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• W2293795453 abstract "Human embryonic stem cells (hESCs) are undifferentiated cells arising from the inner cell mass of the blastocyst, which are able to self-renew or differentiate in vitro into specialised cell types. These pluripotent cells are a powerful tool to study human embryonic development and have great potential in the field of regenerative medicine.Human ESC pluripotency is governed by an intrinsic transcriptional network composed of the three well-known transcription factors OCT4, SOX2 and NANOG, whereas the role of extrinsic cell/microenvironment interactions in the maintenance of hESC stemness has been neglected to some extent. The aim of this work was to develop a systems biology approach oriented on these extrinsic factors and their links with the transcriptional network, in order to uncover some of the fundamental mechanisms underlying the stemness state. The thesis is divided into two complementary approaches: a top-down in silico study and a bottom-up in vitro study. The top-down in silico approach consists of a meta-analysis of hESC transcriptional data, leading to the construction of a hESC transcriptome. These mRNA data served as proxy for proteins in a protein-protein interaction database to build a hESC interactome. This interactome (or protein-protein interaction network) was structurally defined to identify the likely cell surface and extracellular proteins regulating hESC stemness by revealing the ’module organiser’ or hub proteins and the ’module connector’ or bottleneck proteins, along with the extracellular/transcriptional links.The bottom-up in vitro approach was the study of five of the previously identified cell surface/extracellular proteins in hESC fate decision. These candidates, together with OCT4, were stably knocked down using short hairpin (sh)RNAs and lentiviruses. The optimisation of the shRNA lentivirus production led to the development of a method for the direct quantification of these lentiviral particles. The effects of shRNA-mediated knockdown on hESC phenotype were investigated by assessing cell morphology and by determining the expression levels of the following groups of mRNAs: candidate stemness mRNAs, pluripotency mRNAs, as well as trophectoderm, endoderm, mesoderm and ectoderm mRNAs. We found that the candidates could modulate each other’s expression and appeared to regulate hESC commitment into different lineages. Furthermore, the expression levels of some of the candidates were regulated by OCT4. Taken together, these results suggest that by using the novel in silico approach developed during this project, it is possible to identify new stemness factors that could potentially have a role in either maintaining hESC self-renewal or in regulating lineage specification." @default.
• W2293795453 created "2016-06-24" @default.
• W2293795453 creator A5003629568 @default.
• W2293795453 date "2014-09-30" @default.
• W2293795453 modified "2023-09-26" @default.
• W2293795453 title "Defining stemness of human embryonic stem cells : a systems biology approach" @default.
• W2293795453 hasPublicationYear "2014" @default.
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