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Titill
en

The effect of Dlg7 overexpression and silencing on mouse embryonic stem cell differentiation

Útgáfa
Desember 2010
Útdráttur
en

The genetic program controlling stem cell self-renewal and differentiation is yet to be elucidated although significant contributions have been made. Recently, in vitro and in vivo studies have identified several genetic regulatory programs such as various transcription factors, cell cycle inhibitors, genes implicated in chromosomal rearrangements, essential developmental proteins, and signalling pathways that have all been implicated in the process of stem cell self-renewal and differentiation.
Dlg7, a novel cell cycle regulator gene, is expressed in stem cells, including haematopoietic stem cells (HSC), mesenchymal stem cells (MSC) and mouse embryonic stem cells (mESC). Dlg7 is expressed in human haematopoietic stem cells (CD34+CD38-) but much less expression was detected in haematopoietic progenitor cells (CD34+38+) or in fully mature blood cells. Dlg7 is expressed in many leukemic cell lines and in several tumours including bladder, colon and liver, while not being found in healthy adjacent normal tissue. In addition, it is a potential oncogenic target of Aurora-A, and is associated with invasiveness of hepatocellular carcinoma.
The aim of this project was to study the role of Dlg7 in the differentiation of mESC with special emphases on haematopoiesis. We use lentiviral based vectors to silence and overexpress the Dlg7 gene in mESC. Transgenic mESC were differentiated to Embryo Bodies and haematopoietic colony forming units. Effects of genetic changes were monitored using a colony forming unit (CFU) assay, flow-cytometry and Q-PCR.
Dlg7 overexpression and silencing significantly reduces the number of EBs that form in Primary differentiation media compared to scrambled control cell population. Of the EBs that formed a significant larger portion of EB from the scrambled control acquired hematopoietic morphology at later stage of differentiation compared to mESC with Dlg7 overexpression or silencing. BFU-E where in grater numbers with in the CFU population formed from cells with Dlg7 overexpression then in the scrambled control cells.
We can however not say with full certenty that the effect of Dlg7 in hematopoiesis is a direct effect or rather a general effect in differentiation due to the role that Dlg7 plays in the cell cycle.

Birting
4.2.2011


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