Geldanamycin Treatment Reduces Neovascularization in a Mouse Model of Retinopathy of Prematurity

Abstract

Abstract: : Purpose: The benzoquinoid antibiotic geldanamycin has been identified as a potent inhibitor of pp60scr tyrosine kinase. A less toxic, potent, synthetic derivative 17–allylaminogeldanamycin (17–AAG) inhibits the Ras/Raf/MEK and PI3–Kinase signaling pathways and down–regulates vascular endothelial factor expression. Here we investigate the effect of 17–AGG on the development of the retinal vascular system in oxygen–induced retinopathy. Methods: C57/BL6 mice were exposed to 70% oxygen from postnatal day 7 (P7) to day 12 (P12) and recovered at room air thereafter. Beginning with day P12 mice were treated by daily i.p. injection of 17–AGG (12.5 mg/kg BW) microdispersed in an emulsion of 4% Lipoid EPC/5% sucrose in 0.9% NaCl or Wortmannin (100 µg/kg BW) for five days. The retinal vasculature was examined on day 17 and was quantified using a density slicing method evaluating the vasculature after perfusion with FITC–concanavalin A. Vascular and avascular area, blood vessel tufts, and main vessel tortuosity were quantified. The mRNA levels of VEGF were quantitatively analyzed by real–time RT–PCR. Results: The 17–AGG treatment after recovering the mice at room air resulted in a reduction of the vascular area from 36.8% to 30.8% (p=0.02), and in an increase of the avascular area from 7.1% to 20.3% (p=0.0001). Treatment with the PI–3 kinase inhibitor Wortmannin did not alter the vascular pattern compared to the controls. The area of blood vessel tufts and the main vessel turtuosity were not statistically different after treatment with either 17–AGG or Wortmannin. A reduction in VEGF mRNA expression could not be achieved with either of the treatments. Conclusions: We have shown that an i.p. injection of 17–AGG is capable to reduce angioproliferative retinopathy in a mouse model for oxygen–induced retinopathy. Our data support the idea that the mechanism does not involve a reduction of the VEGF mRNA level, and acts not by PI–3 kinase inhibition but via the Ras/Raf/MEK pathway. These data underscore the potential utility of tyrosine kinase inhibitors in oxygen–induced retinopathy.

Keywords: retinopathy of prematurity • retinal neovascularization • vascular cells