Indiana University-Purdue University Indianapolis (IUPUI) / Human pluripotent stem cells (hPSCs) allow for the unprecedented ability to recapitulate early stages of human development, which are otherwise inaccessible to investigation. This is especially true for one of the earliest events in human development, the establishment of a neuroretinal fate from an unspecified pluripotent population. To test the ability of hPSCs to serve in this capacity, hPSCs were generated using mRNA-reprogamming methods and maintained in xenogeneic-free differentiation conditions. These cells were directed to differentiate using a three-dimensional approach to analyze their ability to successfully recapitulate early events in human development in a temporal and developmentally-appropriate fashion. To do so, hPSCs were first directed to an anterior neural phenotype, which was confirmed by analysis of stage-specific neural transcription factors via immunocytochemistry and quantitative RT-PCR. Next, the cells were directed to an optic vesicle-like stage, where the presumptive retinal cells were identified by the expression of specific transcription factors. Finally, three-dimensional optic vesicle-like retinal organoids were identified, isolated, and further analyzed for the expression of markers associated with some of the differentiated cell types of the neural retina. Upon establishment of hPSC-derived retinal organoids, this system was further utilized to study the neurodegeneration in glaucoma and provide insights into the disease mechanisms. Overall, the results of this study help to demonstrate the suitability of hPSC-differentiation approaches as an effective tool to model retinal development and disease.
Identifer | oai:union.ndltd.org:IUPUI/oai:scholarworks.iupui.edu:1805/13409 |
Date | January 2017 |
Creators | Sridhar, Akshayalakshmi |
Contributors | Meyer, Jason |
Source Sets | Indiana University-Purdue University Indianapolis |
Language | en_US |
Detected Language | English |
Type | Thesis |
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