Abstract
Purpose :
To determine the optimal concentration of nanoparticle encapsulated FL2 siRNA (FL2-NP-si), a novel drug targeting the Fidgetin-like 2 (FL2) gene, for the treatment of corneal alkaline chemical injury.
Methods :
Eighty 12-week old, male Sprague-Dawley rats were divided evenly into eight groups: prednisolone, empty nanoparticles, control-NP-si (1 µM, 10 µM, 20 µM) and FL2-NP-si (1 µM, 10 µM, 20 µM). After anesthesia, corneal epithelium was removed and 4 mm discs of 1M NaOH-soaked filter paper were applied for 10 seconds to induce chemical injury to the cornea. Eyes were then treated and clinically assessed for time to re-epithelialization and degree of corneal opacity and hemorrhage, every other day (N=10 per group) for 14 days. At the end of the treatment period, animals were euthanized and eyes collected for histopathologic (N=6 per group) and immunohistochemical (N=4 per group) evaluation, staining for neutrophils, macrophages, and vascular endothelial cells. A qualitative scoring system ranging from 0-4 was used for both clinical and histopathological assessment of opacity, edema, inflammation, neovascularization, and presence of retrocorneal membrane. Nanoparticle-mediated knockdown of FL2 was confirmed by an in vitro qPCR assay. Toxicity assays were performed to assess for apoptosis (TUNEL assay, N=3 per group) and nerve damage (whole mount immunochemical staining, N=2 per group). Statistical analyses were performed using student’s t-test and ANOVA.
Results :
Compared to controls, FL2-treated groups demonstrated enhanced corneal wound healing across a number of parameters. Specifically, the 20 µM FL2-treated group demonstrated the greatest rate of corneal re-epithelialization (p=0.0003 at Day 4 and p=0.0005 at Day 8) as assessed with ImageJ software; enhancement in corneal transparency; histologic improvement with reduction in inflammation, neovascularization, edema, and retrocorneal membrane. Immunohistochemical analysis did not exhibit significant differences between groups. FL2-siRNA was not found to be toxic to nerves or induce apoptosis (p=0.917).
Conclusions :
Based on dose-response, 20 µM FL2-NP-si was found to be the optimal concentration in this rat alkali corneal burn model. FL2-NP-si may offer a novel treatment for alkaline chemical injuries to the cornea.
This is a 2020 ARVO Annual Meeting abstract.