The effects of FK506 combined with natamycin in the treatment of experimental fungal keratitis by suppressing NLRP3 inflammasome activation

Abstract

The aim of this study was to investigate the mechanisms of combination treatment with FK506 and natamycin on alleviating damage of the cornea in mouse model of fungal keratitis. In this study, the mouse model of fungal keratitis was created by intrastromal injection with Fusarium solani or Aspergillus flavus. The mice received 5% natamycin eye drops 6–8 times a day, or the mice received 0.05% FK506 eye drops 2 times per day for 21 consecutive days. Corneal damage was evaluated by H&E staining. The protein expression levels of NLRP3 were detected by immunohistochemistry. Moreover, the markers of inflammasome activation including NLRP3, ASC, caspase-1, IL-1β, and IL-18 were detected by western blot. Histopathological results showed increased corneal thickening, dense inflammatory cell infiltration, and loss of epithelial continuity in the corneas after fungal infection. In addition, NLRP3 positive signals were observed to be obviously increased in the corneas after A. flavus or F. solani infection compared to the control group. Furthermore, the NLRP3 inflammasome is induced by fungal infection, as evidenced by increased protein expression levels of NLRP3, ASC, caspase-1, and downstream cytokines, such as interleukin (IL)-1β and IL-18. However, the corneal damage was alleviated and the activation of the NLRP3 inflammasome was significantly inhibited by drug treatment. Besides, the treatment outcomes were better in combined treatment group than that in single-agent treatment group. In conclusion, FK506 combined with natamycin alleviate fungi-induced corneal damage by suppressing NLRP3 inflammasome activation.