The progression of HO in trauma-induced mice model. (A) Representative imageological examination of Achilles tendons by μCT. (B) Quantitation of BV at different stages. (C, D) H&E and SOFG staining of Achilles tendons were performed at different stages to observe the osteogenesis process, respectively. (E, F) Representative immunohistochemical staining and quantitative analysis of Ocn-positive cells. (G, H) Immunohistochemical staining and quantitative analysis of p-Smad2/3 positive cells. (I) Representative western blot assay and relative quantification of Ocn and p-Smad2/3 protein levels in Achilles tendons. Scale bars: A, 2 mm; C, D, E, G, 50 μm. HO – heterotopic ossification; μCT – microcomputed tomography; BV – bone volume; H&E – hematoxylin and eosin; SOFG – Safranin O and fast green; B – bone; BM – bone marrow; T – tendon; C – cartilage. n=8 per group. All data are present as means ± standard deviation. * P<0.05.