Abstract
In this study we examined the role of the cAMP/protein kinase A (PKA) pathway in affecting IOUD2 ES cell self-renewal and differentiation, Oct4 expression, and cell proliferation. Forskolin, the adenylate cyclase agonist, alone had no effect on ES cell self-renewal. However, when cells were treated with the differentiation-inducing agent retinoic acid, forskolin significantly promoted ES cell self-renewal. Effectively, forskolin rescued cells from a pathway of differentiation. Culturing ES cells in the presence of the phosphodiesterase inhibitor IBMX had no effect on ES cell self-renewal but did increase cell proliferation. In the presence of 100 μM IBMX without LIF, 10 μM forskolin significantly increased ES cell self-renewal. The cell permeable cAMP analog 8-Br-cAMP (1 and 5 mM) promoted ES cell differentiation in the presence of LIF, while in the absence of LIF, it promoted ES cell self-renewal. The effect of the PKA specific inhibitors H89 and KT5720 on Oct4 expression was, again, LIF-dependent. In the presence of LIF, these inhibitors decreased Oct4 expression, while they increased Oct4 expression in the absence of LIF. In general, ES cells maintained on a self-renewal pathway through the presence of LIF show little effect from altered cAMP signaling except at higher levels. However, in strict contrast, when ES cell are on a differentiation pathway through exposure to retinoic acid or the removal of LIF, altering cAMP levels can rescue the self-renewal process promoting Oct4 expression. This study clearly shows that the cAMP/PKA pathway plays a role in ES cell self-renewal pathways.
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Acknowledgments
The authors wish to thank Dr. Austin Smith, Centre for Genome Research, Edinburgh, UK, for the generous gift of his ES cell lines. This work was partly funded by the Millenium Research Fund National University of Ireland Galway.
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This work was partly funded by the Millennium Research Fund National University of Ireland Galway.
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Faherty, S., Fitzgerald, A., Keohan, M. et al. Self-renewal and differentiation of mouse embryonic stem cells as measured by Oct4 expression: the role of the cAMP/PKA pathway. In Vitro Cell.Dev.Biol.-Animal 43, 37–47 (2007). https://doi.org/10.1007/s11626-006-9001-5
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DOI: https://doi.org/10.1007/s11626-006-9001-5