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R. Yang, G.W. McCollum, D.P. Bingaman, J.S. Penn; The Effect of Anecortave Acetate on VEGF Message and Protein Levels in Hypoxic Müller Cells and in Rat OIR . Invest. Ophthalmol. Vis. Sci. 2005;46(13):4177.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:Anecortave acetate (AA) inhibits retinal neovascularization (NV) in a rat model of oxygen–induced retinopathy (OIR) with concommitant upregulation of retinal plasminogen activator inhibitor (PAI)–1. In the present study, using both in vivo and in vitro models, we investigate another mechanism of AA angiostatic action: an effect on VEGF induction by hypoxia. Methods: In vivo studies employed the rat model of OIR. Retinas from vehicle–injected or AA–injected (10% suspension) eyes were harvested 0, 1, 2, 4 or 6 days post–oxygen exposure. Additionally, rat Müller cells were grown to 70–80% confluency and treated with vehicle or 0.1µM–10µM AA under hypoxic conditions (< 2% oxygen) for 24 hr. VEGF concentrations in whole retinal lysates and in culture medium were measured by ELISA. Retinal VEGF mRNA levels were measured by real–time RT–PCR. Results: Retinal VEGF protein levels ranged from 223.68 to 374.58 pg/mg at various times post–exposure for vehicle– injected eyes and 207.24 to 266.34 pg/mg for AA–injected eyes. There was a significant decrease in retinal VEGF of AA–treated eyes at 1 and 2 days post–exposure. There was a significant 1.8–fold decrease of VEGF mRNA in AA–treated vs. vehicle–treated eyes at 6–days post–oxygen exposure. Conditioned medium from hypoxic Müller cells showed a 2–fold increase in VEGF concentration compared to medium from cells cultured in normoxia. VEGF concentration was reduced by 18% and 28% in hypoxic Müller cells treated with 1.0 and 10µM AA, respectively. Conclusions: AA–injection reduces VEGF protein levels in OIR rats at 1 and 2 days post–oxygen exposure providing yet another plausible mechanism for AA inhibition of NV. Furthermore, in vitro experiments suggest that this may result from the direct action of AA on hypoxic Müller cells. However, based on this study, there does not appear to be a simple relationship between VEGF mRNA and protein expression. Future studies designed to quantify specific VEGF mRNA splice–variants are warranted.
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