Printing bone in a gel: using nanocomposite bioink to print functionalised bone scaffolds

Free-form printing offers a novel biofabrication approach to generate complex shapes by depositing hydrogel materials within a temporary supportive environment. However, printed hydrogels typically lack the requisite mechanical properties and functionality of the desired tissue, limiting application and, more importantly, safety and efficacy of the implant. We have developed an innovative nanoclay-based bioink to print high shape fidelity functional constructs for potential skeletal application. Laponite® (LAP) nanoclay was combined with gellan gum (GG) to generate a printable hydrogel that was highly stable in vitro, displayed limited swelling ability compared to the silicate-free control and remained stable over time. An agarose fluid gel was found to provide the requisite support for the deposition of the material ink and preservation of the printed structure prior to crosslinking. Printed C2C12 myoblasts remained viable and displayed extensive proliferation over 21 days in culture. Cell-laden scaffolds demonstrated functionality within 1 day of culture in vitro and that was preserved over 3 weeks. Analysis of absorption and release mechanisms from LAP-GG using model proteins (lysozyme and bovine serum albumin (BSA)) demonstrated the retention capability of the clay-based materials for compound localisation and absence of burst release. Vascular endothelial growth factor (VEGF) was loaded within the agarose fluid gel and absorbed by the material ink via absorbtion during deposition. The 3D printed constructs was implanted on the chorioallantoic membrane of a 10-days old developing chick. Extensive and preferential vasculature infiltration was observed in LAP-GG loaded VEGF constructs compared to controls (p<0.01 and p<0.0001) after only 7 days of incubation. The current studies demonstrate, for the first time, the application of innovative LAP-GG 3D constructs in the generation of growth factor loaded 3D constructs for potential application in skeletal tissue repair