Low-Temperature Three-Dimensional Bioprinted Dual-Factor rhBMP-2/VEGF-165 Biomimetic Scaffolds for Synergistic Bone-Vascularization Repair in Large Segmental Bone Defect

The reconstruction of large segmental bone defects remains a formidable challenge in orthopedic surgery. Benefiting from the rapid advancement of three-dimensional (3D) printing technology, growth factor-loaded artificial bone scaffolds have been extensively studied and have emerged as one of the pivotal strategies for bone defect repair. However, the development of an ideal scaffold system that meets comprehensive clinical requirements still requires sustained efforts. This study designed a biomimetic bone repair scaffold with bioactive properties using low-temperature 3D bioprinting technology and systematically evaluated its osteogenic capacity in large segmental radial bone defects of New Zealand White rabbits. In this study, four types of composite materials were fabricated: polycaprolactone (PCL)/xenogeneic bone powder (XBP) (PB), PCL/XBP with recombinant human bone morphogenetic protein-2 (rhBMP-2) (PBB), PCL/XBP with vascular endothelial growth factor-165 (VEGF-165) (PBV), and PCL/XBP coloaded with rhBMP-2/VEGF-165 (PBBV). Characterization of the scaffolds revealed that the fabricated scaffolds exhibited a well-interconnected porous structure with a porosity of 58.7% and compressive modulus ranging from 61.73 ± 8.11 to 78.72 ± 9.83 MPa. The controlled release profiles showed sustained BMP-2 release (23.6 ± 1.52% to 28.8 ± 2.91% cumulative release over 14 days) and biphasic VEGF-165 release (initial burst followed by sustained release reaching 67.9 ± 4.51% to 75.9 ± 9.44%). In vitro bone marrow mesenchymal stem cells (BMSCs) coculture and in vivo mouse muscle pouch implantation confirmed excellent biocompatibility and early osteogenic-angiogenic potential. Validation in the large segmental rabbit radial defect model demonstrated through multimodal assessments, including micro-CT, histomorphometry, and vascular perfusion angiography, that the PBBV scaffold group (dual-factor loaded scaffolds) exhibits significantly superior osteogenic and angiogenic performance compared to the other groups. In conclusion, the BMP/VEGF coloaded biomimetic scaffold achieves spatiotemporal coordination of vascularization and osteogenesis through innovative structural design and controlled release kinetics. This study addresses critical limitations in the repair of large segmental bone defects, offering a translatable solution combining 3D-printable customization, mechanical support, and biofunctional synergy.