Biofunctional peptide-click PEG-based hydrogels as 3D cell scaffolds for corneal epithelial regeneration

Poly(ethylene glycol) (PEG)-based hydrogels as highly promising 3D cell scaffolds have been widely implemented in the field of tissue regrowth and regeneration, yet the functionalized PEG hydrogel providing dynamic, cell-instructive microenvironments is inherently difficult to obtain. Here, we have exploited the specificity of click reaction to develop a set of hydrogels based on 4-arm PEG tetraazide (4-arm-PEG-N₃) and di-propargylated peptides (GRGDG and GRDGG) with tunable physicochemical properties applicable for 3D cell scaffolds. The azide groups of PEG were reacted with the alkynyl groups of peptides, catalyzed by copper to form triazole rings, thus generating a cross-linked hydrogel. The gelation time and mechanical strength of the hydrogels varied according to the PEG/peptide feed ratio. The resulting hydrogel exhibited a typical porous microstructure and suitable swelling behavior. The in vitro cytotoxicity test indicated that the resulting hydrogels did not cause apparent cytotoxicity against human corneal epithelial cells (HCECs). After co-incubation with HCECs, the density of RGD as well as peptide sequence in the hydrogels remarkably affected the cell attachment, spreadability, and proliferation. Additionally, the proposed hydrogel showed high ocular biocompatibility after being embedded subconjunctivally into rabbit eyes. Overall, these findings highlighted that the biofunctional hydrogels formed by PEG and RGD motifs via a controllable click reaction might be promising 3D cell scaffolds for corneal epithelial regeneration.