ROCK-2 Inhibition Supresses Choroidal Neovascularization

Abstract

Purpose: : Choroidal neovascularization (CNV) is the main cause of severe vision loss in patients with age-related macular degeneration (AMD). Previously we showed that rho kinase (ROCK) inhibition with fasudil reduced CNV size. However, the role of the ROCK isoforms, ROCK-1 and ROCK-2, in AMD is unknown. A novel specific ROCK-2 inhibitor (Surface Logix) was used to address this question.

Methods: : Animals were anesthetized and the pupils were dilated. CNV was induced with an argon laser. Four laser spots were placed in each eye using a slit lamp delivery system and a cover glass as a contact lens. The inhibitor or vehicle was injected intraperitoneally twice a day in three doses (1mg/kg, 10mg/kg, 30mg/kg).One week after laser injury animals were perfused with 10ml PBS and fluorescein-labeled dextran and flatmounts were prepared. The hyperfluorescent area, showing the CNV lesions, were measured with Openlab software.Inhibitors or vehicle were injected intravitreally using an Exmire microsyringe on days 0, 3, 7 after CNV induction. Vascular leakage from the CNV lesions was assessed using fluorescein angiography (FA), using a digital fundus camera.

Results: : Seven days after laser injury, systemic treatment with 10mg/kg (13000±2396µm², n=8) and 30mg/kg (9890±2926µm², n=8) of the ROCK-2 inhibitor caused a significant reduction in CNV size, compared with vehicle-treated controls (21471±1680µm², n=8, P=0.005, P=0.0008) and low dose ROCK-2 inhibitor (1mg/kg, 21972±3014µm², n=8). Intravitreal injection of fasudil or the ROCK-2 inhibitor one week after laser injury caused significantly smaller lesions than the vehicle-treated group (29818±4035µm², 33406±4325µm² vs 58620± 2642µm², n=16, P=0.00003, P=0.0002).FA showed a significantly reduced incidence of the clinically relevant grade IIB lesions with ROCK-2 inhibition (53.1%, n=8), compared with the vehicle-treated animals (84.4%, n=8, P <0.05).

Conclusions: : ROCK-2 inhibition effectively suppresses laser-induced CNV and leakage. Our results indicate that ROCK-2 may become an attractive molecular target in the prevention and treatment of AMD.

Keywords: choroid: neovascularization • age-related macular degeneration