Individual cell bioprinting to generate multi-tissue type condensations for osteochondral tissue regeneration

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Abstract Osteochondral defects of the knee are common worldwide, yet there are few viable options for patients with damaged osteochondral tissue as current treatments do not consistently regenerate functional tissue. The standard of care for osteochondral defect repair is arthroscopic microfracture surgery, but this procedure often results in formation of mechanically inferior fibrocartilage formation. To overcome limitations of this and other surgical procedures, tissue engineering strategies, such as cell-laden biomaterial scaffolds, are promising alternative approaches to treat these defects. However, scaffold-based strategies face several challenges, such as interference with critical cell-cell interactions, potential immune and/or inflammatory reaction to the scaffold and its degradation byproducts, and unsynchronized scaffold degradation rate with that of new tissue formation. New cellular condensation strategies without a scaffold address these issues, however, it is still difficult to precisely control the architecture of the engineered tissues to mimic the sophisticated three-dimensional (3D) structure and organization of natural osteochondral tissues and their structure-derived functions. Recently, 3D bioprinting has been applied in tissue engineering with the potential to create complicated, high-resolution 3D structures. In addition, we have engineered the first technology capable of 3D printing a cell-only bioink and maintaining the printed structure, which is necessary to form cell condensations. The hypothesis of this proposal is that cellular condensation-based prevascularized osteochondral tissue constructs of precisely defined geometries can be directly assembled with human stem cells and endothelial cells via 3D bioprinting into a photocurable liquid-like solid, shear-thinning and rapid self-healing microgel slurry with spatially controlled presentation of tissue specific growth factors. Microgel photocrosslinking after printing will provide temporary mechanical stability for the printed constructs during culture to permit cellular condensation formation. This cell- only bioprinting strategy will be implemented to print seamlessly continuous two-phase osteochondral tissue constructs with a prevascularized bone phase and a cartilage phase. Specifically, this proposal aims to (1) determine the role of microgel properties on the resolution and fidelity of the cell-only 3D printed constructs, (2) engineer prevascularized osteochondral constructs with individual cell-only bioinks by spatiotemporally controlled delivery of vasculogenic, osteogenic and chondrogenic growth factors, and (3) determine the clinical potential of the 3D printed prevascularized osteochondral constructs by evaluation of new osteochondral tissue formation and integration with the host vascular networks and bone and cartilage repair in a full-thickness osteochondral rabbit defect model. This platform strategy has the potential to greatly enhance the lives of those suffering from osteochondral defects and may enable the engineering of other complex functional tissues in the body.