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Title: The role of canonical Wnt/GSK3ß/ß-catenin signalling in stem cell self-renewal and differentiation
Author: Tarafdar, Anuradha
ISNI:       0000 0004 2720 3430
Awarding Body: University of Ulster
Current Institution: Ulster University
Date of Award: 2012
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Canonical Wnt signalling influences the 'sternness' of both embryonic stem (ES) cells and haemopoietic stem cells (RSC). In this study we investigated how modulation of the canonical Wnt pathway affected ES cell self-renewal and haemopoietic differentiation. We observed that Wnt activation suppressed differentiation whereas inhibiting Wnt signalling enhanced differentiation. Relative Quantification using TaqMan Pluripotency Array cards ™ indicated that genes correlating to 'sternness' such as AFP, Tert, Nanog, Zgp 42, Tcf were up-regulated along with characteristic mesodermal markers (Brachyury, Pecam J, Cdh 5, Nodal, Myo DJ) whereas differentiation markers (Myf 5, Foxa 2, Pax 6, Actc, Neuro DJ) were down regulated following activation of the pathway using the GSK3 inhibitor BIO or expression of dominant positive ~-catenin (DP-~C). Chromatin Immunoprecipitation technology demonstrated a complex interplay occurring between ~-cateninlTCF/LEFlBrachyury and Nanog expression following Wnt signalling. In addition during haemopoietic differentiation we demonstrate that at the early hemangioblast stage, the primitive erythroid genes (Pecaml , LM02 and Tie2) , genes related to globin switching (Hbz, Hbb-BHJ, Hba-al , Hba-a2, Hbb-bJ Hbb-b2) and the definitive erythroid genes and haematopoietic regulators PUJ, GATA J, 3 and 4, EPO-R and KLF were all up- regulated following Wnt activation. A higher percentage of cells at this stage expressed Scal , c-Kit, CD44 and CD45 compared to control cells. Furthermore when directed to form multipotent progenitors, Wnt activation resulted in cells having a megakaryotype erythroid progenitor phenotype characterised by high CD4l, CD7l, CD45 and CD24 and Ter1l91ow expression. In colony assays following myeloid differentiation Wnt activation suppressed granulocyte-macrophage colony formation, resulting in a significant increase in erythroid and mixed colonies. Overall this study demonstrates that activation of the canonical Wnt pathway not only maintains 'sternness' during mesodermal differentiation, but also enhances hematopoietic differentiation towards the erythroid lineage, thus establishing an important role for this pathway in the onset of primitive and definitive erythropoiesis.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available