Synthesis and orthogonal photopatterning of hyaluronic acid hydrogels with thiol-norbornene chemistry.

The patterning of chemical and mechanical signals within hydrogels permits added complexity towards their use as cell microenvironments for biomedical applications. Specifically, photopatterning is emerging to introduce heterogeneity in hydrogel properties; however, currently employed systems are limited in the range of properties that can be obtained, as well as in decoupling mechanical properties from changes in chemical signals. Here, we present an orthogonal photopatterning system that utilizes thiol-norbornene chemistry and permits extensive hydrogel modification, including with multiple signals, due to the number of reactive handles accessible for secondary reaction. Hyaluronic acid was functionalized with norbornene groups (NorHA) and reacted with di-thiols to create non-toxic hydrogels with a wide range of mechanical properties. For example, for 4 wt% NorHA at 20% modification, hydrogel mechanics from ≈ 1 kPa up to ≈ 70 kPa could be obtained by simply changing the amount of crosslinker. By limiting the initial extent of crosslinking, NorHA gels were synthesized with remaining pendent norbornene groups that could be reacted with thiol containing molecules in the presence of light and an initiator, including with spatial control. Secondary reactions with a di-thiol crosslinker changed mechanical properties, whereas reaction with mono-thiol peptides had no influence on the gel elastic modulus. This orthogonal chemistry was used sequentially to pattern multiple peptides into a single hydrogel, demonstrating the robustness of this system for the formation of complex hydrogels.