Purpose To determine whether genipin (a natural crosslinker) could reduce the colonization and proliferation of bacteria and fungi in an ex vivo model of corneal infection. Methods This study, using an ex vivo model of bacterial and fungal keratitis, investigated the antimicrobial efficacy of genipin crosslinking. Excised corneoscleral buttons were wounded by scalpel incision and subsequently infected with Staphylococcus aureus , Pseudomonas aeruginosa , or Candida albicans . After inoculation, corneas were treated with genipin for 24 hours at 37°C. Histologic examinations were carried out, and the number of viable colony-forming units (CFU)/cornea was determined. Results Genipin exerts bactericidal action against S. aureus and P. aeruginosa , as well as fungicidal action against C. albicans and significantly reduced the CFU compared to contralateral eyes that received saline treatment ( P < 0.05). Conclusions These data identify genipin as a novel ocular antimicrobial agent that has the potential to be incorporated into the therapeutic armamentarium against microbial keratitis. Translational Relevance This study provided evidence for the antimicrobial and antifungal properties of genipin as an alternative crosslinker that could be used in the management of infectious keratitis.

Infectious keratitis is a vision-threatening microbial infection. The increasing antimicrobial resistance and the fact that severe cases often evolve into corneal perforation necessitate the development of alternative therapeutics for effective medical management. Genipin, a natural crosslinker, was recently shown to exert antimicrobial effects in an ex vivo model of microbial keratitis, highlighting its potential to serve as a novel treatment for infectious keratitis. This study aimed to evaluate the antimicrobial and anti-inflammatory effects of genipin in an in vivo model of Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa) keratitis. Clinical scores, confocal microscopy, plate count, and histology were carried out to evaluate the severity of keratitis. To assess the effect of genipin on inflammation, the gene expression of pro- and anti-inflammatory factors, including matrix metalloproteinases (MMPs), were evaluated. Genipin treatment alleviated the severity of bacterial keratitis by reducing bacterial load and repressing neutrophil infiltration. The expression of interleukin 1B (IL1B), interleukin 6 (IL6), interleukin 8 (IL8), interleukin 15 (IL15), tumor necrosis factor-α (TNF-α), and interferon γ (IFNγ), as well as MMP2 and MMP9, were significantly reduced in genipin-treated corneas. Genipin promoted corneal proteolysis and host resistance to S. aureus and P. aeruginosa infection by suppressing inflammatory cell infiltration, regulating inflammatory mediators, and downregulating the expression of MMP2 and MMP9.

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