Biofabrication Approaches to Engineer Biomimetic Bone Tissue Interfaces

<b>Abstract</b><br/>Bone tissue defects resulting from congenital abnormalities, disease, tumor resection, and traumatic injury can severely affect patient health.[1] In many cases, defects are not limited to bone tissue alone, and damage can also involve surrounding tissues, including periosteum, cartilage, ligament, or tendon. Traditional clinical techniques for bone defect repair often include the implantation of autografts, but donor site morbidity, lack of availability, and considerable medical costs limit their applicability. Tissue engineering (TE) strategies aim to produce functional bone tissue replacements by combining scaffolds, cells, and bioactive signals. Extrusion bioprinting allows for superior control over design among these three critical components and is particularly useful in the case of replicating heterogeneous tissue interfaces.[2] Specifically, bioprinting techniques allow multi-material patterning to manipulate cell-cell or material population distances and patterns, influencing cellular crosstalk and scaffold mechanics.[3] Here, we present biofabrication strategies for the generation of such biomimetic tissue interfaces, including (1) a computational model studying the impact of printed material patterning ratios on resulting scaffold mechanics, (2) a printing approach for interfacial tissues, and (3) a 4D printing strategy to generate microscale tissues among macroscale counterparts. Using these techniques in combination with extrusion printing allows for enhanced interfacial tissue regenerative potential by improving biological and mechanical properties.<br/> <br/>3D stationary solid mechanics models were developed using COMSOL. Osteal and chondral bioinks are co-printed at the interface layer to form the mechanically interlocking interface design, recapitulating the calcified cartilage region of the osteochondral unit using polycaprolactone (PCL) and gelatin methacrylate (GelMA) for bone and cartilage inks, respectively. The COMSOL model simulated the effect of lateral shear force during articulation. This was then validated in vitro using mechanical shear analysis of printed samples.[4] For complex extrusion printing, GelMA Type B was synthesized and dissolved at 5% w/v in PBS with a photoinitiator. Both casted and printed samples were imaged post-fabrication, then treated with varying concentrations and molecular weights of poly-L-lysine to induce shrinking behavior via complex coacervation-like mechanisms.[5] After treatment, hydrogels were imaged again to observe resolution enhancement.<br/> <br/>Computational results broadly demonstrate the ability to predict printed scaffold mechanical properties, specifically that interlocking interfaces have the potential to redirect shear-induced stresses from a single tissue to the entire interface. Extrusion printing allowed complex, multiphase tissues to be constructed and demonstrated that charge-induced shrinking could fabricate microscale periosteal tissue and a macroscale bone composite. Bioprinting allows for the generation of heterogenous bone tissue interfaces with relative ease. Using pre- and post-printing biofabrication approaches, tissue interface mechanics, design, and cellular crosstalk can be readily controlled.<br/> <br/><b>References</b><br/>[1] Cooper, G.M. et al. Plast. Reconstr. Surg. 125(6): 1685-1692, 2010.<br/>[2] Murphy, S.V., Atala, A. Nat. Biotechnol. 32(8):773-85, 2014.<br/>[3] Piard, C. et al. Biomaterials. 222:119423, 2019.<br/>[4] Choe, R. et al. Biofabrication. 14(2):025015, 2022.<br/>[5] Gong, J. et al. Nat. Commun. 11:1267, 2020.<br/> <br/><b>Acknowledgements</b><br/>We acknowledge our funding support from the NIH Center for Engineering Complex Tissues (P41 EB023833) and the OsteoScience Foundation Peter Geistlich Grant.