Abstract
Hypertrophic chondrocytes give rise to osteoblasts during skeletal development; however, the process by which these non-mitotic cells make this transition is not well understood. Prior studies have also suggested that skeletal stem and progenitor cells (SSPCs) localize to the surrounding periosteum and serve as a major source of marrow-associated SSPCs, osteoblasts, osteocytes, and adipocytes during skeletal development. To further understand the cell transition process by which hypertrophic chondrocytes contribute to osteoblasts or other marrow associated cells, we utilized inducible and constitutive hypertrophic chondrocyte lineage tracing and reporter mouse models (Col10a1CreERT2; Rosa26fs-tdTomato and Col10a1Cre; Rosa26fs-tdTomato) in combination with a PDGFRaH2B-GFP transgenic line, single-cell RNA-sequencing, bulk RNA-sequencing, immunofluorescence staining, and cell transplantation assays. Our data demonstrate that hypertrophic chondrocytes undergo a process of dedifferentiation to generate marrow-associated SSPCs that serve as a primary source of osteoblasts during skeletal development. These hypertrophic chondrocyte-derived SSPCs commit to a CXCL12-abundant reticular (CAR) cell phenotype during skeletal development and demonstrate unique abilities to recruit vasculature and promote bone marrow establishment, while also contributing to the adipogenic lineage.
Highlights
Most skeletal elements arise from a cartilage template via the process of endochondral ossification (Long and Ornitz, 2013; Olsen et al, 2000; Ono et al, 2019)
Administration, the tdTOMATO reporter had cleared from the hypertrophic chondrocytes and was exclusively present in a number marrow associated cells; some of those being at a significant distance from the growth plate (Figure 1f)
In this distal region of the marrow, we observed trabecular bone lining cells that co-labeled for tdTOMATO and OCN (Figure 1g; white arrows), suggesting that these cells had transitioned from hypertrophic chondrocytes to osteoblasts
Summary
Most skeletal elements arise from a cartilage template via the process of endochondral ossification (Long and Ornitz, 2013; Olsen et al, 2000; Ono et al, 2019). Mesenchymal stem and progenitor cells condense and differentiate to form chondrocytes or cartilage cells (Hirao et al, 2006; Maes et al, 2012; Olsen et al, 2000; Spencer et al, 2014) These chondrocytes undergo rapid proliferation while depositing an extracellular matrix (ECM) rich in type II collagen (COL2A1), forming the initial immature cartilage rudiments (Ono et al, 2019). The chondrocytes localized nearest the center of each rudiment exit the cell cycle and undergo a process of maturation and hypertrophic differentiation (Capasso et al, 1984; Kong et al, 1993; Kwan et al, 1997; Long and Ornitz, 2013; Olsen et al, 2000; Ono et al, 2019; Reginato et al, 1986; Schmid and Conrad, 1982). Hypertrophic chondrocytes are essential for both establishment of the marrow cavity and bone formation; their precise fate and direct or indirect contribution to bone formation has been questioned (Tsang et al, 2015)
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