Overexpression Of Nanog Research Articles

Single-Cell Analysis of Immune Synapse Formation Associated with Melanoma-T Cell Contact

Extracellular vesicles (EVs) derived from an anaplastic mouse melanoma cell line made by Nanog overexpression of F10 (Nanog +F10 cells) suppressed the metastasis of Nanog +F10 cells. We suspected the involvement of immune cell system and focused on the cytotoxic activity of T cells. T cells are reported to form immune synapses when they come into contact with cancer cells and promote the introduction of cell-killing factors into cancer cells from the sites near immune synapses. However, the dynamic process of formation and loss of immune synapses is still unclear. Especially it is thought to differ depending on the type and the stage of cancer. Therefore, we believe that it is important to analyze the dynamic behavior of immune synapses in order to elucidate their possible role in the metastasis suppression mechanism of Nanog +F10-EVs. In this study, a cytotoxic T cell line (CTLL-2) was used as a T cell model. The purpose of this study was to perform microscopic fluorescence analysis of immune synapses that are expected to be generated when CTLL-2 cells are loaded with Nanog +F10-EVs and then co-cultured with Nanog +F10 cells. We focused on PKCθ, one of the proteins constituting immunological synapses. First, Nanog +F10-EVs were extracted from Nanog +F10 cells by ultracentrifugation. The Nanog +F10-EVs or PBS was added to CTLL-2 cells and cultured overnight. Then, Nanog +F10 cells were co-cultured with the previously treated CTLL-2 cells on poly-L-lysine-coated glass slides. Cells were then fixed in 4% PFA/PBS, permeabilized with 0.1% Triton X-100, and blocked with 3% BSA/PBS. Immunostaining was then performed with drops of PE-labeled PKCθ antibody and Alexa Fluor® 647-labeled Nanog antibody prepared in 1.5% BSA/PBS to a concentration of 10 µg/mL. Observations were made using a confocal microscope LSM510 to observe the contact between CTLL-2 cells and Nanog +F10 cells. The results showed that PKCθ was localized at the plasma membrane in Nanog +F10 cells as expected, but its expression was weak in CTLL-2 cells. When multiple cells formed clusters, the localization was even more pronounced along the cell-cell contact surfaces, with areas of elevated PKCθ expression found on the Nanog +F10 cell side at sites where CTLL-2 cells were likely in contact. On the other hand, in the case of Nanog +F10-EVs loaded CTLL-2 cells, the observed fluorescence intensity suggests that the expression of PKCθ in CTLL-2 cells increased. Furthermore, the expression of PKCθ tended to increase when CTLL-2 cells and Nanog +F10 cells were co-cultured for longer periods of time. It was found that the cytotoxic activity of CTLL-2 cells may be related to this increased expression of PKCθ.

CD57 defines a novel cancer stem cell that drive invasion of diffuse pediatric-type high grade gliomas.

Diffuse invasion remains a primary cause of treatment failure in pediatric high-grade glioma (pHGG). Identifying cellular driver(s) of pHGG invasion is needed for anti-invasion therapies. Ten highly invasive patient-derived orthotopic xenograft (PDOX) models of pHGG were subjected to isolation of matching pairs of invasive (HGGINV) and tumor core (HGGTC) cells. pHGGINV cells were intrinsically more invasive than their matching pHGGTC cells. CSC profiling revealed co-positivity of CD133 and CD57 and identified CD57+CD133- cells as the most abundant CSCs in the invasive front. In addition to discovering a new order of self-renewal capacities, i.e., CD57+CD133- > CD57+CD133+ > CD57-CD133+ > CD57-CD133- cells, we showed that CSC hierarchy was impacted by their spatial locations, and the highest self-renewal capacities were found in CD57+CD133- cells in the HGGINV front (HGGINV/CD57+CD133- cells) mediated by NANOG and SHH over-expression. Direct implantation of CD57+ (CD57+/CD133- and CD57+/CD133+) cells into mouse brains reconstituted diffusely invasion, while depleting CD57+ cells (i.e., CD57-CD133+) abrogated pHGG invasion. We revealed significantly increased invasive capacities in HGGINV cells, confirmed CD57 as a novel glioma stem cell marker, identified CD57+CD133- and CD57+CD133+ cells as a new cellular driver of pHGG invasion and suggested a new dual-mode hierarchy of HGG stem cells.

