Retinal Ganglion Cells With a Glaucoma OPTN(E50K) Mutation Exhibit Neurodegenerative Phenotypes when Derived from Three-Dimensional Retinal Organoids

Abstract

Summary Retinal ganglion cells (RGCs) serve as the connection between the eye and the brain, with this connection disrupted in glaucoma. Numerous cellular mechanisms have been associated with glaucomatous neurodegeneration, and useful cellular models of glaucoma allow for the precise analysis of degenerative phenotypes. Human pluripotent stem cells (hPSCs) serve as powerful tools for studying human disease, particularly cellular mechanisms underlying neurodegeneration. Thus, efforts focused upon hPSCs with an E50K mutation in the Optineurin (OPTN) gene, a leading cause of inherited forms of glaucoma. CRISPR/Cas9 gene editing introduced the OPTN(E50K) mutation into existing lines of hPSCs, as well as generating isogenic controls from patient-derived lines. RGCs differentiated from OPTN(E50K) hPSCs exhibited numerous neurodegenerative deficits, including neurite retraction, autophagy dysfunction, apoptosis, and increased excitability. These results demonstrate the utility of OPTN(E50K) RGCs as an in vitro model of neurodegeneration, with the opportunity to develop novel therapeutic approaches for glaucoma.

Retinal organoids: a window into human retinal development

“…Recently, genome-edited cell lines have been used to generate models of retinal disease in the dish ( Capowski et al, 2014 ; Zheng et al, 2020 ). Patient-derived iPSC lines with disease-causing mutations have been corrected to create ideal isogenic control lines that can be differentiated in parallel to validate disease phenotypes ( Lam et al, 2020 ; Lane et al, 2020 ; VanderWall et al, 2020 ). Edited cell lines are also valuable to test novel regulators of development and disease ( Buskin et al, 2018 ; Deng et al, 2018 ; Eldred et al, 2018 ; Phillips et al, 2014 ).…”

Advancing Drug Discovery for Neurological Disorders Using iPSC-Derived Neural Organoids

“…In parallel to therapy testing, neuroophthalmological disease modeling profited from CRISPR-Cas as well. The introduction of the E50K mutation in the optineurin (OPTN) gene enabled the modeling of glaucoma in retinal organoids [ 79 ] and the knockout of RB1 demonstrated its crucial role in retinal development [ 80 ].…”

Protect, Repair, and Regenerate: Towards Restoring Vision in Glaucoma

“…Organoid-derived RGCs also exhibit photoreceptor-driven action potentials comparable to the earliest light responses recorded from the neonatal mouse retina [ 67 ]. Planar-derived RGCs are transdifferentiated from pluripotent stem cells in culture [ 68 ], and after purification in culture have similar electrophysiological properties to native RGCs; hPSC-RGCs demonstrate characteristic spontaneous and current-evoked activity, indicating functional axons; however, they do not develop the ability to form circuitry as RGCs in organoids do [ 69 – 71 ]. Co-culture of hPSC-RGCs with other cell types improves dendritic complexity and electrophysiological responses [ 72 ].…”