A replicating stem-like cell that contributes to bone morphogenetic protein 2-induced heterotopic bone formation

Abstract

Bone morphogenetic protein 2 (BMP2)-induced heterotopic bone formation (HBF) starts synchronously from zero upon BMP2 induction, which is advantageous for lineage tracking. The studies reported here in GLAST-Cre Ert2 :tdTomato red (TR) floxSTOPflox mice during BMP2-induced HBF show 78.8 ± 11.6% of chondrocytes and 86.5 ± 1.9% of osteoblasts are TR + after approximately 1 week. Clustering after single-cell RNAseq resulted in nine cell types, and analysis revealed one as a highly replicating stem-like cell (RSC). Pseudotiming suggested that the RSC transitions to a mesenchymal stem-like cell that simultaneously expresses multiple osteoblast and chondrocyte transcripts (chondro-osseous progenitor [COP]). RSCs and COPs were isolated using flow cytometry for unique surface markers. Isolated RSCs (GLAST-TR + Hmmr + Cd200 −) and COPs (GLAST-TR + Cd200 + Hmmr −) were injected into the muscle of mice undergoing HBF. Approximately 9% of the cells in heterotopic bone (HB) in mice receiving RSCs were GLAST-TR + , compared with less than 0.5% of the cells in mice receiving COPs, suggesting that RSCs are many times more potent than COPs. Analysis of donor-derived TR + RSCs isolated from the engrafted HB showed approximately 50% were COPs and 45% were other cells, presumably mature bone cells, confirming the early nature of the RSCs. We next isolated RSCs from these mice (approximately 300) and injected them into a second animal, with similar findings upon analysis of HBF. Unlike other methodology, single cell RNAseq has the ability to detect rare cell populations such as RSCs. The fact that RSCs can be injected into mice and differentiate suggests their potential utility for tissue regeneration.

A Population of M2 Macrophages Associated With Bone Formation

“…The earliest cell type in the genesis of ADRβ3+ macrophages is MH6, which appears to undergo further changes to form other M2 macrophages ( Supplemental Figure 4C ). It is interesting that MH6 shows high homology to the Replicating Stem-like Cell, which is a stem-like cell for osteogenesis that goes next to the Chondro-Osseous Progenitor (COP) through an epithelial to mesenchymal transition ( 44 ). The MH6 progenitor leads directly to MH7 ( Supplemental Figure 4 ), that contains markers of hemogenic endothelial cells and MH7 is the progenitor for MH0.…”

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“…14,15 To confirm these findings, researchers led by Alan R. Davis (Baylor College of Medicine, Houston, Texas) employed a mouse reporter model (GLAST-Tomato Red) to identify the exact progenitor population responsible for BMP2-induced heterotopic bone formation. As reported in their STEM CELLS Translational Medicine article, 11 Mejia et al employed highly sensitive single-cell RNA sequencing of fluorescently labeled chondrocytes and osteoblasts after BMP2-induced heterotopic bone formation and identified a highly replicating stem-like cell or RSC as a candidate for study. The authors demonstrated that RSCs undergo an epithelial-to-mesenchymal transition (like neural crest stem cells) to produce mesenchymal stem cell-like cells that express osteoblast and chondrocyte transcripts and generate bone and cartilage tissue.…”

“…Further insight into BMP‐mediated signaling may aid the efficient differentiation of stem/progenitor cells into therapeutically relevant somatic cell types and further our understanding of diverse human pathologies associated with aberrant BMP pathway regulation 10 . In the second of our Featured Articles published this month in STEM CELLS Translational Medicine , Mejia et al identify a highly replicating stem‐like cell (RSC) as the early stem/progenitor cell that initiates BMP2‐induced heterotopic bone formation 11 . In a Related Article published recently in STEM CELLS , Dries et al systematically profiled multiple components of the TGFβ signaling pathway during neural differentiation to reveal the existence of cell‐stage specific networks of individual components 12 …”

“…10 In the second of our Featured Articles published this month in STEM CELLS Translational Medicine, Mejia et al identify a highly replicating stem-like cell (RSC) as the early stem/progenitor cell that initiates BMP2-induced heterotopic bone formation. 11 In a Related Article published recently in STEM CELLS, Dries et al systematically profiled multiple components of the TGFβ signaling pathway during neural differentiation to reveal the existence of cellstage specific networks of individual components. 12…”

Human osteoprogenitor cells obtained from traumatic heterotopic ossification samples showed enhanced osteogenic differentiation potential and ERK/hedgehog signaling than that from normal bone

“…Aberrant enhanced osteogenesis within soft tissues responding to the trauma has long been discussed to be the core mechanism for the development of HO. Despite the diversity of stimulus that cause HO, it has been reported that the osteogenic signals remained greatly elevated during HO progression and committed progenitor cells to enhanced osteogenic behavior 8,9 . In the studies incorporating brain‐trauma or burn‐injury into animal models that recapitulate the trauma‐induced HO, the osteogenic activities of relevant progenitor cells can even be further reinforced by these concomitant injuries that are clinically relevant in the context of HO formation 10,11 .…”