Abstract
Abstract: :
Purpose: To investigate the possible mechanism of retinal ganglion cell death in a mouse model of elevated intraocular pressure (IOP). Methods:Elevated IOP was induced in one eye of each adult C57BL/6J mouse by argon laser photocoagulation of the episcleral and limbal veins. The intraocular pressure of both eyes of each mouse was measured using an indentation tonometer prior to treatment and once per week thereafter. The survival of RGCs was measured by counting the number of RGCs that were labeled retrogradely by DiI applied to the superior colliculus (SC). The reduction in the number of RGCs was compared in the experimental and normal eyes. The mechanism of RGC death after IOP elevation was investigated using TdT–mediated dUTP nick end labeling (TUNEL) staining, immunostaining for cleaved Caspase–3, and Western blot for Bcl–2 and Bax expression. RT–PCR was performed to measure Bcl–2, Bax, Bad, Bak, P53, ICE and Fas. Results: After laser treatment, mean IOP was increased in the treated eyes from the control mean of 13±1.8 mm Hg to 20.0 ± 2.8mm Hg at 4 weeks and 17±2.2 mmHg at 8 weeks. Retinal ganglion cell loss was 16.9 ± 7.8% at two weeks (n = 6, P<0.05) and 22.4% ± 7.5 at 4 weeks (n = 6, P<0.05) after laser photocoagulation. TUNEL staining showed a marked increase in the number of apoptotic nuclei in the ganglion cell layer (GCL) in the treated eyes. Caspase–3 positive cells were observed primarily in the GCL in treated eyes and seldom found in the control eyes. Higher levels of P53, IFN, Bak and Bax were found in eyes with elevated IOP than those in the control group. The expression of the anti–apoptotic genes Bcl–2 and Bad were higher in control group. Conclusions: This model of photocoagulation of the limbal and episcleral vessels in mice results in a consistent, sustained increase in IOP with a reduction in the number of RGCs labeled by DiI in the treated eye. There were more TUNEL–positive RGCs in eyes with elevated IOP. Increased P53, caspase–3, Bax, Bak, IFN, and decreased Bcl–2 were also demonstrated in eyes with elevated IOP, consistent with apoptosis as the mechanism of neuronal cell death.
Keywords: apoptosis/cell death • ganglion cells • intraocular pressure