scholarly journals Toll-Like Receptor 2 Mediates Proliferation, Survival, NF-κB Translocation, and Cytokine mRNA Expression in LIF-Maintained Mouse Embryonic Stem Cells

2010 ◽  
Vol 19 (9) ◽  
pp. 1333-1341 ◽  
Author(s):  
Tammi Taylor ◽  
Young-June Kim ◽  
Xuan Ou ◽  
Wilbert Derbigny ◽  
Hal E. Broxmeyer
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Mrna Expression ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Toll Like Receptor ◽  
Cytokine Mrna ◽  
Cytokine Mrna Expression ◽  
Toll Like Receptor 2

scholarly journals Another face Of DMSO: Effects of dimethyl sulfoxide (DMSO) on the pluripotency and differentiation capacity of mouse embryonic stem cells

2019 ◽  
Author(s):  
JunKoo Yi ◽  
Jae-Jung Ha ◽  
Dae-Hyun Kim ◽  
Zae-Young Ryoo ◽  
Sung-Hyun Kim ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Dimethyl Sulfoxide ◽  
Mrna Expression ◽  
Culture Medium ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Test Methods ◽  
Undifferentiated State ◽  
Dmso Treatment

Abstract Background Mouse embryonic stem cells (mESCs) go through self-renewal in the existence of the cytokine leukemia inhibitory factor (LIF). LIF is add to the mouse stem cells culture medium, and its removal results in fast differentiation. Dimethyl sulfoxide (DMSO) is one of the most general used solvents in drug test. Methods We exposed 4-day mESC cultures to different concentrations of DMSO (0.1%, 0.5%, 1.0%, and 2.0%) to identify the safest dose exhibiting efficacy as a solvent. mESCs grown under general pluripotency conditions in the absence of LIF were treated with DMSO. In addition, as a control for differentiation, mESCs were grown absence of LIF. Results DMSO downregulated the mRNA expression level of pluripotency markers. Moreover, DMSO reduced the mRNA expression levels of ectodermal marker (β-tubulin3) and a mesodermal marker (Hand1) and endodermal markers (Foxa2 and Sox17) in mESCs. These results indicate that DMSO treatment enhances the pluripotency and disrupts the differentiation of mESCs. We also show that members of the Tet oncogene family are critical to inhibiting the differentiation and methylation of mESCs. Conclusion DMSO is appropriate to sustain the pluripotency of mESCs in the absence of LIF, and that mESCs can be sustained in an undifferentiated state using DMSO. Therefore, DMSO may, in part, function as a substitute for LIF.

scholarly journals Another face Of DMSO: Effects of dimethyl sulfoxide (DMSO) on the pluripotency and differentiation capacity of mouse embryonic stem cells

2019 ◽  
Author(s):  
JunKoo Yi ◽  
Jae-Jung Ha ◽  
Dae-Hyun Kim ◽  
Zae-Young Ryoo ◽  
Sung-Hyun Kim ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Dimethyl Sulfoxide ◽  
Mrna Expression ◽  
Culture Medium ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Test Methods ◽  
Undifferentiated State ◽  
Dmso Treatment

Abstract Background Mouse embryonic stem cells (mESCs) go through self-renewal in the existence of the cytokine leukemia inhibitory factor (LIF). LIF is add to the mouse stem cells culture medium, and its removal results in fast differentiation. Dimethyl sulfoxide (DMSO) is one of the most general used solvents in drug test. Methods We exposed 4-day mESC cultures to different concentrations of DMSO (0.1%, 0.5%, 1.0%, and 2.0%) to identify the safest dose exhibiting efficacy as a solvent. mESCs grown under general pluripotency conditions in the absence of LIF were treated with DMSO. In addition, as a control for differentiation, mESCs were grown absence of LIF.Results DMSO downregulated the mRNA expression level of pluripotency markers. Moreover, DMSO reduced the mRNA expression levels of ectodermal marker (β-tubulin3) and a mesodermal marker (Hand1) and endodermal markers (Foxa2 and Sox17) in mESCs. These results indicate that DMSO treatment enhances the pluripotency and disrupts the differentiation of mESCs. We also show that members of the Tet oncogene family are critical to inhibiting the differentiation and methylation of mESCs. Conclusion DMSO is appropriate to sustain the pluripotency of mESCs in the absence of LIF, and that mESCs can be sustained in an undifferentiated state using DMSO. Therefore, DMSO may, in part, function as a substitute for LIF.

