Photoactivated growth factor release from bio-orthogonally crosslinked hydrogels for the regeneration of corneal defects.

Abstract

<i>In situ</i>-forming hydrogels are an attractive option for corneal regeneration, and the delivery of growth factors from such constructs have the potential to improve re-epithelialization and stromal remodeling. However, challenges persist in controlling the release of therapeutic molecules from hydrogels. Here, an <i>in situ</i>-forming bio-orthogonally crosslinked hydrogel containing growth factors tethered <i>via</i> photocleavable linkages (PC-HACol hydrogel) was developed to accelerate corneal regeneration. Epidermal growth factor (EGF) was conjugated to the hydrogel backbone through photo-cleavable (PC) spacer arms and was released when exposed to mild intensity ultraviolet (UV) light (2-5 mW/cm², 365 nm). The PC-HACol hydrogel rapidly gelled within a few minutes when applied to corneal defects, with excellent transparency and biocompatibility. After subsequent exposure to UV irradiation, the hydrogel promoted the proliferation and migration of corneal epithelial cells <i>in vitro</i>. The rate of re-epithelialization was positively correlated to the frequency of irradiation, verified through <i>ex vivo</i> rabbit cornea organ culture studies. In an <i>in vivo</i> rat corneal wound healing study, the PC-HACol hydrogel exposed to UV light significantly promoted re-epithelialization, the remodeling of stromal layers, and exhibited significant anti-scarring effects, with minimal α-SMA and robust ALDH3A1 expression. Normal differentiation of the regenerated epithelia after healing was evaluated by expression of the corneal epithelial biomarker, CK12. The remodeled cornea exhibited full recovery of corneal thickness and layer number without hyperplasia of the epithelium.