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Transient Downregulation of Nanog and Oct4 Induced by DETA/NO Exposure in Mouse Embryonic Stem Cells Leads to Mesodermal/Endodermal Lineage Differentiation.

Mora-Castilla S, Tejedo JR, Tapia-Limonchi R, Díaz I, Hitos AB, Cahuana GM, Hmadcha A, Martín F, Soria B, Bedoya FJ - Stem Cells Int (2014)

Bottom Line: Simultaneously, mRNA of mesodermal markers Flk1 and Mef2c are also regulated by the treatment.Acetylated histone H3 occupancy at the promoter of Nanog is involved in the process of reexpression.Furthermore, Nanog binding to the promoter of Brachyury leads to repression of this gene, thus disrupting mesendoderm transition.

View Article: PubMed Central - PubMed

Affiliation: Andalusian Center for Molecular Biology and Regenerative Medicine (CABIMER), University Pablo de Olavide, Biomedical Research Network (CIBER) of Diabetes and Related Metabolic Diseases, Red-Tercel, Avenida Américo Vespucio S/N, 41092 Seville, Spain ; Sanford Consortium for Regenerative Medicine, University of California San Diego, 2880 Torrey Pines Scenic Drive, La Jolla, CA 92037, USA.

ABSTRACT
The function of pluripotency genes in differentiation is a matter of investigation. We report here that Nanog and Oct4 are reexpressed in two mouse embryonic stem cell (mESC) lines following exposure to the differentiating agent DETA/NO. Both cell lines express a battery of both endoderm and mesoderm markers following induction of differentiation with DETA/NO-based protocols. Confocal analysis of cells undergoing directed differentiation shows that the majority of cells expressing Nanog express also endoderm genes such as Gata4 and FoxA2 (75.4% and 96.2%, resp.). Simultaneously, mRNA of mesodermal markers Flk1 and Mef2c are also regulated by the treatment. Acetylated histone H3 occupancy at the promoter of Nanog is involved in the process of reexpression. Furthermore, Nanog binding to the promoter of Brachyury leads to repression of this gene, thus disrupting mesendoderm transition.

No MeSH data available.


Related in: MedlinePlus

Nanog colocalizes with Gata4 and FoxA2 in definitive endoderm cells. Cells were differentiated according to protocol described in Section 2 and analysed by confocal immunofluorescence (a) for Nanog, Gata4, and FoxA2 at day 8 of differentiation. (b) Analysis showing the percentage of cells coexpressing Nanog-Gata4 and Nanog-FoxA2 in D3 differentiated cells. Data are mean ± ESM from 7–9 fields.
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fig4: Nanog colocalizes with Gata4 and FoxA2 in definitive endoderm cells. Cells were differentiated according to protocol described in Section 2 and analysed by confocal immunofluorescence (a) for Nanog, Gata4, and FoxA2 at day 8 of differentiation. (b) Analysis showing the percentage of cells coexpressing Nanog-Gata4 and Nanog-FoxA2 in D3 differentiated cells. Data are mean ± ESM from 7–9 fields.

Mentions: To test the possibility that the presence of stemness genes in differentiated cell populations might indeed be ascribed to cells escaping from differentiation processes, confocal microscopy immunofluorescence analyses were performed in cells induced to differentiate into DE. Nanog expression increases from 0.65% (data not shown) in NO-treated cells to 54% in differentiated cells population (data calculated from Figure 4(a) ratio Nanog + versus Dapi). Results show that Nanog is heterogeneously expressed in DE-differentiated cells (Figure 4(a)) and that the majority of Nanog positive cells also coexpressed Gata4 and FoxA2 (75.4% and 96.2%, resp.), with the cells expressing only Nanog no more than 7% (Figures 4(a) and 4(b)).


Transient Downregulation of Nanog and Oct4 Induced by DETA/NO Exposure in Mouse Embryonic Stem Cells Leads to Mesodermal/Endodermal Lineage Differentiation.

Mora-Castilla S, Tejedo JR, Tapia-Limonchi R, Díaz I, Hitos AB, Cahuana GM, Hmadcha A, Martín F, Soria B, Bedoya FJ - Stem Cells Int (2014)

Nanog colocalizes with Gata4 and FoxA2 in definitive endoderm cells. Cells were differentiated according to protocol described in Section 2 and analysed by confocal immunofluorescence (a) for Nanog, Gata4, and FoxA2 at day 8 of differentiation. (b) Analysis showing the percentage of cells coexpressing Nanog-Gata4 and Nanog-FoxA2 in D3 differentiated cells. Data are mean ± ESM from 7–9 fields.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4269087&req=5

fig4: Nanog colocalizes with Gata4 and FoxA2 in definitive endoderm cells. Cells were differentiated according to protocol described in Section 2 and analysed by confocal immunofluorescence (a) for Nanog, Gata4, and FoxA2 at day 8 of differentiation. (b) Analysis showing the percentage of cells coexpressing Nanog-Gata4 and Nanog-FoxA2 in D3 differentiated cells. Data are mean ± ESM from 7–9 fields.
Mentions: To test the possibility that the presence of stemness genes in differentiated cell populations might indeed be ascribed to cells escaping from differentiation processes, confocal microscopy immunofluorescence analyses were performed in cells induced to differentiate into DE. Nanog expression increases from 0.65% (data not shown) in NO-treated cells to 54% in differentiated cells population (data calculated from Figure 4(a) ratio Nanog + versus Dapi). Results show that Nanog is heterogeneously expressed in DE-differentiated cells (Figure 4(a)) and that the majority of Nanog positive cells also coexpressed Gata4 and FoxA2 (75.4% and 96.2%, resp.), with the cells expressing only Nanog no more than 7% (Figures 4(a) and 4(b)).

Bottom Line: Simultaneously, mRNA of mesodermal markers Flk1 and Mef2c are also regulated by the treatment.Acetylated histone H3 occupancy at the promoter of Nanog is involved in the process of reexpression.Furthermore, Nanog binding to the promoter of Brachyury leads to repression of this gene, thus disrupting mesendoderm transition.

View Article: PubMed Central - PubMed

Affiliation: Andalusian Center for Molecular Biology and Regenerative Medicine (CABIMER), University Pablo de Olavide, Biomedical Research Network (CIBER) of Diabetes and Related Metabolic Diseases, Red-Tercel, Avenida Américo Vespucio S/N, 41092 Seville, Spain ; Sanford Consortium for Regenerative Medicine, University of California San Diego, 2880 Torrey Pines Scenic Drive, La Jolla, CA 92037, USA.

ABSTRACT
The function of pluripotency genes in differentiation is a matter of investigation. We report here that Nanog and Oct4 are reexpressed in two mouse embryonic stem cell (mESC) lines following exposure to the differentiating agent DETA/NO. Both cell lines express a battery of both endoderm and mesoderm markers following induction of differentiation with DETA/NO-based protocols. Confocal analysis of cells undergoing directed differentiation shows that the majority of cells expressing Nanog express also endoderm genes such as Gata4 and FoxA2 (75.4% and 96.2%, resp.). Simultaneously, mRNA of mesodermal markers Flk1 and Mef2c are also regulated by the treatment. Acetylated histone H3 occupancy at the promoter of Nanog is involved in the process of reexpression. Furthermore, Nanog binding to the promoter of Brachyury leads to repression of this gene, thus disrupting mesendoderm transition.

No MeSH data available.


Related in: MedlinePlus