scholarly journals Mouse Embryonic Stem Cells Express Functional Toll Like Receptor 2.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1473-1473 ◽  
Author(s):  
Tammi Taylor ◽  
Young-June Kim ◽  
Wilbert Derbigny ◽  
Xuan Ou ◽  
Hal E. Broxmeyer
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Cytokine Production ◽  
Nuclear Translocation ◽  
Embryonic Stem ◽  
Cytokine Release ◽  
Toll Like Receptor ◽  
Ligand Activation ◽  
Nuclear Extracts ◽  
Toll Like Receptor 2

Abstract Abstract 1473 Poster Board I-496 Mouse embryonic stem cells (mESCs) are unique in that they give rise to every cell type of the body. Little is known about stimuli that promote mESC differentiation and proliferation. We hypothesized that TLRs are expressed and functional, and when activated by its ligand will influence survival and proliferation of mESCs in the presence of Leukemia Inhibitory Factor (LIF). This study evaluated three mESC lines, R1, CGR8, and E14 to first determine if they express Toll Like Receptors (TLRs) at mRNA and protein levels. Next, we evaluated if the TLR ligands would induce or modulate mESC proliferation and survival of the mESCs in the presence of LIF. We then assessed the E14 mESC line to determine if the mESCs expressed MyD88 an adaptor protein molecule, known to be involved in the TLR pathway and if the TLR ligands would cause inhibitor of kinase kinase alpha/ beta (IKKα/β) phosphorylation and nuclear factor-kappa beta (NF-κβ) nuclear translocation, and cytokine production. In this study we found expression of TLRs 1, 2, 3, 5 and 6 at the mRNA level, but no mRNA expression of TLRs 4, 7, 8 and 9. We also confirmed some of these results by flow analysis. Toll Like Receptor 2 (TLR-2), but not Toll Like Receptor 4 (TLR-4), is expressed on the three mESC lines. Therefore we focused our studies on TLR-2, specifically. Pam3Cys, a synthetic triacyl-lipoprotein and a TLR-2 ligand induced a significant increase in mESC numbers on Day 3 compared to controls and also at Day 4 and Day 5 when compared to controls. Pam3Cys (10ug/ml) also enhanced the survival of mESC colony forming cells subjected to serum withdrawal and then delayed addition of serum. Next we found that E14 mESCs express molecules involved in the TLR Pathway. MyD88 was expressed in mESCs and IKKα/β phosphorylation was enhanced after 15 minute by TLR-2 ligand activation. We found a significant increase of NF-κβ nuclear translocation upon activation by Pam3Cys after 30 minutes which continued for up to an hour. Densitometry analysis of the nuclear extracts from three separate experiments shows a significant 2 fold increase in NF-κβ in the nucleus compared to control mESC nuclear extracts. Since TLR activation of leukocytes enhances cytokine production, and our group has published that mESCs produce cytokines, we studied the effect of Pam3Cys on release of cytokine from our mESC line. Cytokine release in mESCs by TLR-ligand activation was assessed by ELISA, but TLR-2 ligand stimulation did not affect cytokine release of IL-6, TNF-α, or INF-β. We found that there were no significant changes in expression of mESCs markers Oct4, KLF-4, Sox 2, and SSEA1 when compared to cells not activated by Pam3Cys. Thus the mESCs remained in a pluripotent state in the presence of LIF after activation with the TLR-2 ligand. These results demonstrate that mESCs can respond to microbial products, and TLR-2 activation enhances proliferation and survival of the mESCs. This finding expands the role of TLRs and has implications for a better understanding of the responsiveness of embryonic stem cells to certain microbial agents. Disclosures: No relevant conflicts of interest to declare.

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