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• W109650861 abstract "Human embryonic stem cells (hESCs) have great potential for use in regenerative medicine due to their indefinite capacity for self-renewal and their ability to differentiate into cell types derived from all three embryonic germ layers. An important step toward advancing their use in cell-based therapies is development of methods for directed differentiation into specific lineages. Understanding the signaling pathways that underlie hESC differentiation is critical if we are to achieve the full potential of these cells.EphrinB-ephB receptor interactions are a common regulator of multiple somatic stem cells, including neural stem cells (NSCs) and hematopoietic stem cells (HSCs). EphrinB2, in particular, has been identified as a molecular stem cell marker shared in mouse embryonic stem cells (ESCs), NSCs, and HSCs. Its receptor, ephB4, has also been implicated in playing a role in ESC differentiation. Despite evidence that the ephrinB2-ephB4 axis may be involved in ESC fate, this axis has not been carefully studied in hESCs due to a lack of highly specific reagents to interrogate ephrinB2-ephB4 interactions. Here, we characterized the role of ephrinB2 in hESC fate.First, we developed a novel method to specifically tag ephrinB2 hESCs. We showed that we could efficiently pseudotype a lentiviral vector with the Nipah virus (NiV) envelope that could efficiently target ephrinB2+ cells in vitro. These NiV pseudotyped particles specifically transduced a sub-population of hESCs. Next, we sought to further characterize these ephrinB2+ hESCs by FACS-sorting the NiV-transduced GFP+ cells and examining their properties in culture. Passaging of the sorted cells revealed that ephrinB2 does not mark for a stable population of cells, but may be an intrinsic marker of heterogeneity. The ephrinB2+ hESCs demonstrated the abilities to self-renew in vitro, to differentiate into the three germ layers in vitro, and to form teratomas in vivo.Lastly, to further examine the role of ephrinB2 in hESC fate, we generated a hESC line stably expressing an shRNA against ephrinB2 (shEFNB2). Real-time PCR analysis and microarray analysis of embryoid bodies (EBs) derived from shEFNB2 cells demonstrated a severe deficiency in neuro-ectoderm gene expression and an up-regulation of genes involved in mesendoderm specification. Further, functional mesoderm-directed differentiation assays revealed that shEFNB2 hESCs have an increased propensity to differentiate into one specific sub-type of mesenchymal cells. In sum, the findings of this dissertation suggest that the heterogeneity of ephrinB2 expression and perturbation of ephrinB2 signaling may both be manipulated to enhance directed differentiation of hESCs in vitro." @default.
• W109650861 created "2016-06-24" @default.
• W109650861 creator A5024725724 @default.
• W109650861 date "2012-01-01" @default.
• W109650861 modified "2023-09-27" @default.
• W109650861 title "Characterization of the role of ephrinB2 in human embryonic stem cell fate" @default.
• W109650861 hasPublicationYear "2012" @default.
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