We engineered self-oxygenating tissues, which was achieved via the incorporation of hydrophobic oxygen-generating micromaterials into engineered tissues. Self-oxygenation of tissues offered temporary metabolic protection from the formation of anoxic microenvironments by in situ elevating the oxygen tension within living tissues in a homogenous and tissue size-independent manner. This capability was leveraged to transform anoxic stress into hypoxic stimulation, which resulted in the sustained production of high quantities of vascular endothelial growth factor. Self-oxygenation of tissues was therefore proved as a novel and effective strategy to protect and vascularize living implants for organ transplantation and regenerative medicine applications. To maximize the performance of our self-oxygenating tissues, we minimized hydrogen peroxide toxicity by designed incorporation of catalase. Surprisingly, we demonstrated that self-oxygenation of tissues is not only important to maintaining tissues survival, but can also guide cell fate by for example being osteo-inductive. To aid clinical and commercial translation of these micromaterials, we build a custom designed in-air microfluidic set-up that allowed for ultra-high throughput fabrication of micromaterials. We have combined these oxygen generating micromaterials with a new material that we have developed. Specifically, hydrogels were orthogonally post-functionalized with desthiobiotinylated moieties using multivalent neutravidin. In situ exchange of desthiobiotin by biotin enables spatiotemporal material functionalization as demonstrated by the formation of long-range, conformal, and contradirectional biochemical gradients within complex-shaped 3D hydrogels. Temporal control over engineered tissue biochemistry is further demonstrated by timed presentation and sequestration of growth factors using desthiobiotinylated antibodies. The method’s universality is confirmed by modifying hydrogels with biotinylated fluorophores, peptides, nanoparticles, enzymes, and antibodies. Overall, this work provides a facile, cytocompatible, and universal strategy to spatiotemporally functionalize materials, which is of great value as the desired osteogenic bio-ink as well as the vascular nanocoating, which is used for our efforts on bioprinting capillary blood vessel networks. We have created a method based on the photo-annealing of micromaterials to create perfusable vascular beds. Uniquely, this allowed for the creation of high density capillary beds within engineered tissues in a scalable manner. We also invented a novel embedded bioprinting approach based on leveraging aqueous two phase systems, which enabled fast yet stable printing of low viscous materials, which was previously not possible. This allowed for the vast expansion of printable materials and made the print resolution independent of the print nozzle. We combined this print technology with our photoannealable micromaterials to create engineered tissues containing vascular trees composed of large vascular channels that branch into a 3D high density capillary network, and vice versa. We also demonstrated that our mechanism of crosslinking, which is based on oxidative crosslinking of phenolic compounds, not only is effective in creating these clinically-sized and highly perfused tissues, but also orchestrates mechanotransduction to encapsulated cells in a previously unknown manner. This mechanism relies on the fact that instead of placing cells in a material and then form reversible interactions, we crosslinking the material directly to cells and their microenvironment in a semi-permanent manner. Together, ENABLE has provided numerous innovations that represent a highly useful and versatile toolbox to allow the survival and function of clinically-sized implant.
