Characterization and Biological Evaluation of 3D Printed Composite Ink Consisting of Collagen, Hyaluronic Acid and Calcium Phosphate for Bone Regeneration

In large bone defects the self-healing capacity is insufficient, and the current standard treatment, autologous bone grafting, has severe disadvantages such as limited availability and donor site morbidity. Alternatively, clinically available bone graft substitutes lack spatial control over scaffold architecture to anatomically match complicated bone defects. Therefore, the aim in this study was to develop a 3D printable composite biomaterial-ink to promote healing of large bone defects. The composite biomaterial-ink consisted of an organic biopolymer matrix with tyramine modified hyaluronic acid (THA) and collagen type I (Col) mixed with osteoinductive calcium phosphate particles (CaP). The biopolymer was combined with 0, 10, 20 and 30% of either 45-63 µm or 45-106 µm CaP. µCT imaging showed a homogeneous distribution of CaP in the THA-Col hydrogel and all composites were 3D printable. In vitro cell activity assays revealed no indirect cytotoxicity using L929 cells and high cell cytocompatibility using human mesenchymal stromal cells (hMSCs). Additionally, all composites supported in vitro osteogenic differentiation of hMSCs. This study highlights the development of a 3D printable composite biomaterial-ink using CaP and THA-Col hydrogel that holds significant potential to be used as patient-specific bone graft substitute for the regeneration of large bone defects. Statement of significance : This paper introduces a 3D printable composite biomaterial-ink made of osteoinductive calcium phosphate particles combined with matrix biopolymers collagen and hyaluronic acid, which was chemically modified to introduce shear thinning and shape fixation properties for 3D printing. The chemical modification only involves a small percentage of functional groups, preserving hyaluronan's biological properties. We demonstrated printability, the homogeneous distribution of the mineral phase, cytocompatibility and that the composites support osteogenesis of primary human mesenchymal stromal cells from multiple donors. The printability of the composite biomaterial-ink allows the creation of patient-specific implants with controlled geometry on porosity. This study contributes towards engineering personalized implants for replacing autologous bone grafting in all clinical situations where the bone self-healing capacity is insufficient.