Modeling neurodegenerative features of glaucoma using human pluripotent stem cell-derived retinal ganglion cells

Abstract

Purpose : Retinal ganglion cells (RGCs) connect the eye and the brain, allowing for visual perception, and these cells are known to be damaged in various blinding disorders such as glaucoma. The degeneration of RGCs is known to occur in a compartmentalized fashion, with the axonal compartment degenerating via different mechanisms than the somatodendritic compartment. While previous studies have demonstrated the ability to study certain aspects of RGC neurodegeneration with human pluripotent stem cells (hPSCs), they have lacked a focus upon the compartmentalized nature of RGC neurodegeneration. Thus, the goal of this study was to examine how RGC neurodegeneration occurs within different cellular compartments.

Methods : RGCs were differentiated from isogenic control and CRISPR/Cas9 edited OPTN(E50K) hPSCs. RGCs were enriched via Magnetic Activated Cell Sorting (MACS). Initially, RGCs were plated upon laminin-coated coverslips, and studies explored neurodegenerative features within RGCs over the first 4 weeks of maturation including neurite retraction and changes in excitability. Subsequently, RGCs were also plated onto microfluidics slides to allow for the recruitment of RGC axons away from the somatodendritic compartments, which allowed for an analysis of cellular changes within discrete compartments of RGCs.

Results : Glaucomatous OPTN(E50K) RGCs showed reduction in soma size and neurite outgrowth compared to isogenic controls, correlated with an enhanced excitability. Upon growth in microfluidic platforms, RGC axons and dendrites could be readily distinguished, and changes in the complexity of these processes were analyzed and compared for significant changes. Overall, OPTN(E50K) RGCs demonstrated morphological and functional deficits, mimicking some aspects of the phenotypes observed during the progression of glaucomatous neurodegeneration.

Conclusions : The results of this study demonstrate the ability to study neurodegenerative phenotypes in RGCs differentiated from human stem cells, including those with a glaucoma-associated OPTN(E50K) mutation along with the further application of microfluidic platforms that allows for the analysis of neurodegenerative changes in separate neuronal compartments. The results of this study will expand our understanding of the cellular changes that occur in human RGCs, and will support future studies including disease modeling and drug screening.

This is a 2021 ARVO Annual Meeting abstract.