Rock Inhibitor Eye Drops Accelerate Corneal Endothelium Wound Healing In A Primate Model

Abstract

Purpose: : We previously reported that Rho kinase (ROCK) inhibitor Y-27632 promotes wound healing of corneal endothelial cells (CECs) in a rabbit model and showed the possibility of the clinical application for humans who are in a corneal endothelial deficient condition (BJO, 2011). The purpose of this present study was to demonstrate the effect of Y-27632 on wound healing in a primate corneal endothelial dysfunction model.

Methods: : As a corneal endothelial partially-injured model, the corneal endothelium of 6 cynomolgus monkeys (3-5 years-of-age; estimated equivalent human age: 15-20 years) was damaged by transcorneal freezing with a 7-mm-diameter probe. Ten mM of Y-27632 was then applied topically in 1 eye of each animal 6 times daily, while phosphate buffered saline was applied in the fellow eye as a control. The corneal appearance of each eye was then examined by slit-lamp microscopy, and in vivo corneal endothelial cells were examined by noncontact specular microscopy. Four weeks after treatment, the phenotype of the reconstructed corneal endothelium was evaluated by scanning electron microscopy and by immunohistochemical analysis of ZO-1 and Na⁺/K⁺-ATPase.

Results: : Slit-lamp microscopy revealed that both Y-27632 treated and non-treated corneas became hazy after transcorneal freezing, and then recovered their transparency within 4 weeks. Noncontact specular microscopy revealed that corneal endothelial cell density was significantly higher in the Y-27632 group compared to the controls at 4 weeks (3123.7±82.4 and 1833.0±90.8 cells/mm², respectively; p<0.01). Although fairly normal appearing same-size hexagonal cells were observed in the Y-27632 treated eyes, a big variation in cell size and poorly formed cell junctions were observed in the controls by electron microscopy. The percentage of ZO-1 and Na⁺/K⁺-ATPase positive cells among the regenerated area in the Y-27632 group was significantly higher than in the controls (98.7±1.3 vs 48.7±7.6%; p<0.05, and 100.0±0.0 vs 76.6±2.0%, respectively; p<0.01).

Conclusions: : ROCK inhibitor Y-27632 promotes recovery of corneal endothelial cell density and wound healing in terms of both morphology and function. These findings indicate that ROCK inhibitor eye drops might hold the promise of becoming a new pharmaceutical agent for treating corneal endothelial dysfunction.

Keywords: cornea: endothelium • cornea: basic science • drug toxicity/drug effects