The role of PI3K/AKT, MAPK/ERK and NFkappabeta signalling in the maintenance of human embryonic stem cell pluripotency and viability highlighted by transcriptional profiling and functional analysis.
Understanding the molecular mechanism by which pluripotency is maintained in human embryonic stem cells (hESC) is important for the development of improved methods to derive, culture and differentiate these into cells of potential therapeutic use. Large-scale transcriptional comparison of the hES-NCL1 line derived from a day 8 embryo with H1 line derived from a day 5 embryo (WiCell Inc.) showed that only 0.52% of the transcripts analysed varied significantly between the two cell lines. This is within the variability range that has been reported when hESC derived from days 5-6 embryos have been compared with each other. This implies that transcriptional differences between the cell lines are likely to reflect their genetic profile rather than the embryonic stage from which they were derived. Bioinformatic analysis of expression changes observed when these cells were induced to differentiate as embryoid bodies suggested that quite a few of the downregulated genes were components of signal transduction networks. Subsequent analysis using western blotting, flow cytometry and antibody arrays implicated components of the PI3K/AKT kinase, MAPK/ERK and NFkappabeta pathways and confirmed that these components are decreased upon differentiation. Disruption of these pathways in isolation using specific inhibitors resulted in loss of pluripotency and/or loss of viability suggesting the importance of such signalling pathways in embryonic stem cell maintenance.
Armstrong
Lyle
L
Centre for Stem Cell Biology and Developmental Genetics, Institute of Human Genetics, University of Newcastle, Central Parkways, Newcastle upon Tyne, UK.
Hughes
Owen
O
Yung
Sun
S
Hyslop
Louise
L
Stewart
Rebecca
R
Wappler
Ilka
I
Peters
Heiko
H
Walter
Theresia
T
Stojkovic
Petra
P
Evans
Jerome
J
Stojkovic
Miodrag
M
Lako
Majlinda
M
D002470:Cell Survival; D019295:Computational Biology; D004622:Embryo, Mammalian / Q000166:cytology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005260:Female; D006801:Humans; D007621:Karyotyping; D020935:MAP Kinase Signaling System; D008297:Male; D008954:Models, Biological; D016328:NF-kappa B / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D015398:Signal Transduction; D013234:Stem Cells / Q000166:cytology*; D014158:Transcription, Genetic
D016328:NF-kappa B; D019869:Phosphatidylinositol 3-Kinases; D048049:Extracellular Signal-Regulated MAP Kinases