The advent of three dimensionally (3D) printed customized bone grafts using different biomaterials has enabled repairs of complex bone defects in various in vivo models. However, studies related to their clinical translations are truly limited. Herein, 3D printed poly(lactic- co -glycolic acid)/β-tricalcium phosphate (PLGA/TCP) and TCP scaffolds with or without recombinant bone morphogenetic protein −2 (rhBMP-2) coating were utilized to repair primate’s large-volume mandibular defects and compared efficacy of prefabricated tissue-engineered bone (PTEB) over direct implantation (without prefabrication). 18 F-FDG PET/CT was explored for real-time monitoring of bone regeneration and vascularization. After 3-month’s prefabrication, the original 3D-architecture of the PLGA/TCP-BMP scaffold was found to be completely lost, while it was properly maintained in TCP-BMP scaffolds. Besides, there was a remarkable decrease in the PLGA/TCP-BMP scaffold density and increase in TCP-BMP scaffolds density during ectopic (within latissimus dorsi muscle) and orthotopic (within mandibular defect) implantation, indicating regular bone formation with TCP-BMP scaffolds. Notably, PTEB based on TCP-BMP scaffold was successfully fabricated with pronounced effects on bone regeneration and vascularization based on radiographic, 18 F-FDG PET/CT, and histological evaluation, suggesting a promising approach toward clinical translation.

Objectives Mesenchymal stem cells (MSCs) based therapy for bone regeneration has been regarded as a promising method in the clinic. However, hBMSCs with invasive harvesting process and undesirable proliferation rate hinder the extensive usage. HUCMSCs of easier access and excellent performances provide an alternative for the fabrication of tissue‐engineered bone construct. Evidence suggested the osteogenesis ability of hUCMSCs was weaker than that of hBMSCs. To address this issue, a co‐culture strategy of osteogenically and angiogenically induced hUCMSCs has been proposed since thorough vascularization facilitates the blood‐borne nutrition and oxygen to transport in the scaffold, synergistically expediting the process of ossification. Materials and methods Herein, we used osteogenic‐ and angiogenic‐differentiated hUCMSCs for co‐culture in screened culture medium to elevate the osteogenic capacity with in vitro studies and finally coupled with 3D TCP scaffold to repair rat's critical‐sized calvarial bone defect. By dual‐directional induction, hUCMSCs could differentiate into osteoblasts and endothelial cells, respectively. To optimize the co‐culture condition, gradient ratios of dual‐directional differentiated hUCMSCs co‐cultured under different medium were studied to determine the appropriate condition. Results It revealed that the osteogenic‐ and angiogenic‐induced hUCMSCs mixed with the ratio of 3:1 co‐cultured in the mixed medium of osteogenic induction medium to endothelial cell induction medium of 3:1 possessed more mineralization nodules. Similarly, ALP and osteogenesis/angiogenesis‐related genes expressions were relatively higher. Further evidence of bone defect repair with 3D printed TCP of 3:1 group exhibited better restoration outcomes. Conclusions Our work demonstrated a favourable and convenient approach of dual‐directional differentiated hUCMSCs co‐culture to improve the osteogenesis, establishing a novel way to fabricate tissue‐engineered bone graft with 3D TCP for large bone defect augmentation.

Nanocrystalline hydroxyapatite (NHA) is a biocompatible, biodegradable, and osteoconductive bone graft material; however, it lacks osteoinductivity. The present study is aimed at investigating the feasibility of nanocrystalline hydroxyapatite (NHA) as an osteoinductive growth factor carrier. Bone morphogenic protein 2 (BMP2), an osteoinductive growth factor, was incorporated into NHA (BMP2-NHA) using a simple adsorption method. The growth factor loading and release kinetics were profiled using fluorescein-isothiocyanate-labeled bovine serum albumin (FITC-BSA) as a mimic of the osteoinductive growth factor BMP2. The effect of BMP2-NHA on the osteogenic differentiation of C2C12 cells and ectopic bone formation in mice were tested. Confocal laser-scanning microscopy showed that FITC-BSA was diffused throughout the porous structure of NHA. FITC-BSA was efficiently loaded in NHA and sustained release was observed up to 35 days in vitro. BMP2-NHA enhanced the expression of osteogenic markers Runx2, Osterix, Alp, and Col1α1 and ALP activity in C2C12 cells compared to NHA. Similarly, μ-CT and histological examinations showed that BMP2-NHA robustly induced ectopic bone formation in mice. This study suggests that NHA could be used as an effective carrier of osteoinductive growth factors, which ensures osteoinductivity of NHA via sustained release of the growth factor.