Pushing the rheological and mechanical boundaries of extrusion-based 3D bioprinting

A challenge for tissue engineering is producing three-dimensional (3D), vascularized cellular constructs of clinically relevant size, shape and structural integrity. We present an integrated tissue-organ printer (ITOP) that can fabricate stable, human-scale tissue constructs of any shape. Mechanical stability is achieved by printing cell-laden hydrogels together with biodegradable polymers in integrated patterns and anchored on sacrificial hydrogels. The correct shape of the tissue construct is achieved by representing clinical imaging data as a computer model of the anatomical defect and translating the model into a program that controls the motions of the printer nozzles, which dispense cells to discrete locations. The incorporation of microchannels into the tissue constructs facilitates diffusion of nutrients to printed cells, thereby overcoming the diffusion limit of 100-200 μm for cell survival in engineered tissues. We demonstrate capabilities of the ITOP by fabricating mandible and calvarial bone, cartilage and skeletal muscle. Future development of the ITOP is being directed to the production of tissues for human applications and to the building of more complex tissues and solid organs.

Collagen is the primary component of the extracellular matrix in the human body. It has proved challenging to fabricate collagen scaffolds capable of replicating the structure and function of tissues and organs. We present a method to 3D-bioprint collagen using freeform reversible embedding of suspended hydrogels (FRESH) to engineer components of the human heart at various scales, from capillaries to the full organ. Control of pH-driven gelation provides 20-micrometer filament resolution, a porous microstructure that enables rapid cellular infiltration and microvascularization, and mechanical strength for fabrication and perfusion of multiscale vasculature and tri-leaflet valves. We found that FRESH 3D-bioprinted hearts accurately reproduce patient-specific anatomical structure as determined by micro–computed tomography. Cardiac ventricles printed with human cardiomyocytes showed synchronized contractions, directional action potential propagation, and wall thickening up to 14% during peak systole.

Significant research over the past two decades has established that extracellular matrix (ECM) elasticity, or stiffness, impacts fundamental cell processes including spreading, growth, proliferation, migration, differentiation, and organoid formation. Linearly elastic polyacrylamide hydrogels and polydimethylsiloxane (PDMS) elastomers coated with ECM proteins have become widely-used tools for assessing the role of stiffness, and results from these experiments are often assumed to reproduce the effect of the mechanical environment experienced by cells in vivo . However, tissues and ECMs are not linearly elastic materials – they in fact exhibit far more complex mechanical behaviors, including viscoelasticity, or a time-dependent response to loading or deformation, as well as mechanical plasticity and nonlinear elasticity. Recent work has revealed that matrix viscoelasticity regulates these same fundamental cell processes, and importantly can promote behaviors not observed with elastic hydrogels in both 2D and 3D culture microenvironments. These important findings have provided new insights into cell-matrix interactions and have given context as to how these interactions differentially modulate mechano-sensitive molecular pathways in cells. Moreover, these results indicate new design guidelines for the next generation of biomaterials that better match tissue and ECM mechanics for in vitro tissue models and applications in regenerative medicine.