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Sample records for renal epithelial-mesenchymal transition

  1. AM251 Suppresses Epithelial-Mesenchymal Transition of Renal Tubular Epithelial Cells

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    Yoshinaga, Tomoyo; Uwabe, Kenichiro; Naito, Shoichi; Higashino, Kenichi; Nakano, Toru; Numata, Yoshito

    2016-01-01

    Epithelial-mesenchymal transition (EMT) of renal tubular epithelial cells is one of the causative mechanisms of kidney fibrosis. In our study, we screened lipophilic compounds using a lipid library including approximately 200 lipids to identify those that suppressed EMT induced by a transforming growth factor (TGF)-β1 stimulus. Initial screening was performed with the immortalized HK-2 renal tubule epithelial cell line. The most promising compounds were further tested in RPTEC primary renal tubule epithelial cells. We found that the synthetic lipid AM251 suppressed two hallmark events associated with EMT, the upregulation of collagen 1A1 (COL1A1) and downregulation of E-cadherin. Though AM251 is known to act as an antagonist for the cannabinoid receptor type 1 (CB1) and an agonist for the G protein-coupled receptor 55 (GRP55), the suppression of EMT by AM251 was not mediated through either receptor. Microarray analyses revealed that AM251 inhibited induction of several EMT transcription factors such as SNAIL1, which is the key inducer of EMT, and the AP-1 transcription factors FOSB and JUNB. Activation of SMAD2/3 and p38 mitogen-activated protein kinase (MAPK) was inhibited by AM251, with greater inhibition of the latter, indicating that AM251 acted upstream of SMAD/p38 MAPK in the TGF-β signaling pathway. Our findings regarding the effects of AM251 on the TGF-β signaling pathway may inform development of a novel therapeutic agent suppressing EMT, thus preventing kidney fibrosis. PMID:27936102

  2. AM251 Suppresses Epithelial-Mesenchymal Transition of Renal Tubular Epithelial Cells.

    Science.gov (United States)

    Yoshinaga, Tomoyo; Uwabe, Kenichiro; Naito, Shoichi; Higashino, Kenichi; Nakano, Toru; Numata, Yoshito; Kihara, Akio

    2016-01-01

    Epithelial-mesenchymal transition (EMT) of renal tubular epithelial cells is one of the causative mechanisms of kidney fibrosis. In our study, we screened lipophilic compounds using a lipid library including approximately 200 lipids to identify those that suppressed EMT induced by a transforming growth factor (TGF)-β1 stimulus. Initial screening was performed with the immortalized HK-2 renal tubule epithelial cell line. The most promising compounds were further tested in RPTEC primary renal tubule epithelial cells. We found that the synthetic lipid AM251 suppressed two hallmark events associated with EMT, the upregulation of collagen 1A1 (COL1A1) and downregulation of E-cadherin. Though AM251 is known to act as an antagonist for the cannabinoid receptor type 1 (CB1) and an agonist for the G protein-coupled receptor 55 (GRP55), the suppression of EMT by AM251 was not mediated through either receptor. Microarray analyses revealed that AM251 inhibited induction of several EMT transcription factors such as SNAIL1, which is the key inducer of EMT, and the AP-1 transcription factors FOSB and JUNB. Activation of SMAD2/3 and p38 mitogen-activated protein kinase (MAPK) was inhibited by AM251, with greater inhibition of the latter, indicating that AM251 acted upstream of SMAD/p38 MAPK in the TGF-β signaling pathway. Our findings regarding the effects of AM251 on the TGF-β signaling pathway may inform development of a novel therapeutic agent suppressing EMT, thus preventing kidney fibrosis.

  3. Tacrolimus Modulates TGF-β Signaling to Induce Epithelial-Mesenchymal Transition in Human Renal Proximal Tubule Epithelial Cells

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    Jason Bennett

    2016-04-01

    Full Text Available Epithelial-mesenchymal transition (EMT, a process which describes the trans-differentiation of epithelial cells into motile mesenchymal cells, is pivotal in stem cell behavior, development and wound healing, as well as contributing to disease processes including fibrosis and cancer progression. Maintenance immunosuppression with calcineurin inhibitors (CNIs has become routine management for renal transplant patient, but unfortunately the nephrotoxicity of these drugs has been well documented. HK-2 cells were exposed to Tacrolimus (FK506 and EMT markers were assessed by RT PCR and western blot. FK506 effects on TGF-β mRNA were assessed by RT PCR and TGF-β secretion was measured by ELISA. The impact of increased TGF-β secretion on Smad signaling pathways was investigated. The impact of inhibition of TGF-β signaling on EMT processes was assessed by scratch-wound assay. The results presented in this study suggest that FK506 initiates EMT processes in the HK-2 cell line, with altered expression of epithelial and myofibroblast markers evident. Additionally, the study demonstrates that FK506 activation of the TGF-β/ SMAD pathways is an essential step in the EMT process. Overall the results demonstrate that EMT is heavily involved in renal fibrosis associated with CNI nephrotoxicity.

  4. GSTA3 Attenuates Renal Interstitial Fibrosis by Inhibiting TGF-Beta-Induced Tubular Epithelial-Mesenchymal Transition and Fibronectin Expression.

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    Xiao, Yun; Liu, Jishi; Peng, Yu; Xiong, Xuan; Huang, Ling; Yang, Huixiang; Zhang, Jian; Tao, Lijian

    2016-01-01

    Tubular epithelial-mesenchymal transition (EMT) has been widely accepted as the underlying mechanisms of renal interstitial fibrosis (RIF). The production of reactive oxygen species (ROS) plays a vital role in tubular EMT process. The purpose of this study was to investigate the involved molecular mechanisms in TGF-beta-induced EMT and identify the potential role of glutathione S-transferase alpha 3 (GSTA3) in this process. The iTRAQ screening was performed to identify protein alterations of the rats underwent unilateral-ureteral obstruction (UUO). Protein expression of GSTA3 in patients with obstructive nephropathy and UUO rats was detected by immunohistochemistry. Protein and mRNA expression of GSTA3 in UUO rats and NRK-52E cells were determined by Western blot and RT-PCR. siRNA and overexpression plasmid were transfected specifically to assess the role of GSTA3 in RIF. The generation of ROS was measured by dichlorofluorescein fluorescence analysis. GSTA3 protein and mRNA expression was significantly reduced in UUO rats. Immunohistochemical analysis revealed that GSTA3 expression was reduced in renal cortex in UUO rats and patients with obstructive nephropathy. Treating with TGF-β1 down-regulated GSTA3 expression in NRK-52E cells, which have been found to be correlated with the decreased expression in E-cadherin and megalin and increased expression in α-smooth muscle actin. Furthermore, knocking down GSTA3 in NRK-52 cells led to increased production of ROS and tubular EMT, whereas overexpressing GSTA3 ameliorated ROS production and prevented the occurrence of tubular EMT. GSTA3 plays a protective role against tubular EMT in renal fibrosis, suggesting GSTA3 is a potential therapeutic target for RIF.

  5. A new mechanism of action of sulodexide in diabetic nephropathy: inhibits heparanase-1 and prevents FGF-2-induced renal epithelial-mesenchymal transition

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    Masola Valentina

    2012-10-01

    Full Text Available Abstract Background Epithelial-mesenchymal transition of tubular cells is a widely recognized mechanism that sustains interstitial fibrosis in diabetic nephropathy (DN. The signaling of FGF-2, a growth factor involved in this mechanism, is regulated by glycosaminoglycans. Heparanase-1, an endoglycosidase that cleaves heparan sulfate, is implicated in the pathogenesis of diabetic nephropathy and is necessary to FGF-2 for the induction of tubular cells transition. Well known Heparanase-1 inhibitors are heparin(s and sulodexide, a low-molecular weight heparin – dermatan sulphate blend, which is effective in the treatment of DN. Methods We have investigated the inhibition by sulodexide and its components of Heparanase-1 by an ELISA assay. We have analyzed its effect on the epithelial-mesenchymal transition of tubular cells by real time gene expression analysis, zymography and migration assay. Results Results show that sulodexide is an effective heparanase-1 inhibitor, exclusively in virtue to the heparin component, with an IC50 of 5 μg/ml. In FGF-2 treated tubular cells, sulodexide also prevents the over-expression of the mesenchymal markers αSMA, vimentin and fibronectin and the motility increase, i.e. the epithelial-mesenchymal transition of tubular cells. Moreover, sulodexide prevents FGF-2 induced heparanase-1 and MMP9 increase switching off the autocrine loop that FGF-2 activates to support its signal. Conclusions The findings highlight the capacity of sulodexide to inhibit heparanase-1 and to control tubular fibrosis triggered by epithelial-mesenchymal transition. In conclusion, these sulodexide activities support the value of this agent in controlling the progression of nephropathy to renal failure.

  6. Dioscorea alata attenuates renal interstitial cellular fibrosis by regulating Smad- and epithelial-mesenchymal transition signaling pathways.

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    Shu-Fen Liu

    Full Text Available Renal interstitial fibrosis is characterized by increased extracellular matrix (ECM synthesis. Epithelial-mesenchymal transition (EMT in kidneys is driven by regulated expression of fibrogenic cytokines such as transforming growth factor-beta (TGF-β. Yam, or Dioscorea alata (DA is an important herb in Chinese medicine widely used for the treatment of clinical diabetes mellitus. However, the fibrosis regulatory effect of DA is unclear. Thus, we examined TGF-β signaling mechanisms against EMT in rat fibroblast cells (NRK-49F. The characterization of DA water-extracts used various methods; after inducing cellular fibrosis in NRK-49F cells by treatment with β-hydroxybutyrate (β-HB (10 mM, we used Western blotting to examine the protein expression in the TGF-β-related signal protein type I and type II TGF-β receptors, Smads2 and Smad3 (Smad2/3, pSmad2 and Smad3 (pSmad2/3, Smads4, Smads7, and EMT markers. These markers included E-cadherin, alpha-smooth muscle actin (α-SMA, and matrix metalloproteinase-2 (MMP-2. Bioactive TGF-β and fibronectin levels in the culture media were determined using ELISA. Expressions of fibronectin and Snail transcription factor, an EMT-regulatory transcription factor, were assessed by immunofluorescence staining. DA extract dose-dependently (50-200 µg/mL suppressed β-HB-induced expression of fibronectin in NRK-49F cells concomitantly with the inhibition of Smad2/3, pSmad2/3, and Smad4. By contrast, Smad7 expression was significantly increased. DA extract caused a decrease in α-SMA (α-smooth muscle actin and MMP-2 levels, and an increase in E-cadherin expression. We propose that DA extract might act as a novel fibrosis antagonist, which acts partly by down regulating the TGF-β/smad signaling pathway and modulating EMT expression.

  7. Sorafenib ameliorates renal fibrosis through inhibition of TGF-β-induced epithelial-mesenchymal transition.

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    Lining Jia

    Full Text Available This study was to investigate whether sorafenib can inhibit the progression of renal fibrosis and to study the possible mechanisms of this effect.Eight-week-old rats were subjected to unilateral ureteral obstruction (UUO and were intragastrically administered sorafenib, while control and sham groups were administered vehicle for 14 or 21 days. NRK-52E cells were treated with TGF-β1 and sorafenib for 24 or 48 hours. HE and Masson staining were used to visualize fibrosis of the renal tissue in each group. The expression of α-SMA and E-cadherin in kidney tissue and NRK-52E cells were performed using immunohistochemistry and immunofluorescence. The apoptosis rate of NRK-52E cells was determined by flow cytometry analysis. The protein levels of Smad3 and p-Smad3 in kidney tissue and NRK-52E cells were detected by western blot analysis.HE staining demonstrated that kidney interstitial fibrosis, tubular atrophy, and inflammatory cell infiltration in the sorafenib-treated-UUO groups were significantly decreased compared with the vehicle-treated-UUO group (p<0.05. Masson staining showed that the area of fibrosis was significantly decreased in the sorafenib-treated-UUO groups compared with vehicle-treated-UUO group (p<0.01. The size of the kidney did not significantly increase; the cortex of the kidney was thicker and had a richer blood supply in the middle-dose sorafenib group compared with the vehicle-treated-UUO group (p<0.05. Compared with the vehicle-treated-UUO and TGF-β-stimulated NRK-52E groups, the expression of a-SMA and E-cadherin decreased and increased, respectively, in the UUO kidneys and NRK-52E cells of the sorafenib-treated groups (p<0.05. The apoptotic rate of NRK-52E cells treated with sorafenib decreased for 24 hours in a dose-dependent manner (p<0.05. Compared with the vehicle-treated UUO and TGF-β-stimulated NRK-52E groups, the ratio of p-Smad3 to Smad3 decreased in the sorafenib-treated groups (p<0.05.Our results suggest that

  8. Hepatitis B virus X protein promotes renal epithelial-mesenchymal transition in human renal proximal tubule epithelial cells through the activation of NF-κB.

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    Li, Mei; Hu, Liping; Zhu, Fengxin; Zhou, Zhangmei; Tian, Jianwei; Ai, Jun

    2016-08-01

    Hepatitis B virus (HBV)-associated glomerulo-nephritis is the most common extra-hepatic disorder occurring with hepatitis B virus infection. In the present study, we hypothesized that HBV X protein (HBx) may play a critical role in renal interstitial fibrosis, as HBx has been shown to induce epithelial-mesenchymal transition (EMT) in renal cells. For this purpose, we successfully transfected HBx plasmid into human renal proximal tubule epithelial cells (HK-2 cells). We found that transfection with HBx plasmid significantly downregulated E-cadherin expression and upregulated α-smooth muscle actin, collagen I and fibronectin expression in a time- and concentration-dependent manner (at the lower concentrations and earlier time points). HBx also increased nuclear factor-κB (NF-κB) phosphorylation in a time- and concentration-dependent manner (again at the lower concentrations and earlier time points); however, it did not alter the phosphorylation of Smad2, Smad3, p38, phosphoinositide 3-kinase (PI3K) or extracellular signal-regulated kinase (ERK). Thus, the findings of this study demonstrate that HBx promotes EMT in renal HK-2 cells, and the potential underlying mechanisms may involve the activation of the NF-κB signaling pathway.

  9. Epithelial-mesenchymal transition in malignant mesothelioma.

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    Fassina, Ambrogio; Cappellesso, Rocco; Guzzardo, Vincenza; Dalla Via, Lisa; Piccolo, Stefano; Ventura, Laura; Fassan, Matteo

    2012-01-01

    Epithelial-mesenchymal transition is a physiopathological process by which epithelial cells acquire mesenchymal shape and properties. Malignant mesothelioma is histologically characterized by the concomitant presence of epithelioid and sarcomatoid features, the latter being associated to worse prognosis, thus suggesting a role of epithelial-mesenchymal transition in this dual phenotype. We studied 109 malignant mesotheliomas (58 epithelioid, 26 sarcomatoid, and 25 biphasic) by immunohistochemistry and qRT-PCR analysis, and demonstrated a substantial switch from epithelial markers (E-cadherin, β-catenin, and cytokeratins 5/6) to mesenchymal markers (N-cadherin, vimentin, α-smooth muscle actin, Snail, Slug, Twist, ZEB1, ZEB2, S100A4, MMP2, and MMP9) through epithelioid to biphasic and sarcomatoid histotypes. In agreement with these findings, the ectopic expression of miR-205 (a repressor of ZEB1 and ZEB2 expression) in MeT-5A (mesothelial cell line), H2452 (an epithelioid malignant mesothelioma cell line) and MSTO-211H (a biphasic malignant mesothelioma cell line) not only induced a significant reduction of ZEB1 and ZEB2 and a consequent up-regulation of E-cadherin gene expression, but also inhibited migration and invasion. Moreover, miR-205 was significantly down-regulated in biphasic and sarcomatoid histotypes (qRT-PCR and in situ hybridization analyses). Collectively, our findings indicate that epithelial-mesenchymal transition has a significant part in the morphological features of malignant mesothelioma. In particular, miR-205 down-regulation correlated significantly with both a mesenchymal phenotype and a more aggressive behavior.

  10. Epithelial-Mesenchymal Transition in Pancreatic Carcinoma

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    Thomas Wirth

    2010-12-01

    Full Text Available Pancreatic carcinoma is the fourth-leading cause of cancer death and is characterized by early invasion and metastasis. The developmental program of epithelial-mesenchymal transition (EMT is of potential importance for this rapid tumor progression. During EMT, tumor cells lose their epithelial characteristics and gain properties of mesenchymal cells, such as enhanced motility and invasive features. This review will discuss recent findings pertinent to EMT in pancreatic carcinoma. Evidence for and molecular characteristics of EMT in pancreatic carcinoma will be outlined, as well as the connection of EMT to related topics, e.g., cancer stem cells and drug resistance.

  11. Epithelial-Mesenchymal Transition and Breast Cancer

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    Yanyuan Wu

    2016-01-01

    Full Text Available Breast cancer is the most common cancer in women and distant site metastasis is the main cause of death in breast cancer patients. There is increasing evidence supporting the role of epithelial-mesenchymal transition (EMT in tumor cell progression, invasion, and metastasis. During the process of EMT, epithelial cancer cells acquire molecular alternations that facilitate the loss of epithelial features and gain of mesenchymal phenotype. Such transformation promotes cancer cell migration and invasion. Moreover, emerging evidence suggests that EMT is associated with the increased enrichment of cancer stem-like cells (CSCs and these CSCs display mesenchymal characteristics that are resistant to chemotherapy and target therapy. However, the clinical relevance of EMT in human cancer is still under debate. This review will provide an overview of current evidence of EMT from studies using clinical human breast cancer tissues and its associated challenges.

  12. Renal tubular epithelial-mesenchymal transition in kidney fibrosis%肾小管上皮间充质转化与肾脏纤维化

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    王来亮; 罗群

    2014-01-01

    Epithelial-mesenchymal transition ( EMT) , a process by which differentiated epithelial cells under-go a phenotypic conversion that gives rise to the matrix-producing fibroblasts and myofibroblasts, is increasingly recognized as an integral part of tissue fibrogenesis after injury.However, the degree to which renal tubular epithelial EMT contributes to kidney fibrosis remains a matter of intense debate and is likely to be context-dependent.Renal tubular EMT is an adap-tive response of epithelial cells to a hostile or changing microenvironment and is regulated by many factors.Several intrace-llular signal transduction pathways such as transforming growth factor-β( TGF-β)/Smad and Wnt/β-catenin signaling are essential in controlling the process of renal tubular epithelial EMT which are potential targets of antifibrotic therapy present-ly.This review highlights the current understanding of renal tubular epithelial EMT and its underlying mechanisms to stimu-late further discussion on its role in the pathogenesis of renal interstitial fibrosis.

  13. Transcriptional networks in epithelial-mesenchymal transition.

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    Christo Venkov

    Full Text Available Epithelial-mesenchymal transition (EMT changes polarized epithelial cells into migratory phenotypes associated with loss of cell-cell adhesion molecules and cytoskeletal rearrangements. This form of plasticity is seen in mesodermal development, fibroblast formation, and cancer metastasis.Here we identify prominent transcriptional networks active during three time points of this transitional process, as epithelial cells become fibroblasts. DNA microarray in cultured epithelia undergoing EMT, validated in vivo, were used to detect various patterns of gene expression. In particular, the promoter sequences of differentially expressed genes and their transcription factors were analyzed to identify potential binding sites and partners. The four most frequent cis-regulatory elements (CREs in up-regulated genes were SRY, FTS-1, Evi-1, and GC-Box, and RNA inhibition of the four transcription factors, Atf2, Klf10, Sox11, and SP1, most frequently binding these CREs, establish their importance in the initiation and propagation of EMT. Oligonucleotides that block the most frequent CREs restrain EMT at early and intermediate stages through apoptosis of the cells.Our results identify new transcriptional interactions with high frequency CREs that modulate the stability of cellular plasticity, and may serve as targets for modulating these transitional states in fibroblasts.

  14. Epithelial-mesenchymal transition in ovarian carcinoma

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    Ben eDavidson

    2012-04-01

    Full Text Available Ovarian cancer is the most lethal gynecologic malignancy, with the majority of patients dying within 5 years of diagnosis. This poor survival of patients diagnosed with this malignancy is attributed to diagnosis at advanced stage, when the tumor has metastasized, and to chemotherapy resistance, either primary or developing along tumor progression. However, ovarian carcinomas, constituting the vast majority of ovarian cancers, additionally have unique biology, one aspect of which is the ability to co-express epithelial and mesenchymal determinants. Epithelial-mesenchymal transition (EMT, a physiological process by which mesenchymal cells are formed and migrate to target organs during embryogenesis, is involved in cancer cell invasion and metastasis. However, these changes do not fully occur in ovarian carcinoma, and are even reversed in tumor cells present in malignant peritoneal and pleural effusions. This review summarizes current knowledge in this area, including the characteristics of EMT related to adhesion, transcriptional regulation and chemoresistance, and their clinical relevance, as well as the recently observed regulation of EMT by microRNA.

  15. Parathyroid hormone-related protein promotes epithelial-mesenchymal transition.

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    Ardura, Juan Antonio; Rayego-Mateos, Sandra; Rámila, David; Ruiz-Ortega, Marta; Esbrit, Pedro

    2010-02-01

    Epithelial-mesenchymal transition (EMT) is an important process that contributes to renal fibrogenesis. TGF-beta1 and EGF stimulate EMT. Recent studies suggested that parathyroid hormone-related protein (PTHrP) promotes fibrogenesis in the damaged kidney, apparently dependent on its interaction with vascular endothelial growth factor (VEGF), but whether it also interacts with TGF-beta and EGF to modulate EMT is unknown. Here, PTHrP(1-36) increased TGF-beta1 in cultured tubuloepithelial cells and TGF-beta blockade inhibited PTHrP-induced EMT-related changes, including upregulation of alpha-smooth muscle actin and integrin-linked kinase, nuclear translocation of Snail, and downregulation of E-cadherin and zonula occludens-1. PTHrP(1-36) also induced EGF receptor (EGFR) activation; inhibition of protein kinase C and metalloproteases abrogated this activation. Inhibition of EGFR activation abolished these EMT-related changes, the activation of ERK1/2, and upregulation of TGF-beta1 and VEGF by PTHrP(1-36). Moreover, inhibition of ERK1/2 blocked EMT induced by either PTHrP(1-36), TGF-beta1, EGF, or VEGF. In vivo, obstruction of mouse kidneys led to changes consistent with EMT and upregulation of TGF-beta1 mRNA, p-EGFR protein, and PTHrP. Taken together, these data suggest that PTHrP, TGF-beta, EGF, and VEGF might cooperate through activation of ERK1/2 to induce EMT in renal tubuloepithelial cells.

  16. MET: roles in epithelial-mesenchymal transition and cancer stemness

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    Jeon, Hye-Min

    2017-01-01

    In a number of cancers, deregulated MET pathway leads to aberrantly activated proliferative and invasive signaling programs that promote malignant transformation, cell motility and migration, angiogenesis, survival in hypoxia, and invasion. A better understanding of oncogenic MET signaling will help us to discover effective therapeutic approaches and to identify which tumors are likely to respond to MET-targeted cancer therapy. In this review, we will summarize the roles of MET signaling in cancer, with particular focus on epithelial-mesenchymal transition (EMT) and cancer stemness. Then, we will provide update on MET targeting agents and discuss the challenges that should be overcome for the development of an effective therapy. PMID:28164090

  17. Epithelial-Mesenchymal Transition in tumor microenvironment

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    Jing Yingying

    2011-08-01

    Full Text Available Abstract The epithelial to mesenchymal transition (EMT plays crucial roles in the formation of the body plan and also in the tumor invasion process. In addition, EMT also causes disruption of cell-cell adherence, loss of apico-basal polarity, matrix remodeling, increased motility and invasiveness in promoting tumor metastasis. The tumor microenvironment plays an important role in facilitating cancer metastasis and may induce the occurrence of EMT in tumor cells. A large number of inflammatory cells infiltrating the tumor site, as well as hypoxia existing in a large area of tumor, in addition many stem cells present in tumor microenvironment, such as cancer stem cells (CSCs, mesenchymal stem cells (MSCs, all of these may be the inducers of EMT in tumor cells. The signaling pathways involved in EMT are various, including TGF-β, NF-κB, Wnt, Notch, and others. In this review, we discuss the current knowledge about the role of the tumor microenvironment in EMT and the related signaling pathways as well as the interaction between them.

  18. Epithelial-mesenchymal transition: Understanding the basic concept

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    Suresh Babu Ghanta

    2012-01-01

    Full Text Available The epithelial-mesenchymal transition (EMT is described as a rapid and reversible process of change of cell phenotype seen during embryonic development, organ fibrosis, and tumor progression. EMT was first described by Gary Greenberg and Elizabeth Hay in 1982. During EMT the epithelial cells alter their cell polarity, reorganize their cytoskeleton thus become isolated and motile. Depending upon the biological context in which they occur, EMT is divided into three types namely EMT type I, II, III. The article describes the process of EMT implicated in the oral cavity as in palate and root development (type I EMT, gingival fibromatosis and oral sub-mucous fibrosis (type II EMT, and oral squamous cell carcinoma (type III EMT. The reverse process of EMT is called as mesenchymal-epithelial transition seen in association with kidney formation.

  19. Epithelial-mesenchymal transition in breast cancer progression and metastasis

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    Yifan Wang; Binhua P. Zhou

    2011-01-01

    Breast cancer is the most common cancer in women,and approximately 90% of breast cancer deaths are caused by local invasion and distant metastasis of tumor cells.Epithelial-mesenchymal transition (EMT) is a vital process for large-scale cell movement during morphogenesis at the time of embryonic development.Tumor cells usurp this developmental program to execute the multi-step process of tumorigenesis and metastasis.Several transcription factors and signals are involved in these events.In this review,we summarize recent advances in breast cancer researches that have provided new insights in the molecular mechanisms underlying EMT regulation during breast cancer progression and metastasis.We especially focus on the molecular pathways that control EMT.

  20. TRIM37 promotes epithelial-mesenchymal transition in colorectal cancer

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    Hu, Cheng-En; Gan, Jun

    2017-01-01

    There is substantial research on the oncogenic role of tripartite motif containing 37 (TRIM37); however, its importance in colorectal cancer (CRC) remains to be elucidated. The present study used reverse transcription-quantitative polymerase chain reaction, immunohistochemistry and western blotting to detect the expression level of TRIM37 in CRC. The importance of TRIM37 in cell proliferation, invasion and metastasis of CRC were investigated through overexpressing or knocking-down of TRIM37 in CRC cell lines, to observe its function. The present study revealed that TRIM37 was overexpressed in human CRC tissues. High TRIM37 expression resulted in increased CRC proliferation, migration and invasion. Mechanistically, it was confirmed that TRIM37 enhanced invasion and metastasis of CRC via the epithelial-mesenchymal transition pathway. In conclusion, the present study suggested that TRIM3 may contribute to CRC and act as a potential therapeutic target for CRC treatment. PMID:28098873

  1. New insights of epithelial-mesenchymal transition in cancer metastasis

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    Yadi Wu; Binhua P.Zhou

    2008-01-01

    Epithelial-mesenchymal transition (EMT) is a key step during embryonic morphogenesis,heart development,chronic degenerative fibrosis,and cancer metastasis.Several distinct traits have been conveyed by EMT,including cell motility,invasiveness,resistance to apoptosis,and some properties of stem cells.Many signal pathways have contributed to the induction of EMT,such as transforming growth factor-β,Wnt,Hedgehog,Notch,and nuclear factor-κB.Over the last few years,increasing evidence has shown that EMT plays an essential role in tumor progression and metastasis.Understanding the molecular mechanism of EMT has a great effect in unraveling the metastatic cascade and may lead to novel interventions for metastatic disease.

  2. Epithelial-mesenchymal transition in prostate cancer: an overview.

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    Montanari, Micaela; Rossetti, Sabrina; Cavaliere, Carla; D'Aniello, Carmine; Malzone, Maria Gabriella; Vanacore, Daniela; Di Franco, Rossella; La Mantia, Elvira; Iovane, Gelsomina; Piscitelli, Raffaele; Muscariello, Raffaele; Berretta, Massimiliano; Perdonà, Sisto; Muto, Paolo; Botti, Gerardo; Bianchi, Attilio Antonio Montano; Veneziani, Bianca Maria; Facchini, Gaetano

    2017-05-23

    Prostate cancer is a main urological disease associated with significant morbidity and mortality. Radical prostatectomy and radiotherapy are potentially curative for localized prostate cancer, while androgen deprivation therapy is the initial systemic therapy for metastatic prostate disease. However, despite temporary response, most patients relapse and evolve into castration resistant cancer.Epithelial-mesenchymal transition (EMT) is a complex gradual process that occurs during embryonic development and/or tumor progression. During this process, cells lose their epithelial characteristics and acquire mesenchymal features. Increasing evidences indicate that EMT promotes prostate cancer metastatic progression and it is closely correlated with increased stemness and drug resistance.In this review, we discuss the main molecular events that directly or indirectly govern the EMT program in prostate cancer, in order to better define the role and the mechanisms underlying this process in prostate cancer progression and therapeutic resistance.

  3. Epithelial-mesenchymal transition mediated tumourigenesis in the gastrointestinal tract

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    Ammar Natalwala; Robert Spychal; Chris Tselepis

    2008-01-01

    Epithelial-mesenchymal transition (EMT) is a highly conserved process that has been well characterised in embryogenesis.Studies have shown that the aberrant activation of EMT in adult epithelia can promote tumour metastasis by repressing cell adhesion molecules,including epithelial (E)-cadherin.Reduced intracellular adhesion may allow tumour cells to disseminate and spread throughout the body.A number of transcription proteins of the Snail superfamily have been implicated in EMT.These proteins have been shown to be overexpressed in advanced gastrointestinal (GI) tumours including oesophageal adenocarcinomas,colorectal carcinomas,gastric and pancreatic cancers,with a concomitant reduction in the expression of E-cadherin.Regulators of EMT may provide novel clinical targets to detect GI cancers early,so that cancers previously associated with a poor prognosis such as pancreatic cancer can be diagnosed before they become inoperable.Furthermore,pharmacological therapies designed to inhibit these proteins will aim to prevent local and distant tumour invasion.

  4. Mycophenolate mofetil ameliorates diabetic nephropathy through epithelial mesenchymal transition in rats.

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    Xiao, Xiaoyan; Wang, Jie; Chang, Xiangdi; Zhen, Junhui; Zhou, Gengyin; Hu, Zhao

    2015-09-01

    Recent studies in animal models have revealed that mycophenolate mofetil (MMF) has certain protective effects against experimental diabetic nephropathy. The present study therefore aimed to investigate the hypothesis that diabetic nephropathy may be ameliorated by mycophenolate mofetil and benazepril treatment alone or in combination, and identify the potential underlying mechanisms in a rat model. Diabetes was induced in rats by a single intraperitoneal injection of streptozotocin. Rats were subsequently treated with benazepril, MMF or a combination of the two drugs, and blood glucose, normalized kidney weight, urine protein and serum creatinine were determined. The pathological changes in renal tissue were also observed. In addition, indices of epithelial mesenchymal transition, including α‑smooth muscle actin (α‑SMA) and transforming growth factor (TGF)‑β1 expression, were examined. Normalized kidney weight, urine protein and serum creatinine levels were significantly improved in the diabetic rats treated with benazepril or mycophenolate mofetil, compared with those of rats in the untreated diabetic group. Pathological changes in the kidney were detected concurrently with increasing kidney weight and urinary albumin excretion, with a similar trend in variation among groups. In addition, the expression of epithelial mesenchymal transition indices, including α‑SMA and TGF‑β1, in the renal tubule interstitium were significantly decreased in the benazepril‑ and MMF‑treated groups compared with those of the diabetic group. As expected, the aforementioned indices were markedly lower in the benazepril and MMF combined treatment group than those in the single medication groups. These data suggested that MMF may have a protective role in diabetic nephropathy, and that the underlying mechanism may be partially dependent upon the suppression of the epithelial mesenchymal transition. Furthermore, the combination of benazepril and MMF conferred enhanced

  5. Epithelial-mesenchymal transition: a new target in anticancer drug discovery.

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    Marcucci, Fabrizio; Stassi, Giorgio; De Maria, Ruggero

    2016-05-01

    The conversion of cells with an epithelial phenotype into cells with a mesenchymal phenotype, referred to as epithelial-mesenchymal transition, is a critical process for embryonic development that also occurs in adult life, particularly during tumour progression. Tumour cells undergoing epithelial-mesenchymal transition acquire the capacity to disarm the body's antitumour defences, resist apoptosis and anticancer drugs, disseminate throughout the organism, and act as a reservoir that replenishes and expands the tumour cell population. Epithelial-mesenchymal transition is therefore becoming a target of prime interest for anticancer therapy. Here, we discuss the screening and classification of compounds that affect epithelial-mesenchymal transition, highlight some compounds of particular interest, and address issues related to their clinical application.

  6. Effects of artificial cordyceps sinensis on epithelial-mesenchymal transition in the podocytes of diabetic rats

    Institute of Scientific and Technical Information of China (English)

    蔡芸莹

    2013-01-01

    Objective To assess the effects of artificial cordyceps sinensis(Jin shuibao) on the numbers of podocytes and epithelial-mesenchymal transition in diabetic rats. Methods Diabetes was induced by intraperitoneal injection of low dose streptozocin.

  7. The translational significance of epithelial-mesenchymal transition in head and neck cancer

    OpenAIRE

    2014-01-01

    Positive markers of epithelial-mesenchymal transition (EMT) in head and neck cancers complicate clinical management and are associated with reduced survival. We discuss recent translational discoveries in EMT and suggest additional actionable molecular pathways, biomarkers, and clinical agents.

  8. Epithelial-mesenchymal transition in tissue repair and fibrosis.

    Science.gov (United States)

    Stone, Rivka C; Pastar, Irena; Ojeh, Nkemcho; Chen, Vivien; Liu, Sophia; Garzon, Karen I; Tomic-Canic, Marjana

    2016-09-01

    The epithelial-mesenchymal transition (EMT) describes the global process by which stationary epithelial cells undergo phenotypic changes, including the loss of cell-cell adhesion and apical-basal polarity, and acquire mesenchymal characteristics that confer migratory capacity. EMT and its converse, MET (mesenchymal-epithelial transition), are integral stages of many physiologic processes and, as such, are tightly coordinated by a host of molecular regulators. Converging lines of evidence have identified EMT as a component of cutaneous wound healing, during which otherwise stationary keratinocytes (the resident skin epithelial cells) migrate across the wound bed to restore the epidermal barrier. Moreover, EMT plays a role in the development of scarring and fibrosis, as the matrix-producing myofibroblasts arise from cells of the epithelial lineage in response to injury but are pathologically sustained instead of undergoing MET or apoptosis. In this review, we summarize the role of EMT in physiologic repair and pathologic fibrosis of tissues and organs. We conclude that further investigation into the contribution of EMT to the faulty repair of fibrotic wounds might identify components of EMT signaling as common therapeutic targets for impaired healing in many tissues. Graphical Abstract Model for injury-triggered EMT activation in physiologic wound repair (left) and fibrotic wound healing (right).

  9. Regulators of epithelial mesenchymal transition in pancreatic cancer

    Directory of Open Access Journals (Sweden)

    Shin eHamada

    2012-07-01

    Full Text Available Pancreatic cancer is a leading cause of cancer related death due to its invasive nature. Despite the improvement of diagnostic strategy, early diagnosis of pancreatic cancer is still challenging. Surgical resection is the only curative therapy, while vast majority of patients are not eligible for this therapeutic option. Complex biological processes are involved in the establishment of invasion and metastasis of pancreatic cancer and epithelial-mesenchymal transition (EMT has been reported to play crucial role. EMT is part of the normal developmental processes which mobilizes epithelial cells and yields mesenchymal phenotype. Deregulation of EMT inducing molecules in pancreatic cancer is reported, such as multiple cytokines, growth factors and downstream transcriptional factors. In addition to these molecules, non-coding RNA including miRNA also contributes to EMT. EMT of cancer cell also correlates with cancer stem cell properties such as chemoresistance or tumorigenicity, therefore these upstream regulators of EMT could be attractive therapeutic targets and several candidates are examined for clinical application. This review summarizes recent advances in this field, focusing the regulatory molecules of EMT and their downstream targets. Further understanding and research advances will clarify the cryptic mechanism of cancer metastasis and delineate novel therapeutic targets.

  10. Tissue geometry patterns epithelial-mesenchymal transition via intercellular mechanotransduction

    Science.gov (United States)

    Gomez, Esther W.; Chen, Qike K.; Gjorevski, Nikolce; Nelson, Celeste M.

    2010-01-01

    Epithelial-mesenchymal transition (EMT) is a phenotypic change in which epithelial cells detach from their neighbors and become motile. Whereas soluble signals such as growth factors and cytokines are responsible for stimulating EMT, here we show that gradients of mechanical stress define the spatial locations at which EMT occurs. When treated with transforming growth factor (TGF)-β, cells at the corners and edges of square mammary epithelial sheets expressed EMT markers, whereas those in the center did not. Changing the shape of the epithelial sheet altered the spatial pattern of EMT. Traction force microscopy and finite element modeling demonstrated that EMT-permissive regions experienced the highest mechanical stress. Myocardin-related transcription factor (MRTF)-A was localized to the nuclei of cells located in high-stress regions, and inhibiting cytoskeletal tension or MRTF-A expression abrogated the spatial patterning of EMT. These data suggest a causal role for tissue geometry and endogenous mechanical stresses in the spatial patterning of EMT. PMID:20336666

  11. Osteopontin—A Master Regulator of Epithelial-Mesenchymal Transition

    Directory of Open Access Journals (Sweden)

    Anai N. Kothari

    2016-03-01

    Full Text Available Osteopontin (OPN plays an important functional role in both physiologic and pathologic states. OPN is implicated in the progression of fibrosis, cancer, and metastatic disease in several organ systems. The epithelial-mesenchymal transition (EMT, first described in embryology, is increasingly being recognized as a significant contributor to fibrotic phenotypes and tumor progression. Several well-established transcription factors regulate EMT and are conserved across tissue types and organ systems, including TWIST, zinc finger E-box-binding homeobox (ZEB, and SNAIL-family members. Recent literature points to an important relationship between OPN and EMT, implicating OPN as a key regulatory component of EMT programs. In this review, OPN’s interplay with traditional EMT activators, both directly and indirectly, will be discussed. Also, OPN’s ability to restructure the tissue and tumor microenvironment to indirectly modify EMT will be reviewed. Together, these diverse pathways demonstrate that OPN is able to modulate EMT and provide new targets for directing therapeutics.

  12. Targeting ROR1 inhibits epithelial-mesenchymal transition and metastasis.

    Science.gov (United States)

    Cui, Bing; Zhang, Suping; Chen, Liguang; Yu, Jianqiang; Widhopf, George F; Fecteau, Jessie-F; Rassenti, Laura Z; Kipps, Thomas J

    2013-06-15

    Metastasis is responsible for 90% of cancer-related deaths. Strategies are needed that can inhibit the capacity of cancer cells to migrate across the anatomic barriers and colonize distant organs. Here, we show an association between metastasis and expression of a type I receptor tyrosine kinase-like orphan receptor, ROR1, which is expressed during embryogenesis and by various cancers, but not by normal postpartum tissues. We found that expression of ROR1 associates with the epithelial-mesenchymal transition (EMT), which occurs during embryogenesis and cancer metastasis. Breast adenocarcinomas expressing high levels of ROR1 were more likely to have gene expression signatures associated with EMT and had higher rates of relapse and metastasis than breast adenocarcinomas expressing low levels of ROR1. Suppressing expression of ROR1 in metastasis-prone breast cancer cell lines, MDA-MB-231, HS-578T, or BT549, attenuated expression of proteins associated with EMT (e.g., vimentin, SNAIL-1/2, and ZEB1), enhanced expression of E-cadherin, epithelial cytokeratins (e.g., CK-19), and tight junction proteins (e.g., ZO-1), and impaired their migration/invasion capacity in vitro and the metastatic potential of MDA-MB-231 cells in immunodeficient mice. Conversely, transfection of MCF-7 cells to express ROR1 reduced expression of E-cadherin and CK-19, but enhanced the expression of SNAIL-1/2 and vimentin. Treatment of MDA-MB-231 with a monoclonal antibody specific for ROR1 induced downmodulation of vimentin and inhibited cancer cell migration and invasion in vitro and tumor metastasis in vivo. Collectively, this study indicates that ROR1 may regulate EMT and metastasis and that antibodies targeting ROR1 can inhibit cancer progression and metastasis.

  13. Type 2 epithelial mesenchymal transition in vivo: truth or pitfalls?

    Institute of Scientific and Technical Information of China (English)

    XU Xue-feng; DAI Hua-ping

    2012-01-01

    Epithelial-mesenchymal transition (EMT) is a process by which fully differentiated epithelial cells undergo a phenotypic conversion and assume a mesenchymal cell phenotype,including elongated morphology,enhanced migratory and invasiveness capacity,and greatly increased production of extracellular matrix (ECM) components.The EMTs associated with wound healing,tissue regeneration,and organ fibrosis are termed as type 2 EMT.Over the past two decades,emerging evidence suggested that injured epithelial cells,via type 2 EMT,may serve as important sources of fibroblasts and contribute to organ fibrosis,such as kidney,liver,lung and eyes.There is perhaps no doubt that adult epithelial cells can undergo EMT in vitro in response to transforming growth factor (TGF)-β1 and other inflammatory or pro-fibrotic stimuli.However,whether type 2 EMT really occurs in vivo,whethers it is actually a source of functional and activated interstitial fibroblasts and whether it contributes to tissue fibrosis have already been the subjects of heated debate.In this review,we will describe the main features of EMT,the major findings of type 2 EMT in vitro,the evidences for and against type 2 EMT in vivo and discuss the heterogeneity and pitfalls of the techniques used to detect EMT during fibrotic diseases.We suggest that in order to ascertain the existence of type 2 EMT in vivo,different proper phenotype markers of epithelial and mesenchymal cells should be jointly used and cell lineage tracking techniques should be standardized and avoid false positives.Finally,we believe that if EMT really occurs and contributes to tissue fibrosis,efforts should be made to block or reverse EMTto attenuate fibrotic process.

  14. Role of Epithelial-Mesenchyme Transition in Chlamydia Pathogenesis.

    Directory of Open Access Journals (Sweden)

    Joseph U Igietseme

    Full Text Available Chlamydia trachomatis genital infection in women causes serious adverse reproductive complications, and is a strong co-factor for human papilloma virus (HPV-associated cervical epithelial carcinoma. We tested the hypothesis that Chlamydia induces epithelial-mesenchyme transition (EMT involving T cell-derived TNF-alpha signaling, caspase activation, cleavage inactivation of dicer and dysregulation of micro-RNA (miRNA in the reproductive epithelium; the pathologic process of EMT causes fibrosis and fertility-related epithelial dysfunction, and also provides the co-factor function for HPV-related cervical epithelial carcinoma. Using a combination of microarrays, immunohistochemistry and proteomics, we showed that chlamydia altered the expression of crucial miRNAs that control EMT, fibrosis and tumorigenesis; specifically, miR-15a, miR-29b, miR-382 and MiR-429 that maintain epithelial integrity were down-regulated, while miR-9, mi-R-19a, miR-22 and miR-205 that promote EMT, fibrosis and tumorigenesis were up-regulated. Chlamydia induced EMT in vitro and in vivo, marked by the suppression of normal epithelial cell markers especially E-cadherin but up-regulation of mesenchymal markers of pathological EMT, including T-cadherin, MMP9, and fibronectin. Also, Chlamydia upregulated pro-EMT regulators, including the zinc finger E-box binding homeobox protein, ZEB1, Snail1/2, and thrombospondin1 (Thbs1, but down-regulated anti-EMT and fertility promoting proteins (i.e., the major gap junction protein connexin 43 (Cx43, Mets1, Add1Scarb1 and MARCKSL1. T cell-derived TNF-alpha signaling was required for chlamydial-induced infertility and caspase inhibitors prevented both infertility and EMT. Thus, chlamydial-induced T cell-derived TNF-alpha activated caspases that inactivated dicer, causing alteration in the expression of reproductive epithelial miRNAs and induction of EMT. EMT causes epithelial malfunction, fibrosis, infertility, and the enhancement of

  15. Involvement of O-glycosylation defining oncofetal fibronectin in epithelial-mesenchymal transition process

    DEFF Research Database (Denmark)

    Freire-de-Lima, Leonardo; Gelfenbeyn, Kirill; Ding, Yao

    2011-01-01

    The process termed "epithelial-mesenchymal transition" (EMT) was originally discovered in ontogenic development, and has been shown to be one of the key steps in tumor cell progression and metastasis. Recently, we showed that the expression of some glycosphingolipids (GSLs) is down-regulated duri...

  16. Roles and Regulation of Epithelial Splicing Regulatory Proteins 1 and 2 in Epithelial-Mesenchymal Transition

    NARCIS (Netherlands)

    Gottgens, E.L.; Span, P.N.; Zegers, M.M.P.

    2016-01-01

    The transformation of polarized epithelial cells into cells with mesenchymal characteristics by the morphogenetic process of epithelial-mesenchymal transition (EMT) is a well-characterized process essential for embryonic development and associated with cancer progression. EMT is a program driven by

  17. Epithelial-mesenchymal transition: molecular pathways of hepatitis viruses-induced hepatocellular carcinoma progression.

    Science.gov (United States)

    Panebianco, Concetta; Saracino, Chiara; Pazienza, Valerio

    2014-08-01

    Hepatocellular carcinoma is the fifth most common tumor and the third cause of death for cancer in the world. Among the main causative agents of this tumor is the chronic infection by hepatitis viruses B and C, which establish a context of chronic inflammation degenerating in fibrosis, cirrhosis, and, finally, cancer. Recent findings, however, indicate that hepatitis viruses are not only responsible for cancer onset but also for its progression towards metastasis. Indeed, they are able to promote epithelial-mesenchymal transition, a process of cellular reprogramming underlying tumor spread. In this manuscript, we review the currently known molecular mechanisms by which hepatitis viruses induce epithelial-mesenchymal transition and, thus, hepatocellular carcinoma progression.

  18. TEAD transcription factors mediate the function of TAZ in cell growth and epithelial-mesenchymal transition.

    Science.gov (United States)

    Zhang, Heng; Liu, Chen-Ying; Zha, Zheng-Yu; Zhao, Bin; Yao, Jun; Zhao, Shimin; Xiong, Yue; Lei, Qun-Ying; Guan, Kun-Liang

    2009-05-15

    The TAZ transcription co-activator has been shown to promote cell proliferation and to induce epithelial-mesenchymal transition. Recently we have demonstrated that TAZ is phosphorylated and inhibited by the Hippo tumor suppressor pathway, which is altered in human cancer. The mechanism of TAZ-mediated transcription is unclear. We demonstrate here that TEAD is a key downstream transcription factor mediating the function of TAZ. Disruption of TEAD-TAZ binding or silencing of TEAD expression blocked the function of TAZ to promote cell proliferation and to induce epithelial-mesenchymal transition, demonstrating TEAD as a key downstream effector of TAZ. We also identified CTGF, a gene that regulates cell adhesion, proliferation, and migration, as a direct target of TAZ and TEAD. Our study establishes a functional partnership between TAZ and TEAD under negative regulation by the Hippo signaling pathway.

  19. The ectopic expression of Snail in MDBK cells does not induce epithelial-mesenchymal transition

    OpenAIRE

    IZAWA, GENYA; Kobayashi, Wakako; Haraguchi, Misako; Sudo, Akiharu; Ozawa, Masayuki

    2015-01-01

    Epithelial-mesenchymal transition (EMT), a key process in the tumor metastatic cascade, is characterized by the loss of cell-cell junctions and cell polarity, as well as by the acquisition of migratory and invasive properties. However, the precise molecular events that initiate this complex EMT process are poorly understood. Snail expression induces EMT in Madin-Darby canine kidney (MDCK) cells and the human epidermoid carcinoma cell line, A431. Snail is a zinc finger transcription factor and...

  20. Snail2 is an essential mediator of Twist1-induced epithelial-mesenchymal transition and metastasis

    OpenAIRE

    Casas, Esmeralda; Kim, Jihoon; Bendesky, Andrés; Ohno-Machado, Lucila; Wolfe, Cecily J.; Yang, Jing

    2011-01-01

    To metastasize, carcinoma cells must attenuate cell-cell adhesion to disseminate into distant organs. A group of transcription factors, including Twist1, Snail1, Snail2, ZEB1, and ZEB2, have been shown to induce Epithelial-Mesenchymal Transition (EMT), thus promoting tumor dissemination. However, it is unknown whether these transcription factors function independently or coordinately to activate the EMT program. Here we report that direct induction of Snail2 is essential for Twist1 to induce ...

  1. Expression of epithelial-mesenchymal transition regulators SNAI2 and TWIST1 in thyroid carcinomas.

    Science.gov (United States)

    Buehler, Darya; Hardin, Heather; Shan, Weihua; Montemayor-Garcia, Celina; Rush, Patrick S; Asioli, Sofia; Chen, Herbert; Lloyd, Ricardo V

    2013-01-01

    Epithelial-mesenchymal transition is an important mechanism of epithelial tumor progression, local invasion and metastasis. The E-cadherin (CDH1) repressor SLUG (SNAI2) and the basic helix-loop-helix transcription factor TWIST1 inhibit CDH1 expression in poorly differentiated malignancies as inducers of epithelial-mesenchymal transition. Epithelial-mesenchymal transition has been implicated in progression from well to poorly differentiated/anaplastic thyroid carcinoma but the expression of SNAI2 and TWIST1 proteins and their phenotypic association in human thyroid cancers has not been extensively studied. We examined the expression of SNAI2, TWIST1 and CDH1 by immunohistochemistry in a panel of well-differentiated and anaplastic thyroid cancers and by qRT-PCR in thyroid cell lines. Ten normal thyroids, 33 follicular adenomas, 56 papillary thyroid carcinomas including 28 follicular variants, 27 follicular carcinomas and 10 anaplastic thyroid carcinomas were assembled on a tissue microarray and immunostained for SNAI2, TWIST1 and CDH1. Most (8/10) anaplastic thyroid carcinomas demonstrated strong nuclear immunoreactivity for SNAI2 with associated absence of CDH1 in 6/8 cases (75%). TWIST1 was expressed in 5/10 anaplastic thyroid carcinomas with absence of CDH1 in 3/5 (60%) cases. These findings were confirmed in whole sections of all anaplastic thyroid carcinomas and in a separate validation set of 10 additional anaplastic thyroid carcinomas. All normal thyroids, follicular adenomas, papillary and follicular thyroid carcinomas were negative for SNAI2 and TWIST1 (Pcarcinoma and two anaplastic thyroid carcinoma cell lines tested, but the highest levels of CDH1 mRNA were detected in the normal thyroid cell line while the anaplastic thyroid carcinoma cell line demonstrated the highest levels of SNAI2 and TWIST1 mRNA. Our findings support the role of epithelial-mesenchymal transition in the development of anaplastic thyroid carcinoma.

  2. Inhibition of epithelial-mesenchymal transition in A549 cell by transfected Napsin A

    Institute of Scientific and Technical Information of China (English)

    ZHENG Jin-xu; GUAN Shu-hong; XU Qing; LIU Ji-zhu; SONG Ping

    2012-01-01

    Background Epithelial-mesenchymal transition is a cellular process characterized by the loss of cell adhesion,inhibition of E-cadherin expression,and increased cell mobility.Cells without Napsin A are susceptible to transition.Further studies are required to investigate whether this transition can be reversed by restoration of Napsin A.Methods A Napsin A expression vector PLJM1-Napsin A plasmid was constructed and then transfected into the epithelial cell line A549 by lentivirus transfection to obtain A549-PLJM1-Napsin A cell line.Cell proliferation was assayed by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide and cell cycle was measured by flow cytometry.The E-cadherin,type I collagen,and focal adhesion kinase mRNA level was detected by reverse transcription-polymerase chain reaction.The Napsin A,E-cadherin,type I collagen,and focal adhesion kinase protein level in A549 cells was detected by Westen blotting.Results Transforming growth factor-β1 induced epithelial-mesenchymal transition in A549 cells,as demonstrated by significant reduction of E-cadherin mRNA and protein levels (P <0.01) as well as up-regulation of type I collagen (P <0.01 ).Transfection of Napsin A in A549 cells can partially block the transforming growth factor-β1-regulated expression of E-cadherin and type I collagen (P <0.01).In addition,transforming growth factor-β1-induced cell proliferation was inhibited by Napsin A (P <0.01).Further study demonstrated that Napsin A caused Go/G1 arrest and inhibited the expression of focal adhesion kinase (P <0.01),a key protein in the integrin signaling pathway,in the in vitro epithelial-mesenchymal transition model.Conclusions Sustained Napsin A expression in A549 cells can inhibit the transforming growth factor-β1-induced epithelial-mesenchymal transition.This may be due to the Napsin A-mediated inhibition of focal adhesion kinase expression and integrin signaling pathway.

  3. Insulin resistance and necroinflammation drives ductular reaction and epithelial-mesenchymal transition in chronic hepatitis C

    Science.gov (United States)

    Svegliati-Baroni, Gianluca; Faraci, Graziella; Fabris, Luca; Saccomanno, Stefania; Cadamuro, Massimiliano; Pierantonelli, Irene; Trozzi, Luciano; Bugianesi, Elisabetta; Guido, Maria; Strazzabosco, Mario; Benedetti, Antonio; Marchesini, Giulio

    2013-01-01

    Objective To study the mechanism(s) linking insulin resistance (IR) to hepatic fibrosis and the role of the epithelial component in tissue repair and fibrosis in chronic hepatitis C (CHC). Design Prospective observational study. Setting Tertiary care academic centre. Patients 78 consecutive patients with CHC. Main outcome measures IR, calculated by the oral glucose insulin sensitivity during oral glucose tolerance test; necroinflammatory activity and fibrosis, defined according to Ishak’s score; steatosis, graded as 0 (66%). To evaluate the role of the epithelial component in tissue repair and fibrosis, the expansion of the ductular reaction (DR) was calculated by keratin-7 (CK7) morphometry. Nuclear expression of Snail, downregulation of E-cadherin and expression of fibroblast specific protein-1 (FSP1) and vimentin by CK7-positive cells were used as markers of epithelial-mesenchymal transition in DR elements. Results IR, the degree of necroinflammation and expansion of the DR (stratified as reactive ductular cells (RDCs), hepatic progenitor cells and intermediate hepatobiliary cells according to morphological criteria) were all associated with the stage of fibrosis. Nuclear Snail expression, E-cadherin downregulation and vimentin upregulation were observed in RDCs. By dual immunofluorescence for CK7 and FSP1, the number of RDCs undergoing epithelial-mesenchymal transition progressively increased together with the necroinflammatory score. By multivariate analysis, total inflammation and insulin resistance were the only factors significantly predicting the presence of advanced fibrosis (Ishak score ≥3) and the expansion of RDCs. Conclusion This study indicates that IR is associated with the degree of necroinflammatory injury in CHC and contributes to hepatic fibrosis by stimulating the expansion of RDCs that express epithelial-mesenchymal transition markers. PMID:20966027

  4. The Epithelial-Mesenchymal Transition Factor SNAIL Paradoxically Enhances Reprogramming

    Directory of Open Access Journals (Sweden)

    Juli J. Unternaehrer

    2014-11-01

    Full Text Available Reprogramming of fibroblasts to induced pluripotent stem cells (iPSCs entails a mesenchymal to epithelial transition (MET. While attempting to dissect the mechanism of MET during reprogramming, we observed that knockdown (KD of the epithelial-to-mesenchymal transition (EMT factor SNAI1 (SNAIL paradoxically reduced, while overexpression enhanced, reprogramming efficiency in human cells and in mouse cells, depending on strain. We observed nuclear localization of SNAI1 at an early stage of fibroblast reprogramming and using mouse fibroblasts expressing a knockin SNAI1-YFP reporter found cells expressing SNAI1 reprogrammed at higher efficiency. We further demonstrated that SNAI1 binds the let-7 promoter, which may play a role in reduced expression of let-7 microRNAs, enforced expression of which, early in the reprogramming process, compromises efficiency. Our data reveal an unexpected role for the EMT factor SNAI1 in reprogramming somatic cells to pluripotency.

  5. Arctigenin suppresses transforming growth factor-β1-induced expression of monocyte chemoattractant protein-1 and the subsequent epithelial-mesenchymal transition through reactive oxygen species-dependent ERK/NF-κB signaling pathway in renal tubular epithelial cells.

    Science.gov (United States)

    Li, A; Wang, J; Zhu, D; Zhang, X; Pan, R; Wang, R

    2015-01-01

    Transforming growth factor-β1 (TGF-β1) induces expression of the proinflammatory and profibrotic cytokine monocyte chemoattractant protein-1 (MCP-1) in tubular epithelial cells (TECs) and thereby contributes to the tubular epithelial-mesenchymal transition (EMT), which in turn leads to the progression of tubulointerstitial inflammation into tubulointerstitial fibrosis. Exactly how TGF-β1 causes MCP-1 overexpression and subsequent EMT is not well understood. Using human tubular epithelial cultures, we found that TGF-β1 upregulated the expression of reduced nicotinamide adenine dinucleotide phosphate oxidases 2 and 4 and their regulatory subunits, inducing the production of reactive oxygen species. These reactive species activated a signaling pathway mediated by extracellular signal-regulated kinase (ERK1/2) and nuclear factor-κB (NF-κB), which upregulated expression of MCP-1. Incubating cultures with TGF-β1 was sufficient to induce hallmarks of EMT, such as downregulation of epithelial marker proteins (E-cadherin and zonula occludens-1), induction of mesenchymal marker proteins (α-smooth muscle actin, fibronectin, and vimentin), and elevated cell migration and invasion in an EMT-like manner. Overexpressing MCP-1 in cells exposed to TGF-β1 exacerbated these EMT-like changes. Pretreating cells with the antioxidant and anti-inflammatory compound arctigenin (ATG) protected them against these TGF-β1-induced EMT-like changes; the compound worked by inhibiting the ROS/ERK1/2/NF-κB pathway to decrease MCP-1 upregulation. These findings suggest ATG as a new therapeutic candidate to inhibit or even reverse tubular EMT-like changes during progression to tubulointerstitial fibrosis, and they provide the first clues to how ATG may work.

  6. TGF-β induced epithelial-mesenchymal transition modeling

    Science.gov (United States)

    Xenitidis, P.; Seimenis, I.; Kakolyris, S.; Adamopoulos, A.

    2015-09-01

    Epithelial cells may undergo a process called epithelial to mesenchymal transition (EMT). During EMT, cells lose their epithelial characteristics and acquire a migratory ability. Transforming growth factor-beta (TGF-β) signaling is considered to play an important role in EMT by regulating a set of genes through a gene regulatory network (GRN). This work aims at TGF-β induced EMT GRN modeling using publicly available experimental data (gene expression microarray data). The time-series network identification (TSNI) algorithm was used for inferring the EMT GRN. Receiver operating characteristic (ROC) and precision-recall (P-R) curves were constructed and the areas under them were used for evaluating the algorithm performance regarding network inference.

  7. Research on quercetin in inhibiting the epithelial-mesenchymal transition of gastric cancer

    Institute of Scientific and Technical Information of China (English)

    Xue-Feng Jia; Wen-Ming Chen; Dao-Ping Sun; Hai-Bo Zhao; Shu-Long Jiang; Ning Liu; Fan Yang

    2015-01-01

    Objective:To explore the effect of quercetin on the epithelial-mesenchymal transition of human gastric carcinoma BGC-803 cells induced by TGFβ-1 and its mechanism.Methods:Gastric carcinoma BGC-803 cells were divided into three groups, i.e. normal control group, TGFβ-1, and TGFβ-1 + quercetin group. After 12, 24, 48, and 72 h, MTT was used to detect the cell proliferation capability, scratch repair experiment was used to detect the cell migration ability, and Western blot was utilized for determining the expressions of epithelial-mesenchymal transition factor, E-cadherin, N-cadherin and Vimentin. Meanwhile, the activity of PI3K/AKT signal transduction pathway was evaluated.Results:When compared with the TGFβ-1 group, quercetin reduced the proliferation capability and migration ability of human gastric carcinoma BGC-803 cells. In the TGFβ-1 + quercetin group, E-cadherin expression level was up regulated, N-cadherin, Vimentin, AKT, and p-AKT expression levels were down regulated, and the activity of PI3K/AKT signal transduction pathway was reduced. Conclusions:Quercetin can inhibit the EMT process of human gastric cancer cell lines induced by TGFβ-1. The pathogenetic mechanism is probably associated with the regulation of PI3K/AKT signal transduction pathway activity.

  8. Furin promotes epithelial-mesenchymal transition in pancreatic cancer cells via Hippo-YAP pathway.

    Science.gov (United States)

    Zhang, Youli; Zhou, Meng; Wei, Hong; Zhou, Hailang; He, Junbo; Lu, Ying; Wang, Dawei; Chen, Baoding; Zeng, Jian; Peng, Wanxin; Du, Fengyi; Gong, Aihua; Xu, Min

    2017-04-01

    Furin, a well-characterized proprotein convertase, plays an important role in many diseases and links to tumor metastasis. However, the role of furin in pancreatic cancer progression remains to be elucidated. In the present study, we found that furin promotes the growth and the epithelial-mesenchymal transition (EMT) of pancreatic cancer cells. First, we found that furin knockdown significantly inhibited proliferation, invasion and migration in BxPC3 and SW1990 cells, while furin overexpression promoted the above behavior in PANC1 and PaTu8988 cells. Further evidence revealed that furin knockdown resulted in the upregulation of E-cadherin (epithelial marker), and the downregulation of N-cadherin and Vimentin (mesenchymal markers) in BxPC3 and SW1990 cells, whereas furin overexpression remarkably led to the opposite effects in PANC1 and PaTu8988 cells. Furthermore, our data showed that Furin knockdown, furin inhibitor D6R or overexpression significantly affected YAP phosphoration level and total YAP protein level, indicating that furin was involved in Hippo-YAP pathway. It is suggested that furin promotes epithelial-mesenchymal transition in pancreatic cancer cells probably via Hippo-YAP pathway and may be a potential target for anti-pancreatic cancer.

  9. Epithelial-mesenchymal transition (EMT): A biological process in the development, stem cell differentiation, and tumorigenesis.

    Science.gov (United States)

    Chen, Tong; You, Yanan; Jiang, Hua; Wang, Zack Z

    2017-12-01

    The lineage transition between epithelium and mesenchyme is a process known as epithelial-mesenchymal transition (EMT), by which polarized epithelial cells lose their adhesion property and obtain mesenchymal cell phenotypes. EMT is a biological process that is often involved in embryogenesis and diseases, such as cancer invasion and metastasis. The EMT and the reverse process, mesenchymal-epithelial transition (MET), also play important roles in stem cell differentiation and de-differentiation (or reprogramming). In this review, we will discuss current research progress of EMT in embryonic development, cellular differentiation and reprogramming, and cancer progression, all of which are representative models for researches of stem cell biology in normal and in diseases. Understanding of EMT and MET may help to identify specific markers to distinguish normal stem cells from cancer stem cells in future. © 2017 Wiley Periodicals, Inc.

  10. Single cell migration in oral squamous cell carcinoma - possible evidence of epithelial-mesenchymal transition in vivo

    DEFF Research Database (Denmark)

    Jensen, David H; Reibel, Jesper; Mackenzie, Ian C

    2015-01-01

    BACKGROUND: The invasion of cancer cells into the surrounding normal tissue is one of the defining features of cancer. While the phenomena of tumour budding, epithelial-mesenchymal transition and the presence of myofibroblasts have independently been shown to be related to a poor prognosis of ora...

  11. Epithelial-Mesenchymal Transition (EMT) Gene Variants and Epithelial Ovarian Cancer (EOC) Risk

    DEFF Research Database (Denmark)

    Amankwah, Ernest K.; Lin, Hui-Yi; Tyrer, Jonathan P.

    2015-01-01

    women of European ancestry (1,947 cases and 2,009 controls) and identified 793 variants in 278 EMT-related genes that were nominally (P study of 14,525 invasive-cancer patients and 23,447 controls. A P-value ...Epithelial-mesenchymal transition (EMT) is a process whereby epithelial cells assume mesenchymal characteristics to facilitate cancer metastasis. However, EMT also contributes to the initiation and development of primary tumors. Prior studies that explored the hypothesis that EMT gene variants...... contribute to epithelial ovarian carcinoma (EOC) risk have been based on small sample sizes and none have sought replication in an independent population. We screened 15,816 single-nucleotide polymorphisms (SNPs) in 296 genes in a discovery phase using data from a genome-wide association study of EOC among...

  12. The roles of microRNAs and epithelial-mesenchymal transition in colorectal cancer metastasis

    Institute of Scientific and Technical Information of China (English)

    Ping An; Wei Chen; Yu Zhao; Zhongyin Zhou; Hesheng Luo; Ximing Xu

    2014-01-01

    Colorectal cancer (CRC) is the second most common cause of cancer death worldwide. Distant metastasis is the major cause of death in patients with CRC. During progression to metastasis in which malignant cel s disseminate from the primary tumor to seeding other organs, a multistep process is involved. Cancer cel s proliferate, invade microenvironment, en-ter into the blood circulation, then survive and colonize into distant organs. MicroRNAs (miRNAs) and epithelial-mesenchymal transition (EMT) are key regulators and mechanism in tumorigenesis and cancer metastasis. We review the roles of EMT and microRNAs, especial y EMT related microRNAs in the metastatic pathway of CRC. MicroRNAs provide us a set of potential therapeutic applications and molecular target for CRC.

  13. Mechanisms of disease: epithelial-mesenchymal transition and back again: does cellular plasticity fuel neoplastic progression?

    Energy Technology Data Exchange (ETDEWEB)

    Bissell, Mina J; Turley, Eva A.; Veiseh, Mandana; Radisky, Derek C.; Bissell, Mina J.

    2008-02-13

    Epithelial-mesenchymal transition (EMT) is a conversion that facilitates organ morphogenesis and tissue remodeling in physiological processes such as embryonic development and wound healing. A similar phenotypic conversion is also detected in fibrotic diseases and neoplasia, which is associated with disease progression. EMT in cancer epithelial cells often seems to be an incomplete and bi-directional process. In this Review, we discuss the phenomenon of EMT as it pertains to tumor development, focusing on exceptions to the commonly held rule that EMT promotes invasion and metastasis. We also highlight the role of the RAS-controlled signaling mediators, ERK1, ERK2 and PI3-kinase, as microenvironmental responsive regulators of EMT.

  14. Extracellular matrix proteins regulate epithelial-mesenchymal transition in mammary epithelial cells

    Science.gov (United States)

    Chen, Qike K.; Lee, KangAe; Radisky, Derek C.; Nelson, Celeste M.

    2013-01-01

    Mouse mammary epithelial cells undergo transdifferentiation via epithelial-mesenchymal transition (EMT) upon treatment with matrix metalloproteinase-3 (MMP3). In rigid microenvironments, MMP3 upregulates expression of Rac1b, which translocates to the cell membrane to promote induction of reactive oxygen species and EMT. Here we examine the role of the extracellular matrix (ECM) in this process. Our data show that the basement membrane protein laminin suppresses the EMT response in MMP3-treated cells, whereas fibronectin promotes EMT. These ECM proteins regulate EMT via interactions with their specific integrin receptors. α6-integrin sequesters Rac1b from the membrane and is required for inhibition of EMT by laminin. In contrast, α5-integrin maintains Rac1b at the membrane and is required for the promotion of EMT by fibronectin. Understanding the regulatory role of the ECM will provide insight into mechanisms underlying normal and pathological development of the mammary gland. PMID:23660532

  15. Expression Profiles of Epithelial-Mesenchymal Transition-Associated Proteins in Epithelial Ovarian Carcinoma

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    Mi-Kyung Kim

    2014-01-01

    Full Text Available Epithelial-mesenchymal transition (EMT has been suggested to contribute to tumor progression and acquisition of therapeutic resistance. To assess the clinical significance of EMT-associated proteins, we evaluated the expression of Snail and Slug, the key regulators of EMT, in the primary ovarian cancer samples (n=103 by immunohistochemistry. Snail was differentially expressed according to the histologic subtype (P=0.001 and was predominantly expressed in serous and endometrioid types. In the serous and endometrioid adenocarcinomas, the expression of Snail remained high across the stage and grade, suggesting its role in the early phase of carcinogenesis. However, the expression of Snail and Slug was not related to chemoresistance and poor prognosis and did not serve as independent predictive or prognostic marker.

  16. Dynamic Chromatin Modification Sustains Epithelial-Mesenchymal Transition following Inducible Expression of Snail-1

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    Sarah Javaid

    2013-12-01

    Full Text Available Epithelial-mesenchymal transition (EMT is thought to contribute to cancer metastasis, but its underlying mechanisms are not well understood. To define early steps in this cellular transformation, we analyzed human mammary epithelial cells with tightly regulated expression of Snail-1, a master regulator of EMT. After Snail-1 induction, epithelial markers were repressed within 6 hr, and mesenchymal genes were induced at 24 hr. Snail-1 binding to its target promoters was transient (6–48 hr despite continued protein expression, and it was followed by both transient and long-lasting chromatin changes. Pharmacological inhibition of selected histone acetylation and demethylation pathways suppressed the induction as well as the maintenance of Snail-1-mediated EMT. Thus, EMT involves an epigenetic switch that may be prevented or reversed with the use of small-molecule inhibitors of chromatin modifiers.

  17. Dynamic Chromatin Modification Sustains Epithelial-Mesenchymal Transition following Inducible Expression of Snail-1

    Science.gov (United States)

    Javaid, Sarah; Zhang, Jianmin; Anderssen, Endre; Black, Josh C.; Wittner, Ben S.; Tajima, Ken; Ting, David T.; Smolen, Gromoslaw A.; Zubrowski, Matthew; Desai, Rushil; Maheswaran, Shyamala; Ramaswamy, Sridhar; Whetstine, Johnathan R.; Haber, Daniel A.

    2014-01-01

    SUMMARY Epithelial-mesenchymal transition (EMT) is thought to contribute to cancer metastasis, but its underlying mechanisms are not well understood. To define early steps in this cellular transformation, we analyzed human mammary epithelial cells with tightly regulated expression of Snail-1, a master regulator of EMT. After Snail-1 induction, epithelial markers were repressed within 6 hr, and mesenchymal genes were induced at 24 hr. Snail-1 binding to its target promoters was transient (6–48 hr) despite continued protein expression, and it was followed by both transient and long-lasting chromatin changes. Pharmacological inhibition of selected histone acetylation and demethylation pathways suppressed the induction as well as the maintenance of Snail-1-mediated EMT. Thus, EMT involves an epigenetic switch that may be prevented or reversed with the use of small-molecule inhibitors of chromatin modifiers. PMID:24360956

  18. The role of EPH receptors in cancer-related epithelial-mesenchymal transition

    Institute of Scientific and Technical Information of China (English)

    Rui-Xin Li; Zi-Hua Chen; Zhi-Kang Chen

    2014-01-01

    Erythropoietin-producing hepatoma (EPH) receptors are considered the largest family of receptor tyrosine kinases and play key roles in physiological and pathologic processes in development and disease. EPH receptors are often overexpressed in human malignancies and are associated with poor prognosis. However, the functions of EPH receptors in epithelial-mesenchymal transition (EMT) remain largely unknown. This review depicts the relationship between EPH receptors and the EMT marker E-cadherin as wel as the crosstalk between EPH receptors and the signaling pathways involved EMT. Further discussion is focused on the clinical significance of EPH receptors as candidates for targeting in cancer therapeutics. Final y, we summarize how targeted inhibition of both EPH receptors and EMT-related signaling pathways represents a novel strategy for cancer treatment.

  19. The emerging role of exosomes in Epithelial-Mesenchymal-Transition in cancer.

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    Laura Jayne Vella

    2014-12-01

    Full Text Available Metastasis in cancer consists of multiple steps, including Epithelial-Mesenchymal-Transition (EMT, which is characterized by the loss of Epithelial-like characteristics and the gain of Mesenchymal-like attributes including cell migration and invasion. It is clear that the tumour microenvironment can promote the metastatic cascade and that intercellular communication is necessary for this to occur. Exosomes are small membranous vesicles secreted by most cell types into the extracellular environment and they are important communicators in the tumour microenvironment. They promote angiogenesis, invasion and proliferation in recipient cells to support tumour growth and a prometastatic phenotype. Although it is clear that exosomes contribute to cancer cell plasticity, experimental evidence to define exosome induced plasticity as EMT is only just coming to light. This review will discuss recent research on exosomal regulation of the EMT process in the tumour microenvironment.

  20. miR-506 regulates epithelial mesenchymal transition in breast cancer cell lines.

    Science.gov (United States)

    Arora, Himanshu; Qureshi, Rehana; Park, Woong-Yang

    2013-01-01

    Epithelial-mesenchymal transition (EMT) is an important parameter related to breast cancer survival. Among several microRNAs predicted to target EMT-related genes, miR-506 is a novel miRNA found to be significantly related to breast cancer patient survival in a meta-analysis. miR-506 suppressed the expression of mesenchymal genes such as Vimentin, Snai2, and CD151 in MDA-MB-231 human breast cancer cell line. Moreover, NF-κB bound to the upstream promoter region of miR-506 to suppress transcription. Overexpression of miR-506 inhibited TGFβ-induced EMT and suppressed adhesion, invasion, and migration of MDA-MB-231 cells. From these results, we concluded that miR-506 plays a key role in the process of EMT through posttranslational control of EMT-related genes.

  1. miR-506 regulates epithelial mesenchymal transition in breast cancer cell lines.

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    Himanshu Arora

    Full Text Available Epithelial-mesenchymal transition (EMT is an important parameter related to breast cancer survival. Among several microRNAs predicted to target EMT-related genes, miR-506 is a novel miRNA found to be significantly related to breast cancer patient survival in a meta-analysis. miR-506 suppressed the expression of mesenchymal genes such as Vimentin, Snai2, and CD151 in MDA-MB-231 human breast cancer cell line. Moreover, NF-κB bound to the upstream promoter region of miR-506 to suppress transcription. Overexpression of miR-506 inhibited TGFβ-induced EMT and suppressed adhesion, invasion, and migration of MDA-MB-231 cells. From these results, we concluded that miR-506 plays a key role in the process of EMT through posttranslational control of EMT-related genes.

  2. MicroRNAs:regulators of cancer metastasis and epithelial-mesenchymal transition (EMT)

    Institute of Scientific and Technical Information of China (English)

    Xiang-Ming Ding

    2014-01-01

    Tumor metastasis is the main cause of death in patients with solid tumors. The epithelial-mesenchymal transition (EMT) process, in which epithelial cells are converted into mesenchymal cells, is frequently activated during cancer invasion and metastasis. MicroRNAs (miRNAs) are smal , non-coding RNAs that provide widespread expressional control by repressing mRNA translation and inducing mRNA degradation. The fundamental roles of miRNAs in tumor growth and metastasis have been increasingly wel recognized. A growing number of miRNAs are reported to regulate tumor invasion/metastasis through EMT-related and/or non-EMT-related mechanisms. In this review, we discuss the functional role and molecular mechanism of miRNAs in regulating cancer metastasis and EMT.

  3. Targeting Epithelial-Mesenchymal Transition (EMT) to Overcome Drug Resistance in Cancer.

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    Du, Bowen; Shim, Joong Sup

    2016-07-22

    Epithelial-mesenchymal transition (EMT) is known to play an important role in cancer progression, metastasis and drug resistance. Although there are controversies surrounding the causal relationship between EMT and cancer metastasis, the role of EMT in cancer drug resistance has been increasingly recognized. Numerous EMT-related signaling pathways are involved in drug resistance in cancer cells. Cells undergoing EMT show a feature similar to cancer stem cells (CSCs), such as an increase in drug efflux pumps and anti-apoptotic effects. Therefore, targeting EMT has been considered a novel opportunity to overcome cancer drug resistance. This review describes the mechanism by which EMT contributes to drug resistance in cancer cells and summarizes new advances in research in EMT-associated drug resistance.

  4. Clinicopathological and prognostic significance of epithelial mesenchymal transition-related protein expression in intrahepatic cholangiocarcinoma

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    Yao X

    2012-10-01

    Full Text Available Xing Yao,1,* Xiang Wang,1,* Zishu Wang,2,* Licheng Dai,1 Guolei Zhang,1 Qiang Yan,1 Weimin Zhou11Huzhou Central Hospital, Zhejiang Huzhou, 2Department of Medical Oncology, First Affiliated Hospital, Bengbu Medical College, Anhui, People’s Republic of China *These authors contributed equally to this workBackground: The aim of this study was to examine the patterns of expression of epithelial-mesenchymal transition (EMT-related proteins in intrahepatic cholangiocarcinoma. The clinicopathological and prognostic value of these markers was also evaluated.Methods: We detected the expression status of three EMT-related proteins, ie, E-cadherin, vimentin, and N-cadherin, by immunohistochemistry in consecutive intrahepatic cholangiocarcinoma specimens from 96 patients.Results: The frequency of loss of the epithelial marker E-cadherin, and acquisition of mesenchymal markers, vimentin and N-cadherin, in intrahepatic cholangiocarcinoma was 43.8%, 37.5% and 57.3%, respectively. Altered expression of EMT markers was associated with aggressive tumor behavior, including lymph node metastasis, undifferentiated-type histology, advanced tumor stage, venous invasion, and shorter overall survival. Moreover, loss of E-cadherin was retained as an independent prognostic factor for patients with intrahepatic cholangiocarcinoma in multivariate analysis.Conclusion: Our results suggest that the EMT process is associated with tumor progression and a poor outcome in patients with intrahepatic cholangiocarcinoma, and inhibition of EMT might offer novel promising molecular targets for the treatment of affected patients.Keywords: intrahepatic cholangiocarcinoma, epithelial-mesenchymal transition, expression, prognosis, immunohistochemistry

  5. EGCG Suppresses ERK5 Activation to Reverse Tobacco Smoke-Triggered Gastric Epithelial-Mesenchymal Transition in BALB/c Mice

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    Ling Lu

    2016-07-01

    Full Text Available Tobacco smoke is an important risk factor of gastric cancer. Epithelial-mesenchymal transition is a crucial pathophysiological process in cancer development. ERK5 regulation of epithelial-mesenchymal transition may be sensitive to cell types and/or the cellular microenvironment and its role in the epithelial-mesenchymal transition process remain elusive. Epigallocatechin-3-gallate (EGCG is a promising chemopreventive agent for several types of cancers. In the present study we investigated the regulatory role of ERK5 in tobacco smoke-induced epithelial-mesenchymal transition in the stomach of mice and the preventive effect of EGCG. Exposure of mice to tobacco smoke for 12 weeks reduced expression of epithelial markers E-cadherin, ZO-1, and CK5, while the expression of mesenchymal markers Snail-1, Vimentin, and N-cadherin were increased. Importantly, we demonstrated that ERK5 modulated tobacco smoke-mediated epithelial-mesenchymal transition in mice stomach, as evidenced by the findings that tobacco smoke elevated ERK5 activation, and that tobacco smoke-triggered epithelial-mesenchymal transition was reversed by ERK5 inhibition. Treatment of EGCG (100 mg/kg BW effectively attenuated tobacco smoke-triggered activation of ERK5 and epithelial-mesenchymal transition alterations in mice stomach. Collectively, these data suggested that ERK5 was required for tobacco smoke-triggered gastric epithelial-mesenchymal transition and that EGCG suppressed ERK5 activation to reverse tobacco smoke-triggered gastric epithelial-mesenchymal transition in BALB/c mice. These findings provide new insights into the mechanism of tobacco smoke-associated gastric tumorigenesis and the chemoprevention of tobacco smoke-associated gastric cancer.

  6. Pancreatic stellate cells promote epithelial-mesenchymal transition in pancreatic cancer cells

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    Kikuta, Kazuhiro [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Masamune, Atsushi, E-mail: amasamune@med.tohoku.ac.jp [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Watanabe, Takashi; Ariga, Hiroyuki; Itoh, Hiromichi; Hamada, Shin; Satoh, Kennichi [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Egawa, Shinichi; Unno, Michiaki [Department of Hepatobiliary-Pancreatic Surgery, Tohoku University Graduate School of Medicine, Sendai (Japan); Shimosegawa, Tooru [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan)

    2010-12-17

    Research highlights: {yields} Recent studies have shown that pancreatic stellate cells (PSCs) promote the progression of pancreatic cancer. {yields} Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and scattered, fibroblast-like appearance. {yields} PSCs decreased the expression of epithelial markers but increased that of mesenchymal markers, along with increased migration. {yields} This study suggests epithelial-mesenchymal transition as a novel mechanism by which PSCs contribute to the aggressive behavior of pancreatic cancer cells. -- Abstract: The interaction between pancreatic cancer cells and pancreatic stellate cells (PSCs), a major profibrogenic cell type in the pancreas, is receiving increasing attention. There is accumulating evidence that PSCs promote the progression of pancreatic cancer by increasing cancer cell proliferation and invasion as well as by protecting them from radiation- and gemcitabine-induced apoptosis. Because epithelial-mesenchymal transition (EMT) plays a critical role in the progression of pancreatic cancer, we hypothesized that PSCs promote EMT in pancreatic cancer cells. Panc-1 and SUIT-2 pancreatic cancer cells were indirectly co-cultured with human PSCs isolated from patients undergoing operation for pancreatic cancer. The expression of epithelial and mesenchymal markers was examined by real-time PCR and immunofluorescent staining. The migration of pancreatic cancer cells was examined by scratch and two-chamber assays. Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and a scattered, fibroblast-like appearance. The expression of E-cadherin, cytokeratin 19, and membrane-associated {beta}-catenin was decreased, whereas vimentin and Snail (Snai-1) expression was increased more in cancer cells co-cultured with PSCs than in mono-cultured cells. The migration of pancreatic cancer cells was increased by co-culture with PSCs. The PSC-induced decrease of E-cadherin expression was not altered

  7. Proteinase-activated receptor 4 stimulation-induced epithelial-mesenchymal transition in alveolar epithelial cells

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    Araki Hiromasa

    2007-04-01

    Full Text Available Abstract Background Proteinase-activated receptors (PARs; PAR1–4 that can be activated by serine proteinases such as thrombin and neutrophil catepsin G are known to contribute to the pathogenesis of various pulmonary diseases including fibrosis. Among these PARs, especially PAR4, a newly identified subtype, is highly expressed in the lung. Here, we examined whether PAR4 stimulation plays a role in the formation of fibrotic response in the lung, through alveolar epithelial-mesenchymal transition (EMT which contributes to the increase in myofibroblast population. Methods EMT was assessed by measuring the changes in each specific cell markers, E-cadherin for epithelial cell, α-smooth muscle actin (α-SMA for myofibroblast, using primary cultured mouse alveolar epithelial cells and human lung carcinoma-derived alveolar epithelial cell line (A549 cells. Results Stimulation of PAR with thrombin (1 U/ml or a synthetic PAR4 agonist peptide (AYPGKF-NH2, 100 μM for 72 h induced morphological changes from cobblestone-like structure to elongated shape in primary cultured alveolar epithelial cells and A549 cells. In immunocytochemical analyses of these cells, such PAR4 stimulation decreased E-cadherin-like immunoreactivity and increased α-SMA-like immunoreactivity, as observed with a typical EMT-inducer, tumor growth factor-β (TGF-β. Western blot analyses of PAR4-stimulated A549 cells also showed similar changes in expression of these EMT-related marker proteins. Such PAR4-mediated changes were attenuated by inhibitors of epidermal growth factor receptor (EGFR kinase and Src. PAR4-mediated morphological changes in primary cultured alveolar epithelial cells were reduced in the presence of these inhibitors. PAR4 stimulation increased tyrosine phosphorylated EGFR or tyrosine phosphorylated Src level in A549 cells, and the former response being inhibited by Src inhibitor. Conclusion PAR4 stimulation of alveolar epithelial cells induced epithelial-mesenchymal

  8. Expression of Proteins Involved in Epithelial-Mesenchymal Transition as Predictors of Metastasis and Survival in Breast Cancer Patients

    Science.gov (United States)

    2015-01-01

    biology of metastasis could lead to better stratification of recurrence risk. We proposed to study genes related to epithelial-mesenchymal transition (EMT...pathologist at RPCI and a breast biology researcher at the University at Buffalo (Dr. Patricia Masso-Welch) to identify and implement an appropriate plan for...Abstract nr 3593. DOI:1538-7445.AM2012-3593. iv. Roberts MR, Sucheston- Campbell LE, Zirpoli GR, Bandera EV, Ambrosone CB, and Yao S. Single nucleotide

  9. Matrix stiffness drives Epithelial-Mesenchymal Transition and tumour metastasis through a TWIST1-G3BP2 mechanotransduction pathway

    OpenAIRE

    Wei, SC; Fattet, L; Tsai, JH; Guo, Y.; Pai, VH; Majeski, HE; Chen, AC; Sah, RL; Taylor, SS; ENGLER, AJ; Yang, J.

    2015-01-01

    © 2015 Macmillan Publishers Limited. Matrix stiffness potently regulates cellular behaviour in various biological contexts. In breast tumours, the presence of dense clusters of collagen fibrils indicates increased matrix stiffness and correlates with poor survival. It is unclear how mechanical inputs are transduced into transcriptional outputs to drive tumour progression. Here we report that TWIST1 is an essential mechanomediator that promotes epithelial-mesenchymal transition (EMT) in respon...

  10. The Role of BRCA1 in Suppressing Epithelial-Mesenchymal Transition in Mammary Gland and Tumor Development

    Science.gov (United States)

    2015-09-01

    prognosis triple negative (ER-, PR-, and HER2-) invasive carcinomas with high frequencies of metaplastic and medullary differentiation(23-25). To...Clinically, the majority of CL tumors are poor prognosis triple - negative (ER, PR, and HER2) invasive carcinomas with high frequencies of metaplastic and...AWARD NUMBER: W81XWH-13-1-0282 TITLE: The Role of BRCA1 in Suppressing Epithelial-Mesenchymal Transition in Mammary Gland and Tumor

  11. A link between lipid metabolism and epithelial-mesenchymal transition provides a target for colon cancer therapy

    OpenAIRE

    Sánchez-Martínez, Ruth; Cruz-Gil, Silvia; de Cedrón, Marta Gómez; Álvarez-Fernández, Mónica; Vargas, Teodoro; Molina, Susana; García, Belén; Herranz, Jesús; Moreno-Rubio, Juan; Reglero, Guillermo; Pérez-Moreno, Mirna; Feliu, Jaime; Malumbres, Marcos; de Molina, Ana Ramírez

    2015-01-01

    The alterations in carbohydrate metabolism that fuel tumor growth have been extensively studied. However, other metabolic pathways involved in malignant progression, demand further understanding. Here we describe a metabolic acyl-CoA synthetase/stearoyl-CