Synergistic Angio-Osteogenic Effects of Copper-Releasing 3D Biocomposite Scaffolds: A Step Toward Vascularized Bone Regeneration

Critical-sized bone defects present significant clinical challenges due to inadequate vascularization and scaffold integration. This study developed a multifunctional 3D-printed polycaprolactone (PCL)-gelatin (Gel) scaffold reinforced with Bioglass particles (BGPs) or copper dopped BGPs (CuBGPs) to synergistically enhance angiogenesis and bone regeneration in rat model. The scaffolds were fabricated by infiltrating gelatin solutions containing BGPs or CuBGPs into the pores of 3D-printed PCL matrices, followed by freeze-drying. Comprehensive characterization of PCL-gel, PCL-gel-BGPs, and PCL-gel-CuBGPs scaffolds revealed optimal porosity (58.76±5.20, 53.27±11.83, and 54.5±7.61%), contact angle (74.53 ±6.6, 71.76±2.65, and 69.89±4.14), and controlled degradation (44.65±4.73, 47.93±2.51, and 50.58±5.68). MTT study demonstrated dose-dependent enhancement of cell proliferation, with BGPs and CuBGPs significantly improving mesenchymal stem cells (MSCs) viability by day 5. In vivo experiments in rat calvarial defects showed that Cu containing scaffolds promoted greater new bone volume compared to other groups at 12 weeks. Histological and immunohistochemical analyses confirmed robust angiogenesis and woven bone formation, with CuBGPs achieving the highest vasculature. This study provides a detailed and reproducible framework for Cu-doped scaffold fabrication, characterization, and application in critical-sized defect regeneration.