Elucidating the developmental and molecular mechanism of chorioretinal anastomoses in a model of type 3 neovascular age-related macular degeneration

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PROJECT SUMMARY Neovascular age-related macular degeneration (AMD) is a leading cause of vision loss and blindness in the elderly. Among the subtype of neovascular AMD, type 3 neovascularization (type 3 NV) is associated with a guarded visual prognosis and is uniquely characterized by vascular proliferation in both the retina and choroid. A defining feature of advanced type 3 NV is the development of chorioretinal anastomoses (CRA), abnormal direct connections between the retinal and choroidal circulations. Unfortunately, CRA can act as ‘hot spots’ for recurrent exudation which are resilient to available therapies. The developmental mechanism of CRA remains unclear. We established a murine model for type 3 NV, wherein mice harboring a mutation of type IV collagen (Col4a1) develop features of human disease including serous retinal detachment, hemorrhage, atrophy, and pronounced CRA. As in patients, CRA in Col4a1 mutant mice form in adulthood and mature into large de novo fusions between retinal and choroidal circulations, reminiscent of high-flow arteriovenous (AV) shunts. Recently, AV specification has emerged as a potent regulator of vascular patterning, and dysregulation of key genes governing AV identity induces formation of AV shunts. Notably, normalizing AV specification has been shown to reverse AV shunts despite their high-flow state in certain animal models. Elucidating a role for AV specification in CRA may offer novel insights into both disease pathogenesis and therapeutic strategies. Our central hypothesis is that CRA in Col4a1 mutant mice are AV shunts with a conserved developmental origin from either the retina or choroid. We have recently enabled multiphoton imaging of the choroid directly through the sclera of pigmented eyes using optimized multiphoton microscopy. The objectives of this inter-disciplinary research proposal are therefore to: 1) optimize intravital imaging of chorioretinal vascular dynamics with transscleral multiphoton microscopy; 2) determine the origin, blood flow, and arteriovenous identity of CRA in Col4a1 mutant mice; and 3) identify candidate pathways driving CRA in the endothelium using single cell RNA sequencing. The long-term goal of this career development research project is to provide the investigator with a cross- disciplinary skillset in advanced intravital imaging and single-cell transcriptomics that will allow him to investigate fundamental mechanisms of oculovascular dysgenesis toward the discovery of new therapies in vascular disease.