4D Biochemical Photocustomization of Hydrogel Scaffolds for Biomimetic Tissue Engineering.

Programmable engineered tissues and the materials that support them are instrumental to the development of next-generation therapeutics and gaining new understanding of human biology. Toward these ends, recent years have brought a growing emphasis on the creation of "4D" hydrogel culture platforms-those that can be customized in 3D space and on demand over time. Many of the most powerful 4D-tunable biomaterials are photochemically regulated, affording users unmatched spatiotemporal modulation through high-yielding, synthetically tractable, and cytocompatible reactions. Precise physicochemical manipulation of gel networks has given us the ability to drive critical changes in cell fate across a diverse range of distance and time scales, including proliferation, migration, and differentiation through user-directed intracellular and intercellular signaling. This Account provides a survey of the numerous creative approaches taken by our lab and others to recapitulate the dynamically heterogeneous biochemistry underpinning extracellular matrix (ECM)-cell interactions via light-based network (de)decoration with biomolecules (e.g., peptides, proteins) and protein activation/generation. We believe the insights gained from these studies can motivate disruptive improvements to emerging technologies, including low-variability organoid generation and culture, high-throughput drug screening, and personalized medicine. As photolithography and chemical modification strategies continue to mature, access to and control over new and increasingly complex biological pathways are being unlocked. The earliest hydrogel photopatterning efforts selectively encapsulated bioactive peptides and drugs into rudimentary gel volumes. Through continued exploration and refinement, next-generation materials now boast reversible, multiplexed, and/or Boolean logic-based biomolecule presentation, as well as functional activation at subcellular resolutions throughout 3D space. Lithographic hardware and software technologies, particularly those enabling image-guided patterning, allow researchers to precisely replicate complex biological structures within engineered tissue environments. The advent of bioorthogonal click chemistries has expanded 4D tissue engineering toolkits, permitting diverse constructs to be independently customized in the vicinity of any cell that is amenable to hydrogel-based culture. Additionally, the adoption of modern protein engineering techniques including genetic code expansion and chemoenzymatic alteration provides a roadmap toward site-specific modification of nearly any recombinant or isolated protein, affording installation of photoreactive and click handles without sacrificing their bioactivity. While the established bind, release, (de)activate paradigm in hydrogel photolithography continues to thrive alongside these modern engineering techniques, new studies are also demonstrating photocontrol of more complex or nonclassical operations, including engineered material-microorganism interfaces and functional protein photoassembly. Such creative approaches offer exciting new avenues for the field, including spatial control of on-demand biomolecule production from cellular depots and patterned bioactivity using a growing array of split protein pairs. Taken together, these technologies provide the foundation for truly biomimetic photopatterning of engineered tissues.