NANOG regulate the JAK/STAT3 pathway to promote trophoblast cell migration and epithelial-mesenchymal transition (EMT) in hypertensive disorders of pregnancy (HDP) through protein interaction with CDK1.

Hypertensive disorders of pregnancy (HDP) are a common pregnancy disease. NANOG and Cyclin-dependent kinase 1 (CDK1) are essential for regulating the function of cell proliferation and apoptosis. However, the mechanism of action in HDP is yet unclear. The microarray dataset GSE6573 was downloaded from the GEO database. Emt-related gene set was downloaded from Epithelial-Mesenchymal Transition gene database 2.0 were screened differentially expressed genes by bioinformatics analysis. Pathway Commons and Scansite 4.0 databases were used to predict the interaction between proteins. Placental tissue samples were collected from HDP patients and patients with uneventful pregnancies. RT-qPCR, Western blot and immunohistochemistry were used to detect the expression of NANOG, CDK1, MMP-2, MMP-9, EMT markers and the JAK/STAT3 pathway proteins. Transfection NANOG overexpression/knockdown, and CDK1 knockdown into the human chorionic trophoblast cells (HTR-8/Svneo). CCK-8, Transwell and Wound-healing assay were used to evaluate cell proliferation, invasion and migration. CO-IP and GST pull-down assays were used to confirm the protein interaction. A total obtained seven EMT-related differentially expressed genes, wherein NANOG, NODAL and LIN28A had protein interaction. In the HDP patients' tissue found that NANOG and CDK1 had lower expression. NANOG overexpression promoted HTR-8/Svneo proliferation, migration and EMT, while NANOG knockdown had the opposite effect. Further a protein interaction between STAT3 and CDK1 with NANOG. NANOG overexpression downregulated the JAK/STAT3 pathway to promote HTR-8/Svneo proliferation, migration and EMT, which was reversed by CDK1 knockdown. NANOG downregulated the JAK/STAT3 pathway to promote trophoblast cell proliferation, migration and EMT through protein interaction with CDK1.

Gastrulation-stage gene expression in Nipbl+/- mouse embryos foreshadows the development of syndromic birth defects.

In animal models, Nipbl deficiency phenocopies gene expression changes and birth defects seen in Cornelia de Lange syndrome, the most common cause of which is Nipbl haploinsufficiency. Previous studies in Nipbl+/- mice suggested that heart development is abnormal as soon as cardiogenic tissue is formed. To investigate this, we performed single-cell RNA sequencing on wild-type and Nipbl+/- mouse embryos at gastrulation and early cardiac crescent stages. Nipbl+/- embryos had fewer mesoderm cells than wild-type and altered proportions of mesodermal cell subpopulations. These findings were associated with underexpression of genes implicated in driving specific mesodermal lineages. In addition, Nanog was found to be overexpressed in all germ layers, and many gene expression changes observed in Nipbl+/- embryos could be attributed to Nanog overexpression. These findings establish a link between Nipbl deficiency, Nanog overexpression, and gene expression dysregulation/lineage misallocation, which ultimately manifest as birth defects in Nipbl+/- animals and Cornelia de Lange syndrome.

Prediction of synergistic gemcitabine-based combination treatment through a novel tumor stemness biomarker NANOG in pancreatic cancer.

Gemcitabine remains a first-class chemotherapeutic drug for pancreatic cancer. However, due to the rapid development of gemcitabine resistance in pancreatic cancer, gemcitabine alone or in combination with other anti-cancer drugs only showed limited effect in the clinic. It is extremely challenging to effectively and efficiently determine the optimal drug regimens. Thus, identification of appropriate prediction biomarkers is critical for the rational design of gemcitabine-based therapeutic options. Herein, a pancreatic cancer stem cell (PCSC) model exhibiting chemoresistance to gemcitabine was used to test the activity of clinical cancer drugs in the presence or absence of gemcitabine. As determined by combinatorial treatment, several types of drugs resensitized gemcitabine-resistant PCSCs to gemcitabine, with sorafenib (EGFR inhibitor)/gemcitabine and sunitinib (TBK1 inhibitors)/gemcitabine drug combinations being the most preferred treatments for PCSCs. Following the validation of the PCSC model by an antibody array test of 15-gene expression of stemness biomarkers, NANOG showed markedly different expression in PCSCs compared to the parental cells. From comprehensive analysis of stem cell index versus combination index, a stemness-related correlation model was successfully constructed to demonstrate the correlation between NANOG expression and synergism. Cancer cell stemness was ascertained to be highly relevant to NANOG overexpression that can be abrogated by synergized gemcitabine-drug combinations. Therefore, NANOG works as a therapeutic biomarker for predicating efficient combinatorial treatment of gemcitabine in pancreatic cancer.

