Novel hydrogel microfibers and microparticles via interfacial bioorthogonal tetrazine ligation
Date
2022
Authors
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Journal ISSN
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Publisher
University of Delaware
Abstract
Tetrazine ligation, the cycloaddition of s-tetrazine (Tz) with trans-cyclooctene (TCO) derivatives, is biocompatible, does not require catalysis, and proceeds with exceptional rates. In this dissertation, tetrazine ligation is utilized to produce novel hydrogel microfibers and microspheres via a diffusion controlled interfacial polymerization mechanism. Because the reaction is faster than molecular diffusion, materials composition and properties can be readily tuned by varying the identity of the diffusive species. ☐ First, I synthesized hydrogel microfibers that resemble fibrous proteins found in the natural extracellular matrix, with added tunability provided by the synthetic polymer building blocks. Crosslinked, degradable, and cell adhesive hydrogel microfibers were produced via tetrazine ligation-mediated interfacial polymerization. A hydrophobic tris-TCO crosslinker, as well as homo-difunctional poly(ethylene glycol) (PEG)-based macromers with the Tz group conjugated to the PEG backbone via a stable carbamate (PEG-bisTz1) bond or a labile hydrazone (PEG-bisTz2) linkage were synthesized. Microfibers were pulled from the oil-water interface using tris-TCO in ethyl acetate along with bisTz macromers in water. The resultant microfibers exhibited comparable mechanical and thermal properties, but different aqueous stability. Combining PEG-bisTz2 with the PEG-bisTz3 macromer consisting of a non-degradable backbone and a dangling arginine-glycine-aspartic acid (RGD) peptide in the aqueous phase yielded degradable fibers that supported the attachment and growth of primary vocal fold fibroblasts. ☐ Separately, I investigated the utility of tetrazine ligation for producing hydrogel particles with a core-shell pattern. Hyaluronic acid (HA)-based hydrogel microparticles were fabricated and subsequently soaked in a bath containing PEG-bisTz or a Tz-tagged zwitterionic peptide with alternating glutamic acid (E) and arginine (R) residues, poly(ER)-bisTz. While the HA particles readily took up the poly(ER) species, the loading of PEG-bisTz of similar molecular weights was minimal. Paradoxically, the avidity of poly(ER) for HA prevented its release and subsequent ligation with the TCO crosslinker at the particle-water interface. As an alternative strategy, calcium-crosslinked alginate particles were prepared in the presence of PEG-bisTz (7.5 kDa). The bisTz-loaded particles are subsequently soaked in a solution containing the TCO crosslinker to initiate the interfacial reaction. Hydrogel particles with an alginate core and HA shell were prepared using high molecular weight HA-TCO (450 kDa). When a low molecular weight HA-TCO (5 kDa) was used, inward diffusion of the TCO species resulted the creation HA-reinforced alginate particles with a gradient of crosslinking density. ☐ Overall, the novel diffusion-driven polymerization strategy was successfully applied to the synthesis of hydrogel microfibers that are cell-adhesive, mechanically robust and hydrolytically degradable. The diffusion-controlled method was further demonstrated in the fabrication of hydrogel particles with a core-shell structure. Importantly, changing the relative molecular weights of TCO and Tz species provides a means of controlling the diffusion direction, and thus the 3D pattern of the resultant hydrogel particles. Adaptation of this method to other materials with different composition and geometry will afford functional biomaterials with unique properties.
Description
Keywords
Biomaterial, Bioorthogonal, Hydrogel, Patterning, Polymer, Tetrazine ligation