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Sarah Ohlemacher, Akshayalakshmi Sridhar, Yucheng Xiao, Mansoor Sarfarazi, Theodore Cummins, Jason S Meyer; Elucidating Retinal Ganglion Cell Development and Glaucoma Disease Modeling With Human Pluripotent Stem Cells. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3605.
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© ARVO (1962-2015); The Authors (2016-present)
The ability to derive retinal ganglion cells (RGCs) from human pluripotent stem cells (hPSCs) provides an unlimited supply of cells for the study of RGC development and disease, with important implications for drug screening, disease modeling, and cell replacement strategies. However, the ability to derive RGCs from hPSCs has been largely overlooked to date. Thus, the ability of hPSCs to yield a population of RGCs was explored. Additionally, induced pluripotent stem cells (iPSCs) were generated from a glaucoma patient exhibiting an E50K mutation in the Optineurin (OPTN) gene, associated with RGC degeneration leading to normal tension glaucoma. These cells were differentiated to an RGC fate and assayed for phenotypic differences compared to wild type iPSCs.
hPSCs were differentiated to a retinal fate following previously established protocols and RGCs were subsequently characterized for the expression of RGC related features. Fibroblasts were obtained from a glaucoma patient exhibiting an E50K mutation in the OPTN gene, and these fibroblasts were used to generate patient-specific OPTN iPSCs via mRNA reprogramming. OPTN iPSCs were differentiated in a stepwise fashion that allowed for the definitive identification of RGCs based upon gene expression patterns. Phenotypic differences between wild type and OPTN iPSC-derived RGCs were explored, including changes in golgi fragmentation, apoptosis, and autophagy.
Within 40 days of differentiation, RGCs could be readily identified due to the expression of RGC-associated markers as well as their documented transition through a highly enriched retinal progenitor stage. Analysis of hPSC-derived RGCs revealed that these cells expressed numerous morphological, phenotypical, and physiological characteristics of RGCs. In addition, OPTN iPSC-derived RGCs were found to display striking phenotypic differences compared to wild type cells.
The data presented demonstrates the ability of hPSCs to serve as a reliable source of RGCs, including those derived from glaucoma patient sources, as seen by their ability to yield a population of cells possessing a full complement of RGC-associated characteristics. These results will facilitate future studies into the disease-related degeneration of RGCs and as such, will be instrumental as a tool for the study of optic neuropathies, as well as the development of therapeutic approaches.
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