Investigation of a Prevascularized Bone Graft for Large Defects in the Ovine Tibia.

In vivo bioreactors are a promising approach for engineering vascularized autologous bone grafts to repair large bone defects. In this pilot parametric study, we first developed a three-dimensional (3D) printed scaffold uniquely designed to accommodate inclusion of a vascular bundle and facilitate growth factor delivery for accelerated vascular invasion and ectopic bone formation. Second, we established a new sheep deep circumflex iliac artery (DCIA) model as an in vivo bioreactor for engineering a vascularized bone graft and evaluated the effect of implantation duration on ectopic bone formation. Third, after 8 weeks of implantation around the DCIA, we transplanted the prevascularized bone graft to a 5 cm segmental bone defect in the sheep tibia, using the custom 3D printed bone morphogenic protein 2 (BMP-2) loaded scaffold without prior in vivo bioreactor maturation as a control. Analysis by micro-computed tomography and histomorphometry found ectopic bone formation in BMP-2 loaded scaffolds implanted for 8 and 12 weeks in the iliac pouch, with greater bone formation occurring after 12 weeks. Grafts transplanted to the tibial defect supported bone growth, mainly on the periphery of the graft, but greater bone growth and less soft tissue invasion was observed in the avascular BMP-2 loaded scaffold implanted directly into the tibia without prior in vivo maturation. Histopathological evaluation noted considerably greater vascularity in the bone grafts that underwent in vivo maturation with an inserted vascular bundle compared with the avascular BMP-2 loaded graft. Our findings indicate that the use of an initial DCIA in vivo bioreactor maturation step is a promising approach to developing vascularized autologous bone grafts, although scaffolds with greater osteoinductivity should be further studied. Impact statement This translational pilot study aims at combining a tissue engineering scaffold strategy, in vivo prevascularization, and a modified transplantation technique to accelerate large segmental bone defect repair. First, we three-dimensional (3D) printed a 5 cm scaffold with a unique design to facilitate vascular bundle inclusion and osteoinductive growth factor delivery. Second, we established a new sheep deep circumflex iliac artery model as an in vivo bioreactor for prevascularizing the novel 3D printed osteoinductive scaffold. Subsequently, we transplanted the prevascularized bone graft to a clinically relevant 5 cm segmental bone defect in the sheep tibia for bone regeneration.

Osteoinductive 3D printed scaffold healed 5 cm segmental bone defects in the ovine metatarsus.

Autologous bone grafts are considered the gold standard grafting material for the treatment of nonunion, but in very large bone defects, traditional autograft alone is insufficient to induce repair. Recombinant human bone morphogenetic protein 2 (rhBMP-2) can stimulate bone regeneration and enhance the healing efficacy of bone grafts. The delivery of rhBMP-2 may even enable engineered synthetic scaffolds to be used in place of autologous bone grafts for the treatment of critical size defects, eliminating risks associated with autologous tissue harvest. We here demonstrate that an osteoinductive scaffold, fabricated by combining a 3D printed rigid polymer/ceramic composite scaffold with an rhBMP-2-eluting collagen sponge can treat extremely large-scale segmental defects in a pilot feasibility study using a new sheep metatarsus fracture model stabilized with an intramedullary nail. Bone regeneration after 24 weeks was evaluated by micro-computed tomography, mechanical testing, and histological characterization. Load-bearing cortical bridging was achieved in all animals, with increased bone volume observed in sheep that received osteoinductive scaffolds compared to sheep that received an rhBMP-2-eluting collagen sponge alone.

Effect of Electron Beam Sterilization on Three-Dimensional-Printed Polycaprolactone/Beta-Tricalcium Phosphate Scaffolds for Bone Tissue Engineering.

IMPACT STATEMENT: Providing customized geometries and improved control in physical and biological properties, 3D-printed polycaprolactone/beta-tricalcium phosphate (PCL/ß-TCP) composite constructs are of high interest for bone tissue engineering applications. A critical step toward the translation and clinical applications of these types of scaffolds is terminal sterilization, and E-beam irradiation might be the most relevant method because of PCL properties. Through in vitro experimental testing of both physical and biological properties, it is proven in this article that E-beam irradiation is relevant for sterilization of 3D-printed PCL/ß-TCP scaffolds for bone tissue engineering applications.

Systematic characterization of 3D-printed PCL/ß-TCP scaffolds for biomedical devices and bone tissue engineering: influence of composition and porosity.

This work aims at providing guidance through systematic experimental characterization, for the design of 3D printed scaffolds for potential orthopaedic applications, focusing on fused deposition modeling (FDM) with a composite of clinically available polycaprolactone (PCL) and ß-tricalcium phosphate (ß-TCP). First, we studied the effect of the chemical composition (0% to 60% ß-TCP/PCL) on the scaffold's properties. We showed that surface roughness and contact angle were respectively proportional and inversely proportional to the amount of ß-TCP, and that degradation rate increased with the amount of ceramic. Biologically, the addition of ß-TCP enhanced proliferation and osteogenic differentiation of C3H10. Secondly, we systematically investigated the effect of the composition and the porosity on the 3D printed scaffold mechanical properties. Both an increasing amount of ß-TCP and a decreasing porosity augmented the apparent Young's modulus of the 3D printed scaffolds. Third, as a proof-of-concept, a novel multi-material biomimetic implant was designed and fabricated for potential disk replacement.