Genipin corneal crosslinking for the treatment of infectious keratitis in an ex vivo model.

Abstract

Purpose : Infectious keratitis is a major worldwide problem and an important cause of visual impairment and blindness. The increasing emergence of microbial resistance is a global concern and thus, there is an urgent need for the development of alternative therapeutic regimens. Corneal collagen crosslinking has been advocated as adjunctive therapy for the management of microbial keratitis. Genipin (GEN) is a natural plant extract whose properties as a crosslinking agent have been previously demonstrated. The purpose of this study was to investigate the effectiveness of GEN corneal crosslinking, for the treatment of infectious keratitis using an ex vivo pig corneal model designed for the study of bacterial keratitis.

Methods : Excised pig corneoscleral buttons maintained in organ culture. Corneas were infected using 10⁵ Staphylococcus aureus (ATCC 25923) or Pseudomonas aeruginosa (ATCC 27853). Eyes were kept in pairs and thirty-minute post-bacterial inoculation, one eye was treated with saline solution and the contralateral eye was treated with GEN (n=6 pairs for each microorganism). Corneas not exposed to bacteria were used as control. After 24h of incubation, corneas were processed for histological examination. Infected corneas were also homogenized and the resulting suspension was serially diluted and plated onto agar plates in order to determine and compare the colony-forming units (CFU)/cornea.

Results : S.aureus infected corneas showed a visible ina visible invasion in the cornea and bacterial colonies were visible on the corneal surface of all corneas, but were more evident in the corneas treated with saline solution than the corneas treated with GEN. P.aeruginosa treated corneas were also characterized by edema and expressed green fluorescent. Histologically, hematoxylin and eosin and Gram stains confirmed extensive bacterial infiltration throughout the cornea. Bacterial burden of the GEN-treated corneas was diminished and a significant decrease in the CFU/cornea was observed (p<0.05). Control corneas maintained uninfected.

Conclusions : A simple and reproducible ex vivo pig bacterial keratitis model was established and the antibacterial efficacy of GEN corneal crosslinking was evaluated. GEN corneal crosslinking could serve as a potential alternative therapeutic for infectious keratitis. Further studies are needed to fully elucidate the exact antibacterial activity and mechanism of action of GEN.

This is a 2020 ARVO Annual Meeting abstract.