Cancer stem cell subpopulations in primary colon adenocarcinoma
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Aims The cancer stem cell concept proposes that tumor growth and recurrence is driven by a small population of cancer stem cells (CSCs). In this study we investigated the expression of induced-pluripotent stem cell (iPSC) markers and their localization in primary low-grade adenocarcinoma (LGCA) and high-grade adenocarcinoma (HGCA) and their patient-matched normal colon samples. Materials and methods Transcription and translation of iPSC markers OCT4, SOX2, NANOG, KLF4 and c-MYC were investigated using immunohistochemical (IHC) staining, RT-qPCR and in-situ hybridization (ISH). Results All five iPSC markers were detected at the transcriptional and translational levels. Protein abundance was found to be correlated with tumor grade. Based on their protein expression within the tumors, two sub-populations of cells were identified: a NANOG+/OCT4- epithelial subpopulation and an OCT4+/NANOG- stromal subpopulation. All cases were accurately graded based on four pieces of iPSC marker-related data. Conclusions This study suggests the presence of two putative sub-populations of CSCs: a NANOG+/OCT4- epithelial subpopulation and an OCT4+/NANOG- stromal subpopulation. Normal colon, LGCA and HGCA could be accurately distinguished from one another using iPSC marker expression. Once validated, novel combinations of iPSC markers may provide diagnostic and prognostic value to help guide patient management.Keywords:
Homeobox protein NANOG
KLF4
Stem cell marker
Nanog Homeobox Protein
The cancer stem cell theory hypothesizes that cancer stem cells (CSCs), which possess self-renewal and other stem cell properties, are regarded as the cause of tumor formation, recurrence and metastasis. The isolation and identification of CSCs could help to develop novel therapeutic strategies specifically targeting CSCs. In this study, we enriched gastric cancer stem cells through spheroid body formation by cultivating the human gastric cancer cell line MKN-45 in defined serum-free medium. The stemness characteristics of spheroid body-forming cells, including self-renewal, proliferation, chemoresistance, tumorigenicity of the MKN-45 spheroid body-forming cells were evaluated, and the expression levels of stemness genes and related proteins in the MKN-45 spheroid body-forming cells were assessed. Furthermore, immunofluorescence staining for the stem cell markers on spheroid body-forming cells was examined to evaluate the association between stemness factors (Oct4, Sox2, Nanog) and the proposed CSC marker CD44. Our data demonstrated that non-adherent spheroid body-forming cells from the gastric cancer cell line MKN-45 cultured in stem cell-conditioned medium possessed gastric CSC properties, such as persistent self-renewal, extensive proliferation, drug resistance, high tumorigenic capacity and overexpression of CSC-related genes and proteins (Oct4, Sox2, Nanog and CD44), compared with the parental cells. More importantly, CD44-positive cells co-expressing the pluripotency genes Oct4, Sox2 and Nanog may represent gastric CSCs. Further experiments using more refined selection criteria such as a combination of two or multiple markers would be useful to specifically identify and purify CSCs.
Homeobox protein NANOG
Stem cell marker
Nanog Homeobox Protein
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Induced pluripotent stem cells (iPSC) have a great potential, but their clinical application depends on finding strategies to abolish their tumorigenic potential. The use of Oct4, Sox2, Klf4, c-Myc and Nanog to generate iPSC demonstrated the already known importance of these genes to maintain stemness. Therefore, the presence of these genes is responsible for iPSC-derived teratomas. Similar to iPSC, P19 teratocarcinoma cell line also has characteristics of embryonic carcinoma cells and the ability to differentiate into many cell types. We separately silenced the transcription factors Oct4, Sox2, Klf4, c-Myc and Nanog in P19 cells and measured the impact of this silencing in vivo. All silenced cells generated tumors when injected in immunosuppressed mice, but silencing of Oct4, Sox2 and Klf4 generated mainly teratomas with mesoderm tissue. Our results suggest that downregulation of these transcription factors is not enough to avoid the formation of teratomas, but their silencing affect their differentiation potential.
Homeobox protein NANOG
KLF4
Rex1
POU domain
P19 cell
LIN28
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Guo Y, Liu S, Wang P, Zhao S, Wang F, Bing L, Zhang Y, Ling E-A, Gao J & Hao A(2011) Histopathology59, 763–775 Expression profile of embryonic stem cell-associated genes Oct4, Sox2 and Nanog in human gliomas Aims: To investigate whether Oct4, Sox2 and Nanog, three core regulatory factors maintaining pluripotency and self-renewal of embryonic stem cells (ESCs), are coexpressed in human gliomas, and whether their expression might be linked to carcinogenesis and the formation of cancer stem cells (CSCs). Methods and results: Forty cases of human glioma were examined. The expression of Oct4, Sox2 and Nanog was analysed by immunohistochemistry, reverse transcription polymerase chain reaction and western blot. We found a positive correlation between the expression levels of Oct4, Sox2 and Nanog and tumour malignancy. Immunohistochemistry showed that Oct4 and Nanog were expressed in both the nuclei and the cytoplasm of glioma cells, whereas Sox2 was expressed only in the nuclei. Double immunofluorescence staining revealed that a majority of Oct4-positive cells coexpressed Sox2 and Nanog. More than 50% of Oct4-positive cells coexpressed the putative CSC markers CD133 and Nestin. Moreover, some cells exhibited Oct4 and Nanog immunoexpression in the cytoplasm, but the frequency of positive cells did not correlate with tumour malignancy. Conclusions: The present findings suggest that ESC-associated pathways are activated in human gliomas and that these may be involved in glioma progression, a role that is distinct from that in ESCs.
Homeobox protein NANOG
Nanog Homeobox Protein
Rex1
Nestin
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Citations (175)
The expression of the transcription factors Oct4, Sox2, and Nanog is commonly associated with pluripotency of mouse embryonic stem (ES) cells. However, recent observations suggest that ES cell populations are heterogeneous with respect to the expression of Nanog and that individual ES cells reversibly change their Nanog expression level. Furthermore, it has been shown that cells exhibiting a low Nanog level are more likely to undergo differentiation. Applying a novel mathematical transcription factor network model we explore mechanisms and feedback regulations to describe the observed variation of the Nanog levels in mouse ES cells. In particular we show that these variations can occur under different assumptions yielding similar experimental characteristics. Based on model predictions we propose experimental strategies to distinguish between these explanations. Concluding from our results we argue that the heterogeneity with respect to the Nanog concentrations is most likely a functional element to control the differentiation propensity of an ES cell population. Furthermore, we provide a conceptual framework that consistently explains Nanog variability and a potential "gate-keeper" function of Nanog expression with respect to the control of ES cell differentiation.
Homeobox protein NANOG
Rex1
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Citations (93)
NANOG, as a key regulator of pluripotency and acting synergistically with other factors, has been described as a crucial transcription factor in various types of cancer. In meningiomas the expression of this marker has not yet been described. With our study, we aimed to identify and localize NANOG and other possible markers of pluripotency, stem cell properties and differentiation in meningioma tissue, to elucidate a possible effect on tumorigenesis. The gene expression levels of NANOG ( NANOG1 and NANOGP8 ), SOX2 , OCT4 , KLF4 , ABCG2 , CMYC , MSI1 , CD44 , NOTCH1 , NES , SALL4B , TP53 , and EPAS1 were quantitatively examined using RT‐qPCR in 33 surgical specimens of low‐ (WHO grade I) as well as in high‐grade (WHO grade II/III) meningiomas with dural tissue as reference. Immunofluorescence co‐localization analysis following confocal fluorescence microscopy for NANOG, OCT4, SOX2, Nestin, KI‐67, and CD44 was also performed. There was a significant overexpression of NANOG , MSI1 , and EPAS1 and a downregulation of NES in all examined tumors. Subgroup analysis (WHO grade I versus grade II/III) revealed differences in the expression of NANOG , CD44 , and MSI1 . We found 1% NANOG‐positive (NANOG+) cells in low‐grade and 2% in grade II/III meningiomas co‐expressing the other mentioned markers in various compositions. In particular, NANOG+ cells expressing SOX2 and OCT4 were successfully identified (26% low‐grade versus 20% high‐grade). Our data reveal an overexpression of NANOG and other markers of pluripotency and stemness in meningiomas. Such potentially pluripotent “stem cell‐like” cells may have an impact on tumorigenesis and progression in human meningiomas.
Homeobox protein NANOG
Rex1
KLF4
Stem cell marker
Nestin
Nanog Homeobox Protein
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Homeobox protein NANOG
Nanog Homeobox Protein
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Citations (212)
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.
Homeobox protein NANOG
KLF4
Rex1
Nanog Homeobox Protein
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Given that regeneration and repair of tissues following injury require trafficking and differentiation of both resident and non‐resident progenitor stem cells, we tested the hypothesis that adult lungs harbor a limited subset of progenitor adult stem cells. To address this hypothesis, we used adult mice as a source for several tissues including the heart and the lungs. To identify and characterize presence of "candidate" primitive undifferentiated (progenitor) cells in adult tissues, we used Oct3/4, Klf4, Sox2, and Nanog transcription factors as markers. We used RT‐PCR, western blotting and confocal microscopy to determine the expression of these four transcription factors. RT‐PCR and western blotting experiment showed presence of a subset of Klf4+ and Nanog+ primitive cells. The number of Klf4+ and Nanog+ progenitor cells increased in the mice lungs that were challenged with bacterial endotoxin Lipopolysaccharide (LPS, 7.5 mg/kg body weight). These data indicate that adult tissues including lungs harbor a subset of Klf4 and Nanog expressing resident and non‐resident progenitor cells. These findings raise the intriguing possibility of previously unrealized existence of Nanog and Klf4 expressing adult progenitor cells that appear to respond to critical factors, and may contribute to regeneration and repair of tissue post‐injuries. Thus, further investigation into the function of Nanog and Klf4 in vivo may provide insights into the molecular mechanisms involved in the regeneration and repair of tissues in adult.
Homeobox protein NANOG
KLF4
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Recent studies suggest that the regulatory networks controlling the functions of stem cells during development may be abnormally active in human cancers. An embryonic stem cell (ESC) gene signature was found to correlate with a more undifferentiated phenotype of several human cancer types including gliomas, and associated with poor prognosis in breast cancer. In the present study, we used tissue microarrays of 80 low-grade (WHO Grade II) and 98 high-grade human gliomas (WHO Grades III and IV) to investigate the presence of the ESC-related proteins Nanog, Klf4, Oct4, Sox2 and c-Myc by immunohistochemistry. While similar patterns of co-expressed proteins between low- and high-grade gliomas were present, we found up-regulated protein levels of Nanog, Klf4, Oct4 and Sox2 in high-grade gliomas. Survival analysis by Kaplan-Meier analysis revealed a significant shorter survival in the subgroups of low-grade astrocytomas (n = 42) with high levels of Nanog protein (p = 0.0067) and of Klf4 protein (p = 0.0368), in high-grade astrocytomas (n = 85) with high levels of Nanog (p = 0.0042), Klf4 (p = 0.0447), and c-Myc (p = 0.0078) and in glioblastomas only (n = 71) with high levels of Nanog (p = 0.0422) and of c-Myc (p = 0.0256). In the multivariate model, Nanog was identified as an independent prognostic factor in the subgroups of low-grade astrocytomas (p = 0.0039), high-grade astrocytomas (p = 0.0124) and glioblastomas only (p = 0.0544), together with established clinical variables in these tumors. These findings provide further evidence for the joint regulatory pathways of ESC-related proteins in gliomas and identify Nanog as one of the key players in determining clinical outcome of human astrocytomas.
Homeobox protein NANOG
KLF4
Nanog Homeobox Protein
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The regulatory network of factors that center on master transcription factors such as Oct4, Nanog, and Sox2 help maintain embryonic stem (ES) cells and ensure their pluripotency. The target genes of these master transcription factors define the ES cell transcriptional landscape. In this study, we report our findings that Dido1, a target of canonical transcription factors such as Oct4, Sox2, and Nanog, plays an important role in regulating ES cell maintenance. We found that depletion of Dido1 in mouse ES cells led to differentiation, and ectopic expression of Dido1 inhibited differentiation induced by leukemia inhibitory factor withdrawal. We further demonstrated that whereas Nanog and Oct4 could occupy the Dido1 locus and promote its transcription, Dido1 could also target to the loci of pluripotency factors such as Nanog and Oct4 and positively regulate their expression. Through this feedback and feedforward loop, Dido1 is able to regulate self-renewal of mouse ES cells The regulatory network of factors that center on master transcription factors such as Oct4, Nanog, and Sox2 help maintain embryonic stem (ES) cells and ensure their pluripotency. The target genes of these master transcription factors define the ES cell transcriptional landscape. In this study, we report our findings that Dido1, a target of canonical transcription factors such as Oct4, Sox2, and Nanog, plays an important role in regulating ES cell maintenance. We found that depletion of Dido1 in mouse ES cells led to differentiation, and ectopic expression of Dido1 inhibited differentiation induced by leukemia inhibitory factor withdrawal. We further demonstrated that whereas Nanog and Oct4 could occupy the Dido1 locus and promote its transcription, Dido1 could also target to the loci of pluripotency factors such as Nanog and Oct4 and positively regulate their expression. Through this feedback and feedforward loop, Dido1 is able to regulate self-renewal of mouse ES cells
Homeobox protein NANOG
Nanog Homeobox Protein
Rex1
Ectopic expression
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