The Maturation of Human Pluripotent Stem Cell-Derived Retinal Ganglion Cells and Their Degeneration in Glaucoma
thesisposted on 2020-03-10, 16:03 authored by Kirstin VanderWallKirstin VanderWall
In glaucoma, the connection between the eye and the brain is severed leading to the degeneration of retinal ganglion cells (RGCs) and eventual blindness. A need exists to better understand the maturation of human RGCs as well as their degeneration, with the goal of developing new therapeutics for diseases like glaucoma. Human pluripotent stem cells (hPSCs) provide an advantageous model for the study of RGC development and disease as they can be differentiated into RGCs in large, reproducible quantities. Efforts of the current studies initially focused on the development and maturation of RGCs from hPSCs. RGCs derived from hPSCs were a diverse population of cells and matured in a temporal fashion, yielding morphological and functional characteristics similar to their in vivo counterpart. CRISPR/Cas9 gene editing was then utilized to insert the OPTN(E50K) glaucomatous mutation into hPSCs to model RGC degeneration. RGCs harboring this mutation exhibited numerous degenerative phenotypes including neurite retraction and autophagy dysfunction. Within the retina, many cell types contribute to the health and maturation of RGCs including astrocytes. As such, a co-culture system of hPSC-derived RGCs and astrocytes was developed to better understand the interaction between these two cell types. When grown in co-culture with astrocytes, hPSC-derived RGCs demonstrated significantly enhanced and accelerated morphological and functional maturation, indicating an important relationship between these cells in a healthy state. Astrocytes have also been shown to encompass neurodegenerative phenotypes in other diseases of the CNS, with these deficits profoundly effecting the health of surrounding neurons. xix Astrocytes grown from OPTN(E50K)-hPSCs demonstrated cell autonomous deficits and exhibited significant effects on the degeneration of RGCs. Taken together, results of this study demonstrated the utilization of hPSCs to model RGC maturation and degeneration in glaucoma. More so, these results are one of the first to characterize astrocyte deficits caused by the OPTN(E50K) mutation and could provide a new therapeutic target for pharmacological screenings and cell replacement therapies to reverse blindness in optic neuropathies.