Browsing by Subject "stem cell"

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    Analysis of retinal ganglion cell development: from stem cells to synapses
    (2018) Ohlemacher, Sarah K.; Meyer, Jason S.
    Human pluripotent stem cells (hPSCs) have the ability to self renew indefinitely while maintaining their pluripotency, allowing for the study of virtually any human cell type in a dish. The focus of the current study was the differentiation of hPSCs to retinal ganglion cells (RGCs), the primary cell type affected in optic neuropathies. hPSCs were induced to become retinal cells using a stepwise differentiation protocol that allowed for formation of optic vesicle (OV)-like structures. Enrichment of OV like structures allowed for the definitive identification of RGCs. RGCs displayed the proper temporal, spatial, and phenotypic characteristics of RGCs developing in vivo. To test the ability of hPSC-RGCs to serve as a disease model, lines were generated from a patient with an E50K mutation in the Optineurin gene, causative for normal tension primary open angle glaucoma. E50K RGCs displayed significantly higher levels of apoptosis compared to a control lines. Apoptosis was reduced with exposure to neuroprotective factors. Lastly, hPSC-derived RGCs were studied for their ability to develop functional features possessed by mature in vivo RGCs. hPSC-derived RGCs displayed a few immature functional features and as such, strategies in which to expedite synaptogenesis using hPSC-derived astrocytes were explored. Astrocyte and RGG co-cultures displayed expedited synaptic and functional maturation, more closely resembling mature in vivo RGCs. Taken together, the results of this study have important implications for the study of RGC development and by extension, the advancement of translational therapies for optic neuropathies.
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    Retinal Ganglion Cells With a Glaucoma OPTN(E50K) Mutation Exhibit Neurodegenerative Phenotypes when Derived from Three-Dimensional Retinal Organoids
    (Elsevier, 2020-07-14) VanderWall, Kirstin B.; Huang, Kang-Chieh; Pan, Yanling; Lavekar, Sailee S.; Fligor, Clarisse M.; Allsop, Anna R.; Lentsch, Kelly A.; Dang, Pengtao; Zhang, Chi; Tseng, Henry C.; Cummins, Theodore R.; Meyer, Jason S.; Medical and Molecular Genetics, School of Medicine
    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.