NO/beta-catenin crosstalk modulates primitive streak formation prior to embryonic stem cell osteogenic differentiation

Ding, Huawen; Keller, Kevin C.; Martinez, Ivann Kenneth Carvajal; Geransar, Rose; Nieden, Kai O. zur; Nishikawa, Sandra; Rancourt, Derrick E.; Nieden, Nicole I. zur

doi:10.1242/jcs.081703

Cited by 33 publications

(21 citation statements)

References 87 publications

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“…These data coincide with a past study that showed improvement in proliferation of human osteoblast cells (CRL 1543) when the culture medium was supplemented with 0.0195% Tualang honey [30]. The underlying mechanism for the observed improvements is unknown, but it could be related to the active compounds present in Tualang honey or to the presence of H 2 O 2 , a ROS that is essential for modulating stem cell behaviours at the physiological level [45]–[47]. Pan et al (2011) recently suggested that low levels of H 2 O 2 (10–50 µM) could promote rabbit corneal epithelial cell attachment, mobility, and wound repair [48].…”

Section: Discussionsupporting

confidence: 89%

Tualang Honey Improves Human Corneal Epithelial Progenitor Cell Migration and Cellular Resistance to Oxidative Stress In Vitro

et al. 2014

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Stem cells with enhanced resistance to oxidative stress after in vitro expansion have been shown to have improved engraftment and regenerative capacities. Such cells can be generated by preconditioning them with exposure to an antioxidant. In this study we evaluated the effects of Tualang honey (TH), an antioxidant-containing honey, on human corneal epithelial progenitor (HCEP) cells in culture. Cytotoxicity, gene expression, migration, and cellular resistance to oxidative stress were evaluated. Immunofluorescence staining revealed that HCEP cells were holoclonal and expressed epithelial stem cell marker p63 without corneal cytokeratin 3. Cell viability remained unchanged after cells were cultured with 0.004, 0.04, and 0.4% TH in the medium, but it was significantly reduced when the concentration was increased to 3.33%. Cell migration, tested using scratch migration assay, was significantly enhanced when cells were cultured with TH at 0.04% and 0.4%. We also found that TH has hydrogen peroxide (H2O2) scavenging ability, although a trace level of H2O2 was detected in the honey in its native form. Preconditioning HCEP cells with 0.4% TH for 48 h showed better survival following H2O2-induced oxidative stress at 50 µM than untreated group, with a significantly lower number of dead cells (15.3±0.4%) were observed compared to the untreated population (20.5±0.9%, p<0.01). Both TH and ascorbic acid improved HCEP viability following induction of 100 µM H2O2, but the benefit was greater with TH treatment than with ascorbic acid. However, no significant advantage was demonstrated using 5-hydroxymethyl-2-furancarboxaldehyde, a compound that was found abundant in TH using GC/MS analysis. This suggests that the cellular anti-oxidative capacity in HCEP cells was augmented by native TH and was attributed to its antioxidant properties. In conclusion, TH possesses antioxidant properties and can improve cell migration and cellular resistance to oxidative stress in HCEP cells in vitro.

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Section: Discussionsupporting

confidence: 89%

Tualang Honey Improves Human Corneal Epithelial Progenitor Cell Migration and Cellular Resistance to Oxidative Stress In Vitro

et al. 2014

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“…Similar to our results, co-activation of NO, BMP2, and MAPK pathways are involved in osteoblast proliferation and differentiation downstream of phytoestrogeninduced osteogenesis [40]. NO is known to induce osteogenesis of ESCs through activation of canonical Wnt signaling [21]. More recently, our group has reported that Wnt signaling is associated with the osteogenic differentiation of stem cells under various experimental conditions [41][42][43].…”

Section: Discussionsupporting

confidence: 87%

“…A number of groups have demonstrated that NO encourages the differentiation of bone marrow-derived mesenchymal stem cells (MSCs) into osteoprogenitor cells [17][18][19]. However, whether NO is associated with the osteogenic differentiation of ESCs in vitro remains largely unknown [20,21].…”

Section: Introductionmentioning

confidence: 99%

Osteogenic Effect of Inducible Nitric Oxide Synthase (iNOS)-Loaded Mineralized Nanoparticles on Embryonic Stem Cells

Lee¹,

Lee²,

Lee³

et al. 2018

Cell Physiol Biochem

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Background/Aims: This study investigated the effect of inducible nitric oxide synthase-loaded mineralized nanoparticles (iNOS-MNPs) on the osteogenic differentiation of mouse embryonic stem cells (ESCs). Methods: We prepared iNOS-MNPs using an anionic block copolymer template-mediated calcium carbonate (CaCO₃) mineralization process in the presence of iNOS. iNOS-MNPs were spherical and had a narrow size distribution. iNOS was stably loaded within MNPs without denaturation. In order to confirm the successful introduction of iNOS-MNPs into the cytosol of ESCs, intracellular levels of nitric oxide (NO) was determined with a fluorometric analysis. A NO effector molecule, cyclic guanosine 3’,5’ monophosphate (cGMP) was also quantified with a competitive enzyme immunoassay. Cell viability in response to iNOS-MNP treatment was determined using the cell counting kit-8 (CCK-8) assay. Alkaline phosphatase (ALP) activity assay, intracellular calcium quantification assay, and Alizarin red S staining for matrix mineralization were performed to investigate osteogenic differentiation of ESCs. The protein levels of Runt-related transcription factor 2 (RUNX2), osteocalcin (OCN), and osterix (OSX) as osteogenic-related factors were also assessed by immunofluorescence staining and Western blot analysis. The complex pathways associated with iNOS-MNP-derived osteogenic differentiation of ESCs were evaluated by network-based analysis. Results: Cells with iNOS-MNPs displayed a significant increase in NO and cGMP concentration compared with the control group. When cells were exposed to iNOS-MNPs, there were no adverse effects on cell viability. Importantly, iNOS-MNP uptake promoted the osteogenic differentiation of ESCs. Using transcriptome profiling, we obtained 1,836 differentially-induced genes and performed functional enrichment analysis with ClueGO and KEGG. These analyses identified significantly enriched and interconnected molecular pathways such as protein kinase activity, estrogen receptor activity, bone morphogenetic protein (BMP) receptor binding, ligand-gated ion channel activity, and phosphatidylinositol 3-phosphate binding. Conclusion: These findings suggest that iNOS-MNPs can induce osteogenic differentiation in ESCs by integrating complex signaling pathways.

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“…Conversely, it has been shown that NO may act as a positive modulator of ␤-catenin activity under low basal Wnt activation. For instance, expression of iNOS (NOS2) in embryonic stem cells influences cell differentiation and early lineage commitment through activation of ␤-catenin (46). Another study showed that ␤-catenin associates with eNOS in quiescent ECs and that eNOS activation causes cGMP-dependent nuclear translocation of ␤-catenin and influences angiogenesis (47).…”

Section: Discussionmentioning

confidence: 99%

Endothelial NO Synthase-Dependent S-Nitrosylation of β-Catenin Prevents Its Association with TCF4 and Inhibits Proliferation of Endothelial Cells Stimulated by Wnt3a

Zhang

Chidiac

Delisle

et al. 2017

Molecular and Cellular Biology

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Nitric oxide (NO) produced by endothelial NO synthase (eNOS) modulates many functions in endothelial cells. S-nitrosylation (SNO) of cysteine residues on ␤-catenin by eNOS-derived NO has been shown to influence intercellular contacts between endothelial cells. However, the implication of SNO in the regulation of ␤-catenin transcriptional activity is ill defined. Here, we report that NO inhibits the transcriptional activity of ␤-catenin and endothelial cell proliferation induced by activation of Wnt/␤-catenin signaling. Interestingly, induction by Wnt3a of ␤-catenin target genes, such as the axin2 gene, is repressed in an eNOSdependent manner by vascular endothelial growth factor (VEGF). We identified Cys466 of ␤-catenin as a target for SNO by eNOS-derived NO and as the critical residue for the repressive effects of NO on ␤-catenin transcriptional activity. Furthermore, we observed that Cys466 of ␤-catenin, located at the binding interface of the ␤-catenin-TCF4 transcriptional complex, is essential for disruption of this complex by NO. Importantly, Cys466 of ␤-catenin is necessary for the inhibitory effects of NO on Wnt3a-stimulated proliferation of endothelial cells. Thus, our data define the mechanism responsible for the repressive effects of NO on the transcriptional activity of ␤-catenin and link eNOS-derived NO to the modulation by VEGF of Wnt/␤-catenin-induced endothelial cell proliferation.KEYWORDS VEGF, Wnts, cell signaling, endothelial cells, nitric oxide, nitric oxide synthase, S-nitrosylation T he Wnt/␤-catenin signaling pathway is involved in physiological and pathological angiogenesis, the process of blood vessel formation from preexisting vasculature, through its transcriptional regulation (1-3). Canonical Wnt proteins, namely, Wnt1, Wnt3a, Wnt7a, Wnt7b, and Wnt10b, have been demonstrated to activate ␤-catenin signaling in endothelial cells (ECs) and to regulate target genes that affect the angiogenic process (3-6). In addition to its essential structural role in cadherin-based adhesions, ␤-catenin is a regulator of Wnt-mediated gene expression through its association in the nucleus with the T-cell factor/lymphoid-enhancing factor (TCF/LEF) (7-9). In the absence of Wnt stimulation, free cytoplasmic ␤-catenin levels are kept low by a degradation complex that includes glycogen synthase kinase 3␤ (GSK3␤), adenomatous polyposis coli (APC), and axin (10-12). Upon Wnt stimulation, ␤-catenin is stabilized in the cytoplasm and subsequently translocalized to the nucleus, interacts with TCF/LEF, and enhances the expression of genes involved in cell proliferation, differentiation, cell fate, and survival (6, 13-15).In ECs, growth factors such as vascular endothelial growth factor (VEGF) regulate Citation Zhang Y, Chidiac R, Delisle C, Gratton J-P. 2017. Endothelial NO synthase-dependent S-nitrosylation of β-catenin prevents its association with TCF4 and inhibits proliferation of endothelial cells stimulated by Wnt3a. Mol

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NO/beta-catenin crosstalk modulates primitive streak formation prior to embryonic stem cell osteogenic differentiation

Cited by 33 publications

References 87 publications

Tualang Honey Improves Human Corneal Epithelial Progenitor Cell Migration and Cellular Resistance to Oxidative Stress In Vitro

Tualang Honey Improves Human Corneal Epithelial Progenitor Cell Migration and Cellular Resistance to Oxidative Stress In Vitro

Osteogenic Effect of Inducible Nitric Oxide Synthase (iNOS)-Loaded Mineralized Nanoparticles on Embryonic Stem Cells

Endothelial NO Synthase-Dependent S-Nitrosylation of β-Catenin Prevents Its Association with TCF4 and Inhibits Proliferation of Endothelial Cells Stimulated by Wnt3a

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