Comparative Study of Metastasis Suppression Effects of Extracellular Vesicles Derived from Anaplastic Cell Lines, Nanog-Overexpressing Melanoma, and Induced Pluripotent Stem Cells.

Previous studies have demonstrated that extracellular vesicles (EVs) derived from an anaplastic mouse melanoma cell line made using Nanog overexpression of F10 (Nanog+F10) suppressed the metastasis of Nanog+F10. Here, an induced pluripotent stem (iPS) cell line was focused as a more anaplastic cell line, potentially producing EVs with higher metastasis-suppressive effects. The EVs were introduced into the tail vein nine times before introducing Nanog+F10 cells. Two weeks later, the liver and lung were resected and metastatic colonies were quantified. The involvement of macrophages (invasion inhibiting ability, phagocytic activity) and cytotoxic T cells (cytotoxicity) was evaluated using J774.1 and CTLL-2 cell lines. iPS EVs showed similar level effects to Nanog+F10 EVs in every item relevant to metastasis suppression. Differential expression analysis of miRNAs in EVs and functional network database analysis revealed that dominant regulatory miRNAs were predicted. The candidate hub genes most highly associated with the metastasis suppression mechanism were predicted as six genes, including Trp53 and Hif1a, for Nanog+F10 EVs and ten genes, including Ins1 and Kitl, for iPS EVs. Regarding the mechanism, Nanog+F10 EVs and iPS EVs were very different. This suggests synergistic effect when used together as metastasis preventive vaccine.

Targeting Nanog expression increased Cisplatin chemosensitivity and inhibited cell migration in Gastric cancer cells

Regardless of significant advances in cancer treatment, gastric cancer (GC) incidence rate is increasing worldwide. As one of the main transcription factors participating in stemness, Nanog plays a pivotal role in various aspects of tumorigenesis, metastasis, and chemosensitivity. Given that, the current research intended to evaluate the potential effects of Nanog suppression on the GC cell Cisplatin chemosensitivity and in vitro tumorigenesis. First, bioinformatics analysis was performed to evaluate the effect of Nanog expression on GC patients' survival. The MKN-45 human GC cells were transfected with specific siRNA targeting Nanog and/or treated with Cisplatin. Then, to study cellular viability and apoptosis, MTT assay and Annexin V/PI staining were done, respectively. Also, the scratch assay was performed to investigate cell migration, and MKN-45 cell stemness was followed using colony formation assay. Western blotting and qRT-PCR were used for gene expression analysis. The findings demonstrated that Nanog overexpression was significantly correlated with poor survival of GC patients, and siRNA-mediated Nanog silencing strongly increased MKN-45 cell sensitivity to Cisplatin through apoptosis induction. Also, Nanog suppression combined with Cisplatin resulted in the upregulation of the Caspase-3 and Bax/Bcl-2 ratio at mRNA levels and increased Caspase-3 activation. Moreover, reduced expression of Nanog, separately or combined with Cisplatin, inhibited MKN-45 cell migration by downregulating MMP2 mRNA and protein expression levels. The results also evidenced CD44 and SOX-2 downregulation aligned with a decreased rate of MKN-45 cell colony formation ability through treatments. Besides, Nanog downregulation significantly decreased MDR-1 mRNA expression. Taken together, the results of this study indicated that Nanog could be suggested as a promising target combined with Cisplatin-based GC therapies for reducing drug side effects and improving patients’ outcomes.

TRRAP Enhances Cancer Stem Cell Characteristics by Regulating NANOG Protein Stability in Colon Cancer Cells.

NANOG, a stemness-associated transcription factor, is highly expressed in many cancers and plays a critical role in regulating tumorigenicity. Transformation/transcription domain-associated protein (TRRAP) has been reported to stimulate the tumorigenic potential of cancer cells and induce the gene transcription of NANOG. This study aimed to investigate the role of the TRRAP-NANOG signaling pathway in the tumorigenicity of cancer stem cells. We found that TRRAP overexpression specifically increases NANOG protein stability by interfering with NANOG ubiquitination mediated by FBXW8, an E3 ubiquitin ligase. Mapping of NANOG-binding sites using deletion mutants of TRRAP revealed that a domain of TRRAP (amino acids 1898-2400) is responsible for binding to NANOG and that the overexpression of this TRRAP domain abrogated the FBXW8-mediated ubiquitination of NANOG. TRRAP knockdown decreased the expression of CD44, a cancer stem cell marker, and increased the expression of P53, a tumor suppressor gene, in HCT-15 colon cancer cells. TRRAP depletion attenuated spheroid-forming ability and cisplatin resistance in HCT-15 cells, which could be rescued by NANOG overexpression. Furthermore, TRRAP knockdown significantly reduced tumor growth in a murine xenograft transplantation model, which could be reversed by NANOG overexpression. Together, these results suggest that TRRAP plays a pivotal role in the regulation of the tumorigenic potential of colon cancer cells by modulating NANOG protein stability.

Cell-type dependent regulation of pluripotency and chromatin remodeling genes by hydralazine

BackgroundThe generation of induced pluripotent stem cells has opened the field of study for stem cell research, disease modeling and drug development. However, the epigenetic signatures present in somatic cells make cell reprogramming still an inefficient process. This epigenetic memory constitutes an obstacle in cellular reprogramming. Here, we report the effect of hydralazine (HYD) and valproic acid (VPA), two small molecules with proven epigenetic activity, on the expression of pluripotency genes in adult (aHF) and neonatal (nbHF) human fibroblasts.MethodsaHF and nbHF were treated with HYD and/or VPA, and viability and gene expression assays for OCT4, NANOG, c-MYC, KLF4, DNMT1, TET3, ARID1A and ARID2 by quantitative PCR were performed. aHF and nbHF were transfected with episomal plasmid bearing Yamanaka factors (OCT4, SOX2, KLF4 and c-MYC) and exposed to HYD and VPA to determine the reprogramming efficiency. Methylation sensitive restriction enzyme (MSRE) qPCR assays were performed on OCT4 and NANOG promoter regions. Immunofluorescence assays were carried out for pluripotency genes on iPSC derived from aHF and nbHF.ResultsHYD upregulated the expression of OCT4 (2.5-fold) and NANOG (fourfold) genes but not c-Myc or KLF4 in aHF and had no significant effect on the expression of all these genes in nbHF. VPA upregulated the expression of NANOG (twofold) in aHF and c-MYC in nbHF, while it downregulated the expression of NANOG in nbHF. The combination of HYD and VPA canceled the OCT4 and NANOG overexpression induced by HYD in aHF, while it reinforced the effects of VPA on c-Myc expression in nbHF. The HYD-induced overexpression of OCT4 and NANOG in aHDF was not dependent on demethylation of gene promoters, and no changes in the reprogramming efficiency were observed in both cell populations despite the downregulation of epigenetic genes DNMT1, ARID1A, and ARID2 in nbHF.ConclusionsOur data provide evidence that HYD regulates the expression of OCT4 and NANOG pluripotency genes as well as ARID1A and ARID2 genes, two members of the SWI/SNF chromatin remodeling complex family, in normal human dermal fibroblasts.

Modulation of Spheroid Forming Capacity and TRAIL Sensitivity by KLF4 and Nanog in Gastric Cancer Cells.

The expression of pluripotency factors, and their associations with clinicopathological parameters and drug response have been described in various cancers, including gastric cancer. This study investigated the association of pluripotency factor expression with the clinicopathological characteristics of gastric cancer patients, as well as changes in the expression of these factors upon the stem cell-enriching spheroid culture of gastric cancer cells, regulation of sphere-forming capacity, and response to cisplatin and TRAIL treatments by Nanog and KLF4. Nanog expression was significantly associated with the emergence of a new tumor and a worse prognosis in gastric cancer patients. The expression of the pluripotency factors varied among six gastric cancer cells. KLF4 and Nanog were expressed high in SNU-601, whereas SOX2 was expressed high in SNU-484. The expression of KLF4 and SOX2 was increased upon the spheroid culture of SNU-601 (KLF4/Nanog-high) and SNU-638 (KLF4/Nanog-low). The spheroid culture of them enhanced TRAIL-induced viability reduction, which was accompanied by the upregulation of death receptors, DR4 and DR5. Knockdown and overexpression of Nanog in SNU-601 and SNU-638, respectively, did not affect spheroid-forming capacity, however, its expression was inversely correlated with DR4/DR5 expression and TRAIL sensitivity. In contrast, KLF4 overexpression in SNU-638 increased spheroid formation, susceptibility to cisplatin and TRAIL treatments, and DR4/DR5 expression, while the opposite was found in KLF4-silenced SNU-601. KLF4 is supposed to play a critical role in DR4/DR5 expression and responses to TRAIL and cisplatin, whereas Nanog is only implicated in the former events only. Direct regulation of death receptor expression and TRAIL response by KLF4 and Nanog have not been well documented previously, and the regulatory mechanism behind the process remains to be elucidated.

Diallyl Trisulfide Suppresses the Renal Cancer Stem-like Cell Properties via Nanog

Cancer stem-like cells (CSCs), which play an important role in tumor initiation and progression, have been identified in many cancers. Diallyl trisulfide (DATS) is an organosulfur compound extracted from garlic with anticancer activities. Nanog is a transcription factor responsible for maintaining the stemness of CSCs, but its role in the DATS-induced attenuation of renal CSC properties is unknown. In this study, renal CSCs were enriched from human renal cancer cell lines 786-O and ACHN cultured in a serum-free medium (SFM). The properties of CSCs were analyzed by evaluating the ability of the cells in sphere formation and measuring the expression of stem cell markers. We found that downregulation of Nanog inhibited renal CSC properties. DATS suppressed renal CSC activities by reducing tumorsphere formation, decreasing stem cell markers including Nanog, CD44, ALDH1A1, and Oct4, inhibiting cell proliferation and promoting apoptosis. We further revealed that overexpression of Nanog reversed the suppressive effects of DATS on renal CSCs. Taken together, our results demonstrated that DATS inhibited renal CSCs by suppressing Nanog. These novel findings suggested that, through Nanog targeting, DATS can potentially be used as an anti-tumor agent for renal cancer.

Novel Roles of Nanog in Cancer Cells and Their Extracellular Vesicles

The use of extracellular vesicle (EV)-based vaccines is a strategically promising way to prevent cancer metastasis. The effective roles of immune cell-derived EVs have been well understood in the literature. In the present paper, we focus on cancer cell-derived EVs to enforce, more thoroughly, the use of EV-based vaccines against unexpected malignant cells that might appear in poor prognostic patients. As a model of such a cancer cell with high malignancy, Nanog-overexpressing melanoma cell lines were developed. As expected, Nanog overexpression enhanced the metastatic potential of melanomas. Against our expectations, a fantastic finding was obtained that determined that EVs derived from Nanog-overexpressing melanomas exhibited a metastasis-suppressive effect. This is considered to be a novel role for Nanog in regulating the property of cancer cell-derived EVs. Stimulated by this result, the review of Nanog's roles in various cancer cells and their EVs has been updated once again. Although there was no other case presenting a similar contribution by Nanog, only one case suggested that NANOG and SOX might be better prognosis markers in head and neck squamous cell carcinomas. This review clarifies the varieties of Nanog-dependent phenomena and the relevant signaling factors. The information summarized in this study is, thus, suggestive enough to generate novel ideas for the construction of an EV-based versatile vaccine platform against cancer metastasis.

Epidermal stem cells participate in the repair of scalds via Nanog and Myc regulation.

Epidermal stem cells (EpSCs) with high expression of regulatory factor Nanog can promote wound healing. The aim of the present study was to investigate the effectiveness and mechanism of epidermal stem cells (EpSCs) in healing scalds and the underlying molecular mechanism. Mouse EpSCs were isolated from skin tissues and cultured invitro. First, the proliferative ability of EpSCs was determined via the upregulation and downregulation of Nanog expression levels in EpSCs using the MTS‑assay. Second, a wound healing assay of the EpSCs with different Nanog expression levels was performed to investigate cell migratory capacities. Third, the protein expression levels of various proteins in EpSCs with Nanog overexpression or knockdown, were determined. Finally, the transfected EpSCs were applied to the rat scald model to observe their effect on scald healing. Subsequently, wound scores, re‑epithelialization and capillary density were determined histologically. The results demonstrated that Nanog overexpression enhanced the proliferative ability of EpSCs via cellular (c)‑Myc. Moreover, the LV‑Nanog group of EpSCs with increased Nanog expression levels exhibited improved healing abilities in the wound healing test than control group. Using western blotting, it was demonstrated that EpSCs that were transfected with a Nanog‑overexpression vector expressed high Nanog protein expression levels, whereas small interfering RNA‑Nanog‑transfected EpSCs exhibited low Nanog protein expression levels. Furthermore, c‑Myc expression was synchronized with Nanog expression. It was also revealed that as the expression levels of c‑Myc increased, p53 expression levels also increased. In the rat scald model, Nanog‑overexpressing EpSCs enhanced wound closure and re‑epithelialization. The EpSCs with Nanog knockdown exhibited the opposite effects. The present study therefore indicated that Nanog may have a positive effect on scald healing in rats, which supports its use in EpSC‑based treatments against scalds. Furthermore, it was suggested that c‑Myc potentially serves a key role in this process and that this process avoids cancerization by relying on the supervision of p53.

NANOG regulates epithelial-mesenchymal transition via AMPK/mTOR signalling pathway in ovarian cancer SKOV-3 and A2780 cells.

NANOG engages with tumour initiation and metastasis by regulating the epithelial–mesenchymal transition (EMT) in epithelial ovarian cancer (EOC). However, its role in association with pAMPKα, and its clinical significance in EOC have not been elucidated even though AMPK is known to degrade NANOG in various human cancers. Hence, we investigated the role of pAMPKα and its association with NANOG as potential prognostic biomarkers in EOC. Both NANOG and pAMPKα expression were significantly overexpressed in EOCs comparing nonadjacent normal epithelial tissues, benign tissues, and borderline tumours. NANOG overexpression was significantly associated with poor disease‐free survival (DFS) and overall survival (OS), whereas pAMPKα overexpression was associated with good DFS and OS. Importantly, multivariate analysis revealed that the combination of high NANOG and low pAMPKα expression was a poor independent prognostic factor for DFS and was associated with platinum resistance. In ovarian cancer cell lines, siRNA‐mediated NANOG knockdown diminished migration and invasion properties by regulating the EMT process via the AMPK/mTOR signalling pathway. Furthermore, treatment with AMPK activator suppressed expression of stemness factors such as NANOG, Oct4 and Sox2. Collectively, these findings established that the combination of high NANOG and low pAMPKα expression was associated with EOC progression and platinum resistance, suggesting a potential prognostic biomarker for clinical management in EOC patients.

NANOG regulates tumor metastasis via AMPK/mTOR signaling pathway in patients with epithelial ovarian cancer (252)

<b>Objectives:</b> NANOG engages with tumor initiation and metastasis by regulating EMT (epithelial-mesenchymal transition) in epithelial ovarian cancer (EOC). However, its role in association with pAMPK and its clinical significance in EOC has not been elucidated, even though pAMPK is known to degrade NANOG in various human cancer. Therefore, here we investigated the role of pAMPK and its association with NANOG as a potential prognostic, predictive biomarker in EOC. <b>Methods:</b> Immunohistochemistry of NANOG and pAMPK in epithelial ovarian cancer tissues were performed to define the clinical significance with TMA. Moreover, ovarian cancer cell lines were used for functional studies to investigate the underlying mechanisms. <b>Results:</b> Both NANOG and pAMPK expression were significantly overexpressed in EOCs compared to nonadjacent normal epithelial tissues, benign tissues, and borderline tumors. NANOG overexpression was significantly associated with poor DFS and OS, whereas pAMPK overexpression was associated with good DFS and OS. Importantly, multivariate analysis revealed that combining high NANOG and low pAMPK expression was an independent prognostic factor for DFS and associated with poor platinum resistance. In ovarian cancer cell lines, NANOG knockdown diminished migration and invasion property by regulating the EMT process via the AMPK/mTOR signaling pathway. Furthermore, treatment of AMPK activator suppressed expression of stemness-related genes, such as <i>NANOG, Oct4</i>, and <i>Sox2</i>. <b>Conclusions:</b> Collectively, our findings established that a combination of high NANOG and low pAMPK expression was associated with EOC progression and platinum resistance, suggesting a potential predictive biomarker for clinical management in EOC patients. Also, NANOG knockdown inhibited tumor metastasis and invasion by regulating EMT via the AMPK/mTOR signaling pathway.

Expression of Sox2, Oct4, and Nanog in Human Lung Cancer Non-small-cell

Background: Cancer stem cells are a subpopulation of tumor cells with self-renewal capacity that promote tumorigenesis, resistance to chemotherapy, and metastasis. Sox2, Oct4, and Nanog are three pluripotent transcription factors expressed in embryonic stem cells and cancer stem cells. Materials and Methods: This study aimed to evaluate the expression of Sox2, Nanog, and Oct4, and analyze their clinical significance in human non-small-cell lung cancer (NSCLC). Expression of Sox2, Nanog, and Oct4 was assayed in cancer tissues and their corresponding paracancerous tissues from 30 patients with NSCLC. RT-PCR was used to analyze the expression of these genes. The correlation between the expression of these three genes and clinical parameters including disease stage, smoking, lymph node, and cancer subgroups (adenocarcinoma and squamous) were analyzed. Results: All three genes were expressed simultaneously in 76.6% of tumor samples. A significant correlation was observed between the expression of Sox2, Nanog, and Oct4 in the cancer tissues in comparison to the paracancerous tissues (P&lt;0.000). Expression of Sox2 and Oct4 gene had a positive correlation with the stage of cancer (Sox2 P=0.01, Oct4 P=0.0007), while the expression profile of Nanog showed a significant positive correlation with sex (P=0.0063), smoking (P=0.0253), tumor stages (P=0.0003), and tumor type (P=0.0085). Conclusion: Evaluating the expression of Sox2, Nanog, and Oct4 genes in NSCLC might have some implications for diagnosis and prognosis; they might be also promising treatment targets. The correlations between prognosis and pathological features and Nanog overexpression in NSCLC suggest Nanog is a potential indicator of the early stage of NSCLC.

Reprogramming neurons for regeneration: The fountain of youth.

Neurons in the central nervous system (CNS) are terminally differentiated cells that gradually lose their ability to support regeneration during maturation due to changes in transcriptomic and chromatin landscape. Similar transcriptomic changes also occur during development when stem cells differentiate into different types of somatic cells. Importantly, differentiated cells can be reprogrammed back to induced pluripotent stems cells (iPSCs) via global epigenetic remodeling by combined overexpression of pluripotent reprogramming factors, including Oct4, Sox2, Klf4, c-Myc, Nanog, and/or Lin28. Moreover, recent findings showed that many proneural transcription factors were able to convert non-neural somatic cells into neurons bypassing the pluripotent stage via direct reprogramming. Interestingly, many of these factors have recently been identified as key regulators of CNS neural regeneration. Recent studies indicated that these factors could rejuvenate mature CNS neurons back to a younger state through cellular state reprogramming, thus favoring regeneration. Here we will review some recent findings regarding the roles of genetic cellular state reprogramming in regulation of neural regeneration and explore the potential underlying molecular mechanisms. Moreover, by using newly emerging techniques, such as multiomics sequencing with big data analysis and Crispr-based gene editing, we will discuss future research directions focusing on better revealing cellular state reprogramming-induced remodeling of chromatin landscape and potential translational application.

Anti-Metastatic Function of Extracellular Vesicles Derived from Nanog-Overexpressing Melanoma.

A metastatic melanoma cell line B16-F10 (F10) was modified to a more undifferentiated state by Nanog overexpression. The produced cell line Nanog+F10 showed a higher metastatic potential than F10. Instead of whole cells, the extracellular vesicles (EVs) therefrom were investigated about their possible role as an autovaccine against metastasis. EVs from Nanog+F10 cells (Nanog+F10-EVs) could suppress the metastasis, contrasting the EVs from less metastatic F10 cells (F10-EVs) enhanced metastasis. The involvement of TGF-β1 in the role of Nanog+F10-EVs was analyzed, as TGF-β1 was a secretory cytokine being affected most intensively by Nanog overexpression. It was suggested to be crucial that the TGF-β1 concentration in Nanog+F10-EVs should be as low as 1.6 pg/μg for its metastasis-suppressive role. In response to Nanog+F10-EVs, immunoreaction was observed in liver, indicating the specific decrease in the number of tumor-promotive CD163-positive macrophages. These indicate a possibility of Nanog+F10-EVs as a novel autovaccine candidate against melanoma metastasis.

Nanog mediated by FAO/ACLY signaling induces cellular dormancy in colorectal cancer cells

Dormant cancer cells drive recurrence and drug resistance, which lead to poor prognosis in colorectal cancer (CRC). The mechanisms that regulate the entry of cancer cells into dormancy remain to be extensively studied. Nanog is a master transcription factor to maintain the self-renewal and pluripotency of stem cells. Since dormant cancer cells are similar to quiescent cancer stem cells, the correlation between dormant state and Nanog in CRC is worth to be explored. Serum deprivation is a common method to establish experimental cellular dormancy model. Here, we verified that serum deprivation-induced CRC cells to enter a cellular dormancy state, characterized by no proliferation, no death, no senescence, resistance to chemotherapy, high expression of dormant markers, metabolic suppression, and recovery to active status. Interestingly, we further identified that Nanog was upregulated in dormant CRC cells. Nanog knockdown could destroy the dormant state of serum-deprived CRC cells while Nanog overexpression could induce dormancy in CRC cells. Mechanistically, Nanog was regulated through a fatty acid oxidation (FAO)/ATP citrate lyase (ACLY)-dependent pathway. FAO increased ACLY expression to promote the synthesis of acetyl-CoA, which was transferred by P300 to accelerate H3K27 acetylation of Nanog promoter. Then, Nanog upregulation increased the transcription of P21 and P27, which promoted the dormancy of CRC cells. Our findings revealed that Nanog could induce cellular dormancy in CRC cells and unlocked a specific mechanism to govern the process.

Lack of CD44 and Sox-2 Overexpression as Two Independent Favourable Prognostic Factors in HPV Positive Patients with Oropharyngeal Cancers

Introduction: In our earlier publications, in the group of 63 patients with oropharyngeal cancer, we have found HPV16 infection (assessed by qPCR) in 25 tumours (39.7%), immunohistochemical overexpression of CD44, CD98, ALDH1/2 and Nanog in, respectively: 43 (68.2%), 30 (47.6%), 33 (52.4%), and 53 (84.1%) cancers. Analysing CD44, CD98, ALDH1/2, we have also shown that lack of CD44 overexpression indicates excellent prognosis in patients with HPV16 positivity. The aim of the present study was to compare prognostic potential of Nanog, Oct3/4, Sox-2 expression in relation to CD44, CD98, ALDH1/2 immunoreactivity (assessed by us earlier) and clinicopathological features in the subgroups of patients: with HPV16 positivity and HPV16 negativity. Methods: Status of Oct3/4 and Sox-2 expression was assessed for 63 patients with oropharyngeal cancers based on immunohistochemistry. In survival analysis, two endpoints were applied: overall survival (OS) and disease-free survival (DFS). Results: Overexpression of Oct3/4 and Sox-2 was found in 0 (0.0%) and 27 (42.9%) of patients. In the subgroup with HPV16 positivity, the DFS for patients with lack of Sox-2 overexpression was significantly (p = 0.003) higher than for patients with Sox-2 overexpression. In the subgroup with HPV16 negativity, Nanog and Sox-2 immunoexpression did not significantly influence OS and DFS. In multivariate analysis performed for the subgroup with HPV16 positivity, lack of CD44 overexpression (p = 0.012) and lack of Sox-2 overexpression (p = 0.027) were positive independent prognostic factors. Conclusion: Based on CD44 and Sox-2 immunoreactivity, it is possible to differentiate the prognosis of HPV16-positive patients with oropharyngeal cancers.