Reparative Outcomes in Corneal Infection: Linking Adjunctive Tβ4 Treatment to Nerve Regeneration and Visual Function

Purpose: Previous studies have shown that adjunctive thymosin beta-4 (Tβ4) with ciprofloxacin reduces bacterial keratitis severity, enhances wound repair, and promotes a return to homeostasis. However, its impact on corneal nerves and visual function, two critical but often overlooked determinants of long-term outcomes, remains unexplored. The present study addresses this gap by evaluating whether adjunctive Tβ4 supports nerve regeneration and restores visual function after infection. Methods: Bacterial keratitis was induced in C57BL/6 mice by inoculating wounded corneas with Pseudomonas aeruginosa American Type Culture Collection strain 19660. Topical treatments (PBS, Tβ4 monotherapy, ciprofloxacin monotherapy, adjunctive Tβ4 + ciprofloxacin) were administered three times daily, starting 24 hours post infection. Ocular disease severity was assessed by clinical scoring. Visual recovery was evaluated by measuring visual acuity (cycle/degree) and contrast sensitivity (%). Corneal sensitivity testing assessed nerve function, while β-III tubulin immunofluorescence was used to visualize and quantify corneal nerves in flat mounts. Results: Adjunctive Tβ4 + ciprofloxacin treatment markedly improved visual acuity and contrast sensitivity and significantly enhanced corneal sensitivity and nerve regeneration compared with PBS, Tβ4, and ciprofloxacin monotherapies. Combination therapy restored nerve density and architecture to levels comparable to those in uninfected controls. Conclusions: Adjunctive Tβ4 treatment promotes meaningful reparative outcomes, restoring both visual function and corneal nerve integrity during P. aeruginosa keratitis. By demonstrating that functional vision and nerve regeneration can be restored with combination therapy, this study highlights the importance of incorporating these end points into therapeutic evaluation and supports further development of Tβ4-based adjunctive strategies.