Extended Data Figure 1. Removal of periosteum reduces bone formation and callus vascularization.

(a) Histological characterization of the murine bone autograft healing model. At the host-graft junction cartilage (Safranin O⁺) is formed at PFD7. Note absence of CD31⁺ blood vessels in these regions. Near the graft centre new woven bone (bright pink on H&E staining) is deposited, cartilage is absent and blood vessels are abundant. By PFD14, the cartilage at the host-graft junction is gradually being replaced by bone, while the woven bone near the graft centre appears mature (representative images of 4 mice). Scale bars, 200μm in host-graft junction images, 100μm in graft centre images, 50μm in magnifications. (b) MicroCT-based visualization and quantification of newly formed bone around control autografts, autografts from which the periosteum was removed or devitalized allografts (no living cells) at PFD28 (n=3 mice). Coverage ratio represents percentage of graft surface covered by new bone. (c) Dual energy microCT-based visualization and quantification of vascularization in a 250μm-wide region around autografts and allografts at PFD14 (n=5 mice for autograft, n=6 mice for devitalized allograft). (d) CD31 immunohistochemical visualization and quantification of vascularization in a 250μm-wide region around autografts and allografts at PFD14 (n=3 mice). Scale bars, 500μm. b: bone, c: cartilage, ft: fibrous tissue, g: graft, h: host, m: muscle, p: periosteum. Mean ± s.e.m. One-way ANOVA with Bonferroni post-hoc test (b), two-tailed Student’s t-test (c,d).