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School of Medicine and Health Sciences Poster Presentations
Retinoic Acid Signaling in Embryonic Stem Cells
Poster Number
266
Document Type
Poster
Publication Date
3-2016
Abstract
This project aims to improve our understanding of how embryonic stem cells (ESCs) are induced to form neurons, which is central for developing methods for using ESCs in repair and regeneration. It is known that retinoic acid (RA) signaling promotes neural differentiation of pluripotent stem cells; however, it is not clear when and where this occurs – does RA directly induce neurons from the ESCs themselves, or does it promote the proliferation or differentiation of an intermediate subtype of neural-specific stem cell. To assess this question, DNA constructs were made using destabilized green fluourescent proten (dGFP) that will allow us to observe the differentiation of embryonic stem cells (ESC) into neurons in real time. We made three constructs to allow for labeling of ESC as they mature into neurons. The first and second constructs express GFP and destabilized GFP respectively, and include a neomycin cassette for cell selection. The third uses a conditional variant of Cre (CreERT) that was transfected in combination with a reporter (floxed tdTomato) and a hygromycin cassette for selection. When Tamoxifen is added, the activated Cre will activate the tdTomato reporter, labeling RA activated cells and their progeny. These experiments have been initiated and results from these transfections are forthcoming. If successful, we can define the lineage relationship between RA-activated cells and mature neurons. Understanding how pluripotent stem cells become neurons also has implications for tumorogeneisis, specifically the uncontrolled proliferation in glioblastoma, which is critical for developing new treatments and to guide neurosurgical intervention.
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Open Access
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Retinoic Acid Signaling in Embryonic Stem Cells
This project aims to improve our understanding of how embryonic stem cells (ESCs) are induced to form neurons, which is central for developing methods for using ESCs in repair and regeneration. It is known that retinoic acid (RA) signaling promotes neural differentiation of pluripotent stem cells; however, it is not clear when and where this occurs – does RA directly induce neurons from the ESCs themselves, or does it promote the proliferation or differentiation of an intermediate subtype of neural-specific stem cell. To assess this question, DNA constructs were made using destabilized green fluourescent proten (dGFP) that will allow us to observe the differentiation of embryonic stem cells (ESC) into neurons in real time. We made three constructs to allow for labeling of ESC as they mature into neurons. The first and second constructs express GFP and destabilized GFP respectively, and include a neomycin cassette for cell selection. The third uses a conditional variant of Cre (CreERT) that was transfected in combination with a reporter (floxed tdTomato) and a hygromycin cassette for selection. When Tamoxifen is added, the activated Cre will activate the tdTomato reporter, labeling RA activated cells and their progeny. These experiments have been initiated and results from these transfections are forthcoming. If successful, we can define the lineage relationship between RA-activated cells and mature neurons. Understanding how pluripotent stem cells become neurons also has implications for tumorogeneisis, specifically the uncontrolled proliferation in glioblastoma, which is critical for developing new treatments and to guide neurosurgical intervention.
Comments
Presented at: GW Research Days 2016