scholarly journals Valproic Acid Inhibits Proliferation and Reduces Invasiveness in Glioma Stem Cells Through Wnt/β Catenin Signalling Activation

Genes ◽  
2018 ◽  
Vol 9 (11) ◽  
pp. 522 ◽  
Author(s):  
Gabriele Riva ◽  
Chiara Cilibrasi ◽  
Riccardo Bazzoni ◽  
Massimiliano Cadamuro ◽  
Caterina Negroni ◽  
...  
Keyword(s):  
Stem Cells ◽  
Valproic Acid ◽  
Histone Deacetylase Inhibitors ◽  
Cell Signalling ◽  
Luciferase Reporter ◽  
Signalling Pathways ◽  
Boyden Chamber ◽  
Glioma Stem Cells ◽  
Bioinformatics Analyses ◽  
E Cadherin

Glioblastoma is the most common malignant brain tumour in adults. The failure of current therapies can be ascribed to glioma stem cells (GSCs), which can rapidly repopulate the tumour following the initial treatment. The study of histone deacetylase inhibitors, such as valproic acid (VPA), is becoming an attractive field in cancer research. However, the exact mechanisms underlying its anti-cancer effect remain to be elucidated due to its pleiotropic effects on several cell-signalling pathways. Ingenuity Pathway Analysis (IPA) bioinformatics analysis was performed on genome-wide data regarding GSCs methylome to identify the signalling pathways mainly affected by methylation changes induced by VPA. Real time PCR and luciferase reporter assay were used to better investigate VPA effects on Wnt/β-catenin signalling pathway. VPA effect on GSC proliferation was evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) and Trypan blue assays. Finally, VPA impact on GSC motility was demonstrated by Boyden chamber assay and further confirmed evaluating the expression levels or localisation, through western blot or immunofluorescence, of Twist1, Snail1, E-Cadherin and N-Cadherin. The bioinformatics analyses performed on GSCs methylome highlighted that Wnt/β-catenin signalling was affected by the methylation changes induced by VPA, which could influence its activation status. In particular, we pointed out a general activation of this pathway after VPA exposure, which was accompanied by an inhibitory potential on GSCs proliferation. Finally, we also proved VPA’s ability to inhibit GSCs invasion through Snail1 and Twist1 downregulation and E-Cadherin relocalisation. VPA treatment may represent a new, interesting therapeutic approach to affect GSC proliferation and motility, but further investigations are certainly needed.

scholarly journals The U2AF2 /circRNA ARF1/miR-342–3p/ISL2 feedback loop regulates angiogenesis in glioma stem cells

2020 ◽  
Vol 39 (1) ◽  
Author(s):  
Yang Jiang ◽  
Jinpeng Zhou ◽  
Junshuang Zhao ◽  
Haiying Zhang ◽  
Long Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Development ◽  
Feedback Loop ◽  
Rna Binding ◽  
Patient Survival ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Glioma Stem Cells ◽  
Poor Patient ◽  
Poor Patient Survival

Abstract Background Glioma is the most common and lethal primary brain tumor in adults, and angiogenesis is one of the key factors contributing to its proliferation, aggressiveness, and malignant transformation. However, the discovery of novel oncogenes and the study of its molecular regulating mechanism based on circular RNAs (circRNAs) may provide a promising treatment target in glioma. Methods Bioinformatics analysis, qPCR, western blotting, and immunohistochemistry were used to detect the expression levels of ISL2, miR-342–3p, circRNA ARF1 (cARF1), U2AF2, and VEGFA. Patient-derived glioma stem cells (GSCs) were established for the molecular experiments. Lentiviral-based infection was used to regulate the expression of these molecules in GSCs. The MTS, EDU, Transwell, and tube formation assays were used to detect the proliferation, invasion, and angiogenesis of human brain microvessel endothelial cells (hBMECs). RNA-binding protein immunoprecipitation, RNA pull-down, dual-luciferase reporter, and chromatin immunoprecipitation assays were used to detect the direct regulation mechanisms among these molecules. Results We first identified a novel transcription factor related to neural development. ISL2 was overexpressed in glioma and correlated with poor patient survival. ISL2 transcriptionally regulated VEGFA expression in GSCs and promoted the proliferation, invasion, and angiogenesis of hBMECs via VEGFA-mediated ERK signaling. Regarding its mechanism of action, cARF1 upregulated ISL2 expression in GSCs via miR-342–3p sponging. Furthermore, U2AF2 bound to and promoted the stability and expression of cARF1, while ISL2 induced the expression of U2AF2, which formed a feedback loop in GSCs. We also showed that both U2AF2 and cARF1 had an oncogenic effect, were overexpressed in glioma, and correlated with poor patient survival. Conclusions Our study identified a novel feedback loop among U2AF2, cARF1, miR-342–3p, and ISL2 in GSCs. This feedback loop promoted glioma angiogenesis, and could provide an effective biomarker for glioma diagnosis and prognostic evaluation, as well as possibly being used for targeted therapy.

Connexin 43 Reverses Malignant Phenotypes of Glioma Stem Cells by Modulating E-Cadherin

Stem Cells ◽  
2012 ◽  
Vol 30 (2) ◽  
pp. 108-120 ◽  
Author(s):  
Shi-Cang Yu ◽  
Hua-Liang Xiao ◽  
Xue-Feng Jiang ◽  
Qing-Liang Wang ◽  
Yan Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Connexin 43 ◽  
Glioma Stem Cells ◽  
E Cadherin

scholarly journals miR-92a-3p Promoted EMT via Targeting LATS1 in Cervical Cancer Stem Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Shuangyue Liu ◽  
Liping Chu ◽  
Mingzhu Xie ◽  
Lisha Ma ◽  
Hongmei An ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cervical Cancer ◽  
Stem Cells ◽  
Western Blotting ◽  
Cyclin E ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Gene Assay ◽  
Cell Cycle Transition ◽  
E Cadherin

miR-92a-3p (microRNA-92a-3p) has been reported to be dysregulated in several cancers, and as such, it is considered to be a cancer-related microRNA. However, the influence of miR-92a-3p on biological behaviors in cervical cancer (CC) still remains unclear. Quantitative real-time PCR was used to detect miR-92a-3p levels in CC stem cells. Here, Cell Counting Kit-8 (CCK8) assay, Transwell cell invasion assay and flow cytometry assay were used to characterize the effects that miR-92a-3p and large tumor suppressor l (LATS1) had on proliferation, invasion and cell cycle transition. The luciferase reporter gene assay was used to verify the targeting relationship between miR-92a-3p and LATS1. Western Blotting was used to investigate the related signaling pathways and proteins. Data from The Cancer Genome Atlas (TCGA) showed that miR-92a-3p was upregulated in CC tissues and closely associated with overall survival. miR-92a-3p promoted proliferation, invasion and cell cycle transition in CC stem cells. The luciferase reporter assay showed that miR-92a-3p bound to the 3′-untranslated region (3′-UTR) of the LATS1 promoter. LATS1 inhibited proliferation, invasion and cell cycle transition. Results measured by Western Blotting showed that LATS1 downregulated expressions of transcriptional co-activator with PDZ-binding motif (TAZ), vimentin and cyclin E, but upregulated the expression of E-cadherin. Re-expression of LATS1 partly reversed the effects of miR-92a-3p on proliferation, invasion and cell cycle transition, as well as on TAZ, E-cadherin, vimentin, and cyclin E. miR-92a-3p promoted the malignant behavior of CC stem cells by targeting LATS1, which regulated TAZ and E-cadherin.

scholarly journals A comparative cell biological analysis reveals only limited functional homology between the NS5A proteins of hepatitis C virus and GB virus B

2008 ◽  
Vol 89 (8) ◽  
pp. 1911-1920 ◽  
Author(s):  
Jamel Mankouri ◽  
Andrew Milward ◽  
Kenneth R. Pryde ◽  
Lucile Warter ◽  
Annette Martin ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Cell Signalling ◽  
Pi3k Pathway ◽  
Luciferase Reporter ◽  
Signalling Pathways ◽  
New World Monkeys ◽  
Structural Protein ◽  
Reporter System ◽  
Early Endosomes

GB virus B (GBV-B) is the closest relative to hepatitis C virus (HCV) with which it shares a common genome organization, however, unlike HCV in humans, it generally causes an acute resolving hepatitis in New World monkeys. It is important to understand the factors regulating the different disease profiles of the two viruses and in this regard, as well as playing a key role in viral RNA replication, the HCV NS5A non-structural protein modulates a variety of host-cell signalling pathways. We have shown previously that HCV NS5A, expressed either alone, or in the context of the complete polyprotein, inhibits the Ras-extracellular-signal-regulated kinase (Erk) pathway and activates the phosphoinositide 3-kinase (PI3K) pathway. In this report, we investigate whether these functions are shared by GBV-B NS5A. Immunofluorescence analysis revealed that a C-terminally FLAG-tagged GBV-B NS5A exhibited a punctate cytoplasmic distribution. However, unlike HCV NS5A, the GBV-B protein did not partially co-localize with early endosomes. Utilizing a transient luciferase reporter system, we observed that GBV-B NS5A failed to inhibit Ras–Erk signalling, however GBV-B NS5A expression did result in the elevation of β-catenin-dependent transcription via activation of the PI3K pathway. These effects of GBV-B and HCV NS5A on the PI3K and Ras–Erk pathways were confirmed in cells harbouring subgenomic replicons derived from the two viruses. Based on these data we speculate that the differential effects of the two NS5A proteins on cellular signalling pathways may contribute to the differences in the natural history of the two viruses.

scholarly journals RETRACTED ARTICLE: MicroRNA-27a-3p Modulates the Wnt/β-Catenin Signaling Pathway to Promote Epithelial-Mesenchymal Transition in Oral Squamous Carcinoma Stem Cells by Targeting SFRP1

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Bin Qiao ◽  
Bao-Xia He ◽  
Jing-Hua Cai ◽  
Qian Tao ◽  
Alfred King-yin Lam
Keyword(s):  
Stem Cells ◽  
Signaling Pathway ◽  
Epithelial Mesenchymal Transition ◽  
Squamous Carcinoma ◽  
Luciferase Reporter ◽  
Mesenchymal Transition ◽  
Oral Squamous Carcinoma ◽  
Related Proteins ◽  
Emt Markers ◽  
E Cadherin

AbstractThis study aimed to elucidate how microRNA27a-3p (miR-27a-3p) modulates the Wnt/β-catenin signaling pathway to promote the epithelial-mesenchymal transition (EMT) in oral squamous carcinoma stem cells (OSCSCs) by targeting secreted frizzled-related protein 1 (SFRP1). Flow cytometry was used to sort OSCSCs from the SCC-9 and Tca8113 cell lines. The OSCSCs were randomly assigned into the miR-27a-3p inhibitors group, the miR-27a-3p inhibitors-NC group, the si-SFRP1 group, the si-SFRP1 + miR-27a-3p inhibitors group and the blank group. A luciferase reporter, immunofluorescence and Transwell assays were performed to detect luciferase activity, SFRP1, and cell migration and invasion, respectively. The mRNA expression of miR-27a-3p, SFRP1 and EMT markers (E-cadherin, N-cadherin, vimentin and ZEB1) were detected using qRT-PCR. The protein expression of SFRP1, EMT markers and the proteins of the Wnt/β-catenin signaling pathway was detected by Western blotting. OSCSCs showed up-regulated miR-27a-3p, Wnt/β-catenin signaling pathway-related proteins, vimentin, N-cadherin and ZEB1 and down-regulated SFRP1 and E-cadherin. MiR-27a-3p targeted SFRP1. Down-regulated miR-27a-3p resulted in increased E-cadherin and SFRP1 but decreased vimentin, N-cadherin, ZEB1, the Wnt/β-catenin signaling pathway-related proteins, and invasive and migratory cells. Silenced SFRP1 reversed this effect. We found that miR-27a-3p modulated the Wnt/β-catenin signaling pathway to promote EMT in OSCSCs by down-regulating SFRP1.

scholarly journals The SRSF1/circATP5B/miR-185-5p/HOXB5 Feedback Loop Regulates the Proliferation of Glioma Stem Cells via the IL6-mediated JAK2/STAT3 Signaling Pathway

2020 ◽  
Author(s):  
Junshuang Zhao ◽  
Yang Jiang ◽  
Haiying Zhang ◽  
Jinpeng Zhou ◽  
Lian Chen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Feedback Loop ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Bioinformatic Analysis ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Glioma Stem Cells ◽  
Stat3 Signaling

Abstract BackgroundGlioma is the most common and malignant tumor of central nervous system. The tumor initiation, self-renewal, and multi-lineage differentiation abilities of glioma stem cells (GSCs) are responsible for glioma proliferation and recurrence. Although circular RNAs (circRNAs) play vital roles in the progression of glioma, the detailed mechanisms remain unknown. MethodsqRT-PCR, western blotting, immunohistochemistry, and bioinformatic analysis were performed to detect the expression of circATP5B, miR-185-5p, HOXB5, and SRSF1. Patient-derived GSCs were established, and MTS, EDU, neurosphere formation, and limiting dilution assays were used to detect the proliferation of GSCs. RNA-binding protein immunoprecipitation, RNA pull-down, luciferase reporter assays, and chromatin immunoprecipitation assays were used to detect these molecules' regulation mechanisms. ResultsWe found circATP5B expression was significantly upregulated in GSCs and promoted the proliferation of GSCs. Mechanistically, circATP5B acted as miR-185-5p sponge to upregulate HOXB5 expression. HOXB5 was overexpressed in glioma and transcriptionally regulated IL6 expression and promoted the proliferation of GSCs via JAK2/STAT3 signaling. Moreover, RNA binding protein SRSF1 could bind to and promote circATP5B expression and regulate the proliferation of GSCs, while HOXB5 also transcriptionally regulated SRSF1 expression. ConclusionsOur study identified the SRSF1/circATP5B/miR-185-5p/HOXB5 feedback loop in GSCs. This provides an effective biomarker for glioma diagnosis and prognostic evaluation.

scholarly journals Cped1 promotes chicken SSCs formation with the aid of histone acetylation and transcription factor Sox2

2018 ◽  
Vol 38 (5) ◽  
Author(s):  
Chen Zhang ◽  
Fei Wang ◽  
Qisheng Zuo ◽  
Changhua Sun ◽  
Jing Jin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Transcription Factor ◽  
Histone Acetylation ◽  
Histone Deacetylase Inhibitors ◽  
Embryonic Stem ◽  
Control Area ◽  
Luciferase Reporter ◽  
Marker Genes

Spermatogonial stem cells (SSCs) may apply to gene therapy, regenerative medicine in place of embryonic stem cells (ESCs). However, the application of SSCs was severely limited by the low induction efficiency and the lack of thorough analysis of the regulatory mechanisms of SSCs formation. Current evidences have demonstrated multiple marker genes of germ cells, while genes that specifically regulate the formation of SSCs have not been explored. In our study, cadherin-like and PC-esterase domain containing 1 (Cped1) expressed specifically in SSCs based on RNA-seq data analysis. To study the function of Cped1 in the formation of SSCs, we successfully established a CRISPR/Cas9 knockout system. The gene disruption frequency is 37% in DF1 and 25% in ESCs without off-target effects. Knockout of Cped1 could significantly inhibit the formation of SSCs in vivo and in vitro. The fragment of −1050 to −1 bp had the activity as Cped1 gene promoter. Histone acetylation could regulate the expression of Cped1. We added 5-azaeytidi (DNA methylation inhibitors) and TSA (histone deacetylase inhibitors) respectively during the cultivation of SSCs. TSA was validated to promote the transcription of Cped1. Dual-luciferase reporter assay revealed that active control area of the chicken Cped1 gene is −296 to −1 bp. There are Cebpb, Sp1, and Sox2 transcription factor binding sites in this region. Point-mutation experiment results showed that Sox2 negatively regulates the transcription of Cped1. Above results demonstrated that Cped1 is a key gene that regulates the formation of SSCs. Histone acetylation and transcription factor Sox2 participate in the regulation of Cped1.

scholarly journals The SRSF1/circATP5B/miR-185-5p/HOXB5 feedback loop regulates the proliferation of glioma stem cells via the IL6-mediated JAK2/STAT3 signaling pathway

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Junshuang Zhao ◽  
Yang Jiang ◽  
Haiying Zhang ◽  
Jinpeng Zhou ◽  
Lian Chen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Feedback Loop ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Bioinformatic Analysis ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Glioma Stem Cells ◽  
Stat3 Signaling

Abstract Background Glioma is the most common and malignant tumor of central nervous system. The tumor initiation, self-renewal, and multi-lineage differentiation abilities of glioma stem cells (GSCs) are responsible for glioma proliferation and recurrence. Although circular RNAs (circRNAs) play vital roles in the progression of glioma, the detailed mechanisms remain unknown. Methods qRT-PCR, western blotting, immunohistochemistry, and bioinformatic analysis were performed to detect the expression of circATP5B, miR-185-5p, HOXB5, and SRSF1. Patient-derived GSCs were established, and MTS, EDU, neurosphere formation, and limiting dilution assays were used to detect the proliferation of GSCs. RNA-binding protein immunoprecipitation, RNA pull-down, luciferase reporter assays, and chromatin immunoprecipitation assays were used to detect these molecules’ regulation mechanisms. Results We found circATP5B expression was significantly upregulated in GSCs and promoted the proliferation of GSCs. Mechanistically, circATP5B acted as miR-185-5p sponge to upregulate HOXB5 expression. HOXB5 was overexpressed in glioma and transcriptionally regulated IL6 expression and promoted the proliferation of GSCs via JAK2/STAT3 signaling. Moreover, RNA binding protein SRSF1 could bind to and promote circATP5B expression and regulate the proliferation of GSCs, while HOXB5 also transcriptionally regulated SRSF1 expression. Conclusions Our study identified the SRSF1/circATP5B/miR-185-5p/HOXB5 feedback loop in GSCs. This provides an effective biomarker for glioma diagnosis and prognostic evaluation.

scholarly journals Valproic Acid Improves Porcine Parthenogenetic Embryo Development Through Transient Remodeling of Histone Modifiers

2015 ◽  
Vol 37 (4) ◽  
pp. 1463-1473 ◽  
Author(s):  
Yongye Huang ◽  
Lin Yuan ◽  
Tianye Li ◽  
Anfeng Wang ◽  
Zhanjun Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Valproic Acid ◽  
Histone Deacetylase Inhibitors ◽  
Formation Rate ◽  
Embryonic Stem ◽  
Cell Stage ◽  
Alternative Source ◽  
Histone Modifiers ◽  
Parthenogenetic Embryos

Background/Aims: Parthenogenetic embryos are useful in many applications, such as being an alternative source of embryonic stem cells that would avoid ethical problems. Aberrance in epigenetic reprogramming is considered the major reason for the developmental failure of parthenogenetic embryos. Many histone deacetylase inhibitors have been shown to improve the reprogramming of stem cells and embryos. Here, the relationship between histone modification and parthenogenetic embryonic development was explored. Methods: Valproic acid (VPA) treatment was applied during the culture of parthenogenetic embryos. The abundance of histone modifiers was examined by immunofluorescence and quantified by Image-pro software. Results: The acH3K9 level in in vitro fertilized embryos was significantly higher than parthenogenetic embryos. VPA treatment improved both the blastocyst formation rate and the acH3K9 level in parthenogenetic embryos. The signal intensities of acH4K5 and H3K4me2 were also enhanced in VPA treated embryos. The H3K27me2 level was decreased in the VPA treated embryos at the 2-cell stage. However, the enhancement in the acH3K9, acH4K5 and H3K4me2 level, or the decrease in the H3K27me2 level disappeared shortly after VPA withdrawal. Conclusion: Optimizing histone modifications for a short time following activation was sufficient to enhance the in vitro development of parthenogenetic embryos.

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