A major problem in the treatment of large bone defects is the inability to provide an adequate blood supply to the implantation site. Therefore, a bone regeneration strategy that provides an adequate supply of vessels would address this need. Cobalt (Co2+), because of its ability to induce hypoxia, has been used to accelerate new vessel formation. In this study, we used a freeze-drying technique to fabricate a scaffold that consisted of Co2+-doped calcium phosphate (CaP) [e.g., hydroxyapatite (HA)] and natural silk fiber through an optimized alternate mineralization process. The composition and structure of the scaffold were confirmed by X-ray diffraction (XRD), Fourier transform infrared (FTIR), inductively coupled plasma (ICP), and scanning electron microscope (SEM). The data showed that the scaffolds promoted differentiation of adipose-derived mesenchymal stem cells (ADSCs) toward endothelial and osteoblast linages. We observed improved angiogenesis and bone formation with the fabricated scaffolds compared with the control groups. Computed tomography (CT) scans and radiographic imaging, in addition to histology and immunohistochemical analyses, showed the presence of angiogenesis and bone regeneration after implantation of the ADSC-seeded scaffolds in a critical size calavarial bone defect in a Wistar rat model. We obtained the best in vitro and in vivo results by doping 2% Co2+ in HA. Taken together, we propose that the Co2+-doped HA/silk fibroin (SF) scaffold would be a good candidate to induce angiogenesis and bone formation both in vitro and in vivo.
Keywords: angiogenesis; bone tissue engineering; cobalt; osteogenesis; silk fibroin.