Theranostics of Photoreceptor-RPE-Choroid Neurovascular Unit in Eye Diseases:

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TITLE: Theranostics of Photoreceptor-RPE-Choroid Neurovascular Unit in Eye Diseases PI: Robert J. Zawadzki, Ph.D. SUMMARY Inherited and age-related macular degeneration (AMD) are currently responsible for severe vision impairment in over 2 million US residents, with prevalence expected to double by 2040 as the population ages. Degeneration occurs in photoreceptor cells, retinal pigment epithelial (RPE) cells, and the choroidal vasculature, a complex of tightly interdependent tissues in the posterior eye. This project will continue our in vivo investigations of morphology and function of the photoreceptor-RPE-choroid complex in animal models of retinal degenerations, including the RPE mitochondrial dysfunction model and Doyne Honeycomb Retinal Dystrophy. These models recapitulate major hallmarks of inherited and age-related macular degeneration: degeneration of RPE cells and age-related increase in extracellular deposits between the RPE and Bruch’s membrane, which separates the RPE from the choroidal capillary bed. The project will use innovative, cellular- level resolution in vivo probing of retinal structure and function combined with additional functional tests to characterize age-related changes in major components of the photoreceptor-RPE-choroid neurovascular unit (PRC-NVU). These studies will be performed longitudinally in cohorts of mice with genetic defects and wild- type controls, fed with a high-fat diet (to accelerate disease progression). The studies will test the hypothesis that primary defects in RPE-Bruch’s membrane cause secondary deterioration of photoreceptors and choriocapillaris vasculature. The studies include measures of photoreceptor structure and function (using Temporal Speckle Averaging - Optical Coherence Tomography (TSA-OCT), OCT-based Optoretinography (ORG)), and full-field Electroretinography (ffERG)), mapping of RPE mosaic and Bruch’s membrane morphology with water driven transient changes in BrM scattering, RPE cell autofluorescence intensity and emission spectrum, the redox status of RPE cells, and choriocapillaris vascular morphology using TSA-OCT Angiography. We will analyze these multidimensional longitudinal data to discover potential structural and functional biomarkers of PRC-NVU degeneration and evaluate its predictive value for disease progression. At the termination of in vivo experiments, the choroid-RPE from one eye of each mouse will be imaged with a custom high-resolution ex vivo confocal microscope equipped with an Optical Coherence Microscopy (OCM) unit and a LiveCell stage top incubation system allowing imaging of a “live” RPE cells flat-mounted with the sclera (to allow validation of in vivo observations and evaluation of Fluorescence Lifetime of RPE organelles); the retina from the second eye of each mouse will be evaluated by conventional histological and biochemical measurements performed on light or electron microscopes, to allow validation of morphological in vivo findings. A novel method for delivering drugs to the RPE via near-infrared light-degradable nanoparticles will be used to locally target therapeutic agents to ailing RPE cells, and spatially resolved imaging will be used to determine if the therapeutic agents slow and stop disease progression. By combining longitudinal, in vivo imaging and optical nanotherapies, these studies will lay a foundation for locally targeted drug delivery in human ocular disease.