Conditional ablation of VEGFR-1 in photoreceptors induces retinal angiomatous proliferation: A transgenic mouse model of AMD

Abstract

Purpose: To determine whether retinal angiomatous proliferation (RAP, a subtype of age related macular degeneration, or AMD) is developed spontaneously without mechanical injury (e.g. subretinal injection) by conditional ablation of VEGF receptor-1 (VEGFR-1, or Flt-1) in the murine photoreceptors.

Methods: We interbred iCre-75+ mice (Cre recombinase expressed specifically in phtotoreceptors) with floxed Flt-1 mice. Cre expression in above Cre lines was identified by interbreeding with mT/mG mice expressing tomato/EGFP in all tissues. At 21days to 3 months after born, the fundi were observed in vivo by fluorescein angiography (FA) and indocyanine green (ICG) angiography using the Heidelberg Retina Angiograph. Histology was performed by hematoxylin and eosin (H&E) stainining and Transmission electron microscope (TEM). Cre expression in the iCre-75+ lines was identified by interbreeding with mT/mG mice expressing tomato/EGFP in all tissues; The sFlt-1 expression was analyzed by immunohisochemistry (IHC) and in situ hybridization.

Results: Cre expression, as indicated by tomato deletion resulting in exclusive Egfp fluorescence, was restricted to photoreceptors in floxed mT/mG mice crossed with the iCre-75+ line. At 1-3 months of age, both homozygous (iCre-75+ flt-1lox/lox, 8/14 eyes, 57%, P<0.05) and heterozygous (13/24 eyes, 54%, P<0.05) photoreceptor-specific Flt-1 knockout mice (iCre-75+ flt-1lox/+) developed RAP, compared with 10% (1/10 eyes) of littermate controls (iCre-75+ flt-1+/+). RAP started from the inner retina (arising from retinal vessels) at early stage and invaded the photoreceptor layer, eventually anastomosing with the choroidal neovasculature.

Conclusions: FLT-1 knockdown in the photoreceptor layers by selective Cre/lox FLT-1 ablation can induce RAP in the early stage. The transgenic mice were developed in this study could be potentially used as a murine RAP model.

Keywords: 412 age-related macular degeneration • 609 neovascularization