In vivo prevascularized scaffold enhanced vascularization of regenerated bone within a segmental tibial defect in rats (A) Tissue sections through the segmental tibia defect immunostained for osteocalcin (OCN), a hormone secreted by osteoblasts. These sections were taken from rats 8 weeks after implantation with either a blank or prevascularized scaffold. Higher magnification photograph of boxed areas are shown to the right. Scale bars, 1000 μm n = 3 in each group (B) Integrated optical density (IOD) of OCN-positive staining in tissue sections from rats 8 weeks after implantation of scaffold (C) BMP2 and VEGFA mRNA expression levels in regenerated bone tissue 2, 4, 6, and 8 weeks after implantation of scaffold. Expression was quantified by RT-PCR. The housekeeping gene GAPDH served as an internal control. n = 3 in each group. (D) Western blots of lysates derived from regenerated bone tissue at the indicated times after implantation. n = 3 in each group. (E, F) Quantification of BMP2 and VEGFA protein expression levels in regenerated bone tissue at the indicated times after implantation of scaffold. Expression was evaluated by western blotting. The housekeeping gene GAPDH served as an internal control. Significant differences were evaluated by Student's t tests; ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001, compared with the blank group. #p < 0.05, compared with the control.