The Effect And Mechanism Of Genistein On Retinal Neovascularization

Retinal neovascularization occurring as a complication of diabetic retinopathy, central retinal vein occlusion and age-related macular degeneration can cause severe vision loss and blindness. Though pathogenesis led to neovascularization is still unclear, it probably results from retinal ischemia and/or hypoxia. Recent studies have implicated that VEGF overexpression is sufficient to cause intraretinal and subretinal neovascularization.Genistein, a soy derived isoflavone, has recently attracted much attention of the medical scientific community. This compound was found to be a potent agent in both prophylaxis and treatment of cancer as well as other chronic diseases. The effects of genistein include inhibition of angiogenesis, tyrosine kinase phosphorylation, DNA synthesis and cell cycle arrest. But the inhibitory effects of genistein on retinal neovascularization were unknown.The quantifying mouse model of oxygen-induced retinal neovascularization that was established by smith L was suitable for the study of pathogenesis and therapeutic intervention for the retinal neovascularization. One-week-old (P7) C57BL/6 mice were exposed to 75% oxygen for 5 days and then to room air. At PI4, genistein (0,50,100,200mg/kg weight body) treatment was begun. Mice of the same age kept in room air as controls. The proliferated neovascular response was quantitated by counting the nuclei of endothelial cells of new vessels extending from the retina into the vitreous in 6 u m sagittal cross sections. VEGFand hypoxia-induced factor lα (HIF-lct) were determined on the cross-sections after immunofluorescence stain. That were found are a mean of 23.92 ?4.43 neovascular nuclei per cross-section in the oxygen-treated retina, 20.92 ?4.70 nuclei in the 50mg/kg weight body genistein-treated retinal, 17.16 ?.98 nuclei in the l00mg/kg weight body genistein-treated retinal, 14.2 ?3.42 nuclei in the 200mg/kg weight body genistein-treated retinal, and less than 1 nucleus in the control group. Oxygen-treated mouse developed severe retinal hypoxia, dramatic up-regulation of retinal neovascularization. Genistein inhibited hypoxia induced retinal neovascularization in a concentration-dependent manner. VEGF and HIF-1α stain were found stronger in the inner retinal layer of oxygen-treated mouse than in that of controls, and these were found weaker in the inner retinal layer of genistein-treated mouse than in that of oxygen-treated mouse.In order to explore mechanism of genistein inhibited effects, the time course changes of VEGF protein expression induced by cobalt chloride (CoCl2) and hypoxia and the effects of genistein on CoCl2- and hypoxia-induced VEGF expression in rabbit retinal pigment epithelium (RPE) cells were studied. Judged by relativefluorescence using a confocal scanning laser microscope coupled to a computer, VEGF protein expression exposed for different periods to CoCl2 or hypoxia was investigated. CoCl2 was found to significantly elevate VEGF protein expression. At 4 h after CoCl2 treatment, the expression of VEGF protein was about three times as much as that at the start of treatment. Genistein (50, 100 and 200 u M) inhibited VEGF protein expression elicited by CoCl2 in a concentration-dependent manner. Hypoxia (5% CO_{22/95% N2} could markedly increase VEGF protein expression. The elevationof VEGF protein expression was gradual and time-dependently. At 6 h, the highest expression of VEGF protein was observed, it was about three times as much as that at the start of treatment. After preincubation with 50, 100 and 200 u M genistein respectively, the hypoxia-evoked VEGF expression wasconcentration-dependently suppressed.These results indicated that genistein could be an effective agent in the prevention and treatment of intraretinal and subretinal neovascularization. The mechanism may involve in inhibition of VEGFandHIF-lα.