UC Santa Barbara

This dissertation is divided into two main sections: the first focuses on retinal diseases and their associated in-vitro models, while the second explores advancements in cell therapies for treating retinal diseases. The eyes include the complex neurosensory system that facilitates our visual perception. The cells responsible for transmitting visual information from the eye to the brain include photosensitive rods and cones, interneurons, and ganglion cells, which operate in concert. This information is carried to the brain via the optic nerve bundle, located towards the back of the eye. The neural retina is situated above the pigmented monolayer, known as the retinal pigmented epithelium (RPE), which serves both as a protective barrier shielding the neural retina from adjacent blood vessels and as a support system. The RPE also provides nutrients and protection to maintain the health and functionality of nearby photoreceptors. Mutations affecting various cell types in the eye have been identified, and while our understanding of conditions such as Age-Related Macular Degeneration (AMD) and Retinitis Pigmentosa (RP) has significantly improved in recent years, some remain poorly understood. First, we will review the current landscape describing several inherited retinal dystrophies and relevant stem-cell-derived models (Chapter I). Second, we describe an in-vitro model of stem-cell derived RPE that recapitulates several features of Danon disease (DD), a multi systemic disease caused by 2 mutations in Lysosome Associated Membrane Protein 2 (LAMP2) that can affect a patient’s vision. We provide mechanistic insights on how loss of LAMP2 can impair RPE function and longevity (Chapter II). To address the growing number of patients with retinal diseases, multiple research groups have utilized stem cell biology to generate cell banks of affected cell types. The goal is to replace degenerative tissue with healthy and viable cells, offering potential treatment options for patients. One such project targets AMD and includes replacing dysfunctional RPE with healthy ones. Several challenges have emerged including long-term storage, production and scalability. Here we describe methods to produce viable and functional RPE cells at scales necessary to treat the afflicted population of individuals suffering from AMD (Chapter III). Additionally, we address storage difficulties by detailing methods to cryopreserve clinical grade RPE implants that yield viable and functional cells post thaw (Chapter IV). The final section of this dissertation provides insights into additional work and future directions presented herein (Chapter V). We build upon previous studies describing retinal diseases associated with several dystrophies, understanding the pathogenicity of DD associated vision loss and develop methodologies to improve cellular therapies targeting AMD