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Sarah Ohlemacher, Akshayalakshmi Sridhar, Yucheng Xiao, Theodore Cummins, Jason S Meyer; Functional Maturation and Long-Term Survival of Human Pluripotent Stem Cell-Derived Retinal Ganglion Cells. Invest. Ophthalmol. Vis. Sci. 2016;57(12):6070.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal ganglion cells (RGCs) play an essential role in transmitting visual information from the eye to the visual thalamus in the brain. They are also the primary cell type affected in traumatic retinal injuries as well as optic neuropathies. Human pluripotent stem cells (hPSCs) provide an attractive source of cells that can be used for patient-specific cell replacement therapies, drug screening, and disease modeling. However, little is known about the functional maturation of these hPSC-derived RGCS. As such, the development of these neurons was studied over time, and methods in which to influence their maturation and functionality were tested.
hPSCs were differentiated toward a retinal lineage in a stepwise fashion as previously described, yielding highly enriched populations of optic vesicle-like neurospheres. Within 40 days of differentiation, presumptive RGCs were identified and further characterized based upon morphology and expression of RGC-associated factors. These RGCs were grown in culture for prolonged periods of time, and the emergence of specific RGC subtypes was examined. Additionally, RGC maturation was explored by immunocytochemistry and electrophysiology, including the formation of presumptive synaptic connections. Candidate factors were added to RGCs to aid in the synaptic maturation of these cells, with analyses performed by microscopy and electrophysiology.
Presumptive RGCs differentiated in a temporally-appropriate manner and expressed an array of RGC-associated features. Moreover, subtypes of RGCs were identifiable over time in prolonged cultures, including melanopsin-expressing ipRGCs. The maturation of hPSC-derived RGCs was analyzed by neurite outgrowth and the development of synaptic-like structures in prolonged cultures. Furthermore, the development of hPSC-derived RGCs was influenced by the addition of critical factors capable of enhancing RGC differentiation and maturation.
The data presented demonstrates the ability to derive RGCs from hPSCs though a stepwise differentiation protocol, including multiple RGC subtypes. Moreover, the functional maturation of these cells was observed in prolonged cultures and found to increase under the influence of candidate factors. These results will facilitate the use of hPSC-derived RGCs to study optic neuropathies, the development of drug therapeutics and for cell replacement therapies.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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