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Expression profiling in the characterization of differentiating human embryonic stem cells
thesisposted on 2017-02-14, 00:14 authored by Yu, Qing Cissy
During Embryogenesis, the harbinger of germ layer formation is the primitive streak (PS), a transient structure oriented along the embryonic anteroposterior axis, through which epiblast cells egress to form mesoderm and endoderm. Hematopoietic and vascular lineages arise in multiple sites and at different times during embryogenesis from extra embryonic and lateral plate mesoderm, derivatives of the posterior and mid regions of the PS. In vitro differentiation of human embryonic stem cells (hESCs) has been demonstrated to recapitulate aspects of vertebrate development, and we have established an in vitro hESC differentiation system to study the initiation of gastrulation as well as the development of early hematopoietic mesoderm in a defined, serum-free medium, showing that exogenously added TGFb family molecules induce primitive streak-like cells that subsequently differentiate into mesodermal and endodermal lineages. In order to ascertain the precise isolation of mesodermal precursors, a targeted hESC line in which GFP was inserted by homologous recombination in the locus of the MIXL1 homeobox gene (MIXL1GFP/W) was generated and used as guidance for the identification of viable, PS-like, cells. In this study, microarray-based transcriptional profiling was used to examine gene expression during mesoderm induction from MIXL1GFP/W hESCs in isolated mesodermal populations. The aim of this study was to identify novel cell surface markers and growth factors that would facilitate the identification and generation of hematopoietic lineages from hESCs. During this study, transcriptional profiling method was also used to assist the characterization of hESCs derived cell phenotypes, such as neuronal, endothelial, cardiac and endodermal pancreatic progenitor cells.