VEGF Is Required for Neuroretina Survival and Function

Abstract

Purpose: : Agents targeting VEGF have led to significant vision improvements in many patients with wet macular degeneration. However, the role of VEGF in the adult retina remains undefined. The goal of these studies was to investigate the effect of VEGF neutralization on the structure and function of the normal, adult retina.

Methods: : VEGF and its receptors were localized in adult retina using VEGF-lacZ/+ and VEGFR2-lacZ/+ mice as well as by immunohistochemistry. Expression of VEGF in isolated Müller cells was examined by PCR and western-blot analysis. VEGFR2 activation was examined by immunoprecipitation with antiVEGFR2 followed by Western blotting with anti-phosphotyrosine. Systemic VEGF neutralization was accomplished by adenoviral delivery of sFlt1. Vascular changes were assessed by fundus angiography, FITC-dextran perfusion and TEM. Retinal cell apoptosis was determined by TUNEL and the thickness of the inner nuclear (INL) and outer nuclear layers. Visual function was measured by electrogram (ERG). A role for VEGF autocrine signaling in Müller cell survival was investigated in vitro using VEGF siRNA in Müller cells (M10M1).

Results: : VEGF was expressed by astrocytes and pericytes in the ganglion cell layer and in the INL, and VEGFR2 was demonstrated to be phosphorylated in the adult retina by immunoprecipitation with anti-VEGFR2 and blotting with anti-phosphotyrosine. After 14 days of VEGF neutralization, while there was no detectable effect on the vasculature of the INL, there was marked apoptosis of cells in the INL and photoreceptors, leading to reduced inner and outer nuclear layer thickness. Four weeks of VEGF neutralization led to visual dysfunction as evidenced as reduced a- and b-waves in ERGs. siRNA suppression of VEGF expression in Müller cells led to the induction of the pro-apoptotic gene bax and increased apoptosis.

Conclusions: : These data implicate VEGF in the maintenance of neural retina structure and function, and point to a need for caution in the prolonged use of intraocular anti-VEGF therapy.

Keywords: vascular endothelial growth factor • retina • pathology: experimental