The Inhibitory Effect Of Riluzole On Retinal Neovascularization

Objective: 1. To explore the inhibitory effects of riluzole on the proliferation of the human umbilical vein endothelial cell culture (HUVECs) induced by VEGF in vitro. 2. To establish the modified oxygen-induced retinopathy (OIR) mouse model, investigate the inhibitory effects of riluzole on the retinal neovascularization and explore the potential mechanism of its effects.Methods: 1.Human umbilical vein endothelial cell cultures were treated with VEGF to induce endothelial cell proliferation in the presence or absence of riluzole, using MTT assay observe the effect. 2. Twenty three-day-old neonatal C57BL/6 mice were fed with their nursing mothers (no less than two) until postnatal day (P) 7. From P7 to P12, mice were raised in a (75±2)% oxygen chamber for 5 days. After exposure to hyperoxia, mice were raised for another 5 days (P12-P17) in a room-air environment, thus set up the modified oxygen-induced retinopathy (OIR) model. Then those mice were divided into two groups randomly, the other ten mice of the same age were raised in room air as normal controls. Each mouse in the treatment group received an intraperitoneal injection of 10mg/kg of riluzole daily form P12-P17, and the control animals received volume and osmolarity-matched saline injection. At P17, all animals were killed and both eyes were enucleated, fixed with 4% paraformaldehyde, and embedded in paraffin, stained with HE. The proliferative neovascular response was quantified by counting the endothelial cell nuclei of new vessels beyond the inner limiting membrane of retina. The expression of retina PKC-βII and VEGF were detected with immunohistochemistry to investigate the protential mechanism of the inhibitory effects of riluzole.Results: 1. The proliferation of the HUVECs induced by VEGF was suppressed by riluzole 0.1-10μM。2. There was a mean of (0.28±0.66) neovascular nuclei per cross-section in the normal control compared to(31.14±10.00)nuclei per cross-section in the hyperoxia control group ( t=21.77,P<0.001 ), suggested the modified OIR model was successfully established. There was a mean of (31.14±10.00)neovascular nuclei per cross-section in the hyperoxia control compared to(6.50±3.26)nuclei per cross-section in the riluzole treated group (t=16.66,P<0.001), suggested that riluzole has significant inhibitory effects on retinal neovascularization. Immunohistochemistry of the retinal sections revealed PKC-βII and VEGF overexpression in the retina of the hypoxia control than the normal control and riluzole treated group, suggested that riluzole can successfully inhibit the expression of PKC-βII and VEGF in retina.Conclusions: Riluzole can inhibit the proliferation of the HUVECs induced by VEGF in vitro. Retinal neovascularization can be successfully inhibited by intraperitoneal injection of riluzole, the PKC-βII and VEGF expression in retina was partial inhibited. The mechanism of inhibitory effect of riluzole may contribute to inhibiting the activity of PKC-βII and thus the expression of some important growth factors, thus as VEGF. All suggested that riluzole may have potential therapeutic benefits in retinal vascular disease.