSC activation. These findings offer potential therapeutic strategies to promote joint tissue regeneration by enhancing beneficial signals between macrophages and synovial MSCs.

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Section: Discussionsupporting

confidence: 89%

Macrophage depletion in inflamed rat knees prevents the activation of synovial mesenchymal stem cells by weakening Nampt and Spp1 signaling

Kodama,

Endo,

Sekiya

2024

Preprint

View full text Add to dashboard Cite

show abstract

Macrophage depletion in inflamed rat knees prevents the activation of synovial mesenchymal stem cells by weakening Nampt and Spp1 signaling

Kodama,

Endo,

Sekiya

2024

Inflamm Regener

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Background Macrophages and mesenchymal stem cells (MSCs) engage in crucial interplay during inflammation and have significant roles in tissue regeneration. Synovial MSCs, as key players in joint regeneration, are known to proliferate together with macrophages in synovitis. However, the crosstalk between synovial MSCs and macrophages remains unclear. In this study, we investigated changes in the activation of synovial MSCs in inflamed rat knees following selective depletion of macrophages with clodronate liposomes. Methods Acute inflammation was induced in rat knee joints by injection of carrageenan (day 0). Clodronate liposomes were administered intra-articularly on days 1 and 4 to deplete macrophages, with empty liposomes as a control. Knee joints were collected on day 7 for evaluation by histology, flow cytometry, and colony-forming assays. Concurrently, synovial MSCs were cultured and subjected to proliferation assays, flow cytometry, and chondrogenesis assessments. We also analyzed their crosstalk using single-cell RNA sequencing (scRNA-seq). Results Clodronate liposome treatment significantly reduced CD68-positive macrophage numbers and suppressed synovitis. Immunohistochemistry and flow cytometry showed decreased expression of CD68 (a macrophage marker) and CD44 and CD271 (MSC markers) in the clodronate group, while CD73 expression remained unchanged. The number of colony-forming cells per 1000 nucleated cells and per gram of synovium was significantly lower in the clodronate group than in the control group. Cultured synovial MSCs from both groups showed comparable proliferation, surface antigen expression, and chondrogenic capacity. scRNA-seq identified seven distinct synovial fibroblast (SF) subsets, with a notable decrease in the Mki67+ SF subset, corresponding to synovial MSCs, in the clodronate group. Clodronate treatment downregulated genes related to extracellular matrix organization and anabolic pathways in Mki67+ SF. Cell–cell communication analysis revealed diminished Nampt and Spp1 signaling interaction between macrophages and Mki67+ SF and diminished Ccl7, Spp1, and Csf1 signaling interaction between Mki67+ SF and macrophages in the clodronate group. Spp1 and Nampt promoted the proliferation and/or chondrogenesis of synovial MSCs. Conclusions Macrophage depletion with clodronate liposomes suppressed synovitis and reduced the number and activity of synovial MSCs, highlighting the significance of macrophage-derived Nampt and Spp1 signals in synovial MSC activation. These findings offer potential therapeutic strategies to promote joint tissue regeneration by enhancing beneficial signals between macrophages and synovial MSCs.

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Applications of single-cell RNA sequencing in rheumatoid arthritis

Figueiredo

2024

Front. Immunol.

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Single cell RNA sequencing (scRNA-seq) is a relatively new technology that provides an unprecedented, detailed view of cellular heterogeneity and function by delineating the transcriptomic difference among individual cells. This will allow for mapping of cell-type-specific signaling during physiological and pathological processes, to build highly specific models of cellular signaling networks between the many discrete clusters that are present. This technology therefore provides a powerful approach to dissecting the cellular and molecular mechanisms that contribute to autoimmune diseases, including rheumatoid arthritis (RA). scRNA-seq can offer valuable insights into RA unique cellular states and transitions, potentially enabling development of novel drug targets. However, some challenges that still limit its mainstream utilization and include higher costs, a lower sensitivity for low-abundance transcripts, and a relatively complex data analysis workflow relative to bulk or traditional RNA-seq. This minireview explores the emerging application of scRNA-seq in RA research, highlighting its role in producing important insights that can help pave the way for innovative and more effective therapeutic strategies.

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Synovial macrophage diversity and activation of M-CSF signaling in post-traumatic osteoarthritis

Cited by 3 publications

References 66 publications

Macrophage depletion in inflamed rat knees prevents the activation of synovial mesenchymal stem cells by weakening Nampt and Spp1 signaling

Macrophage depletion in inflamed rat knees prevents the activation of synovial mesenchymal stem cells by weakening Nampt and Spp1 signaling

Macrophage depletion in inflamed rat knees prevents the activation of synovial mesenchymal stem cells by weakening Nampt and Spp1 signaling

Applications of single-cell RNA sequencing in rheumatoid arthritis

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