In vivo prevascularization strategy enhances neovascularization of ll-tricalcium phosphate scaffolds in bone regeneration

Background: Neovascularization is critical for bone regeneration. Numerous studies have explored prevasculari-zation preimplant strategies, ranging from calcium phosphate cement (CPC) scaffolds to co-culturing CPCs with stem cells. The aim of the present study was to evaluate an alternative in vivo prevascularization approach, using preimplant-prepared macroporous beta-tricalcium phosphate (ll-TCP) scaffolds and subsequent transplantation in bone defect model.Methods: The morphology of ll-TCPs was characterized by scanning electron microscopy. After 3 weeks of pre-vascularization within a muscle pouch at the lateral size of rat tibia, we transplanted prevascularized macroporous ll-TCPs in segmental tibia defects, using blank ll-TCPs as a control. Extent of neovascularization was determined by angiography and immunohistochemical (IHC) evaluations. Tibia samples were collected at different time points for biomechanical, radiological, and histological analyses. RT-PCR and western blotting were used to evaluate angio-and osteo-specific markers.Results: With macroporous ll-TCPs, we documented more vascular and supporting tissue invasion in the macro -porous ll-TCPs with prior in vivo prevascularization. Radiography, biomechanical, IHC, and histological analyses revealed considerably more vascularity and bone consolidation in ll-TCP scaffolds that had undergone the pre-vascularization step compared to the blank ll-TCP scaffolds. Moreover, the prevascularization treatment remarkably upregulated mRNA and protein expression of BMP2 and vascular endothelial growth factor (VEGF) during bone regeneration.Conclusion: This novel in vivo prevascularization strategy successfully accelerated vascular formation to bone regeneration. Our findings indicate that prevascularized tissue-engineered bone grafts have promising potential in clinical applications.The translational potential of this article: This study indicates a novel in vivo prevascularization strategy for growing vasculature on ll-TCP scaffolds to be used for repair of large segmental bone defects, might serve as a promising tissue-engineered bone grafts in the future.