Alkaline Phosphatase-Triggered Spatiotemporal Repair of Corneal Injury with TB500 Peptide Hydrogel

Corneal injury remains a significant clinical challenge due to the limited regenerative capacity of the cornea and the difficulties associated with maintaining drug retention at the injury site. This study presents a novel spatiotemporal repair strategy for corneal wounds, utilizing an alkaline phosphatase (ALP)-triggered, lesion-responsive peptide hydrogel that incorporates TB500, a biologically active peptide with the amino acid sequence LKKTETQ, which has not been previously explored for corneal disease treatment. The hydrogel is designed through enzyme-instructed self-assembly (EISA) of a phosphorylated peptide precursor, Nap-YpYY-TB500, which undergoes site-specific dephosphorylation by elevated ALP levels at the wound site, triggering nanofiber formation and gelation in situ. Among three candidate sequences, Nap-YpYY-TB500 exhibited optimal gelation kinetics, nanostructure, and therapeutic efficacy. In vitro, the hydrogel promoted human corneal epithelial cell (HCEC) migration, proliferation, and tight junction recovery, while also enhancing myofibroblastic differentiation and cytoskeletal reorganization of human corneal stromal fibroblasts (HCSFs). In an alkali burn model, the hydrogel significantly accelerated epithelial regeneration, reduced inflammation, and improved corneal barrier function. Our work represents the first ocular application of TB500 and underscores the potential of enzyme-responsive, self-assembling peptide hydrogel as a localized and sustained delivery system for corneal repair.