Fig. 7: Cxcl12-creER⁺ BMSCs transform into a skeletal stem cell-like state.

a Inducible partial cell ablation of Cxcl12-creER⁺ stromal cells using an inducible diphtheria toxin fragment A (DTA) allele. Flow cytometry-based verification of iDTA-mediated cell ablation of Cxcl12-creER⁺ stromal cells. n = 3 (tdT), n = 6 (DTA) mice. *p < 0.05, two-tailed, Mann–Whitney’s U test. b, c DTA-mediated ablation of Cxcl12-creER⁺ cells during cortical bone regeneration. Rosa26^{lsl-tdTomato/iDTA} (Control) and Cxcl12-creER; Rosa26^{lsl-tdTomato/iDTA} (DTA) femur diaphyseal cortical bones, 14 days after cortical drill-hole surgery (pulsed at 7 days before surgery). b Cortical bone injured areas. Upper panels: alkaline phosphatase (ALP) staining. Gray: DIC. Scale bar: 200 µm. Lower panels: 3D-µCT. Scale bar: 1 mm. c Bone volume/tissue volume (BV/TV, left) and bone mineral density (BMD, right) of the injured area. n = 6 (Control), n = 4 (DTA) mice. **p < 0.01, two-tailed, Mann–Whitney’s U test. d–i Single-cell RNA-seq analyses of lineage-traced Cxcl12-creER⁺ cells. Cxcl12^CE-tdTomato⁺ cells were isolated from ablated (ABL) femurs at 14 days after marrow ablation. e UMAP-based visualization of major classes of mesenchymal Cxcl12^CE-tdTomato⁺ cells (Clusters 0–9). Red dotted contour: reticular cells, green dotted contour: preosteoblasts. n = 1825 cells pooled from n = 7 mice. f Feature plots of skeletal stem cell marker genes. g Heatmap of representative pseudotime-dependent genes. h Pseudotime and single cell trajectory analysis by Monocle. i Most significant Gene Ontology (GO) terms (Biological Process) based on DAVID functional annotation of differentially expressed genes (DEGs). Black: general, red: Wnt signaling-related, blue: bone-related terms. All data are presented as mean ± s.d. Source data are provided as a Source Data file.