Accidents, osteoporosis or cancer can cause severe bone damage requiring grafts to heal. All current grafting methods have disadvantages including scarcity and infection/rejection risks. An alternative is therefore needed. Hydroxyapatite/calcium carbonate (HA/CC) scaffolds mimic the mineral bone composition but lack growth factors present in auto- and allografts, limiting their osteoinductive capacity. We hypothesize that this will increase the osteogenicity and osteoinductivity of scaffolds through the presence of growth factors. The objectives of this study are to develop and mass-produce grafts with enhanced osteoinductive capacity. HA/CC scaffolds were cultured together with umbilical cord mesenchymal stem cells in bioreactors so that they adhere to the surface and deposit growth factors. Cells growing on the scaffolds are confirmed by Alamar blue assays, SEM, and confocal microscopy. ELISA and IHC are used to assess the growth factor content of the finished product. It has been confirmed that cells attach to the scaffolds and proliferate over time when grown in bioreactors. Dynamic seeding of cells is clearly advantageous for cell deposits, equalizing the amount of cells on each scaffold granule. Hydroxyapatite/calcium carbonate scaffolds support cell-growth. This should be confirmed by further research, including Quantification of BMPs and other indicators of osteogenic differentiation such as Runx2, osteocalcin and ALP is pending, and amounts are expected to be increased in enhanced scaffolds and in-vivo implantation.
Bone defects require implantable graft substitutes, especially porous and biodegradable biomaterial for tissue regeneration. The aim of this study was to fabricate and assess a 3D-printed biodegradable hydroxyapatite/calcium carbonate scaffold for bone regeneration. A 3D-printed biodegradable biomaterial containing calcium phosphate and aragonite (calcium carbonate) was fabricated using a Bioplotter. The physicochemical properties of the material were characterised. The materials were assessed in vitro for cytotoxicity and ostegenic potential and in vivo in rat intercondylar Φ3mm bone defect model for 3 months and Φ5mm of mini pig femoral bone defects for 6 months. The results showed that the materials contained hydroxyapatite and calcium carbonate, with the compression strength of 2.49± 0.2 MPa, pore size of 300.00 ± 41mm, and porosity of 40.±3%. The hydroxyapatite/aragonite was not cytotoxic and it promoted osteogenic differentiation of human umbilical cord matrix mesenchymal stem cells in vitro. After implantation, the bone defects were healed in the treatment group whereas the defect of controlled group with gelatin sponge implantation remained non-union. hydroxyapatite/aragonite fully integrated with host bone tissue and bridged the defects in 2 months, and significant biodegradation was followed by host new bone formation. After implantation into Φ5mm femoral defects in mini pigs hydroxyapatite/aragonite were completed degraded in 6 months and fully replaced by host bone formation, which matched the healing and degradation of porcine allogenic bone graft. In conclusion, hydroxyapatite/aragonite is a suitable new scaffold for bone regeneration. The calcium carbonate in the materials may have played an important role in osteogenesis and material biodegradation.Materials and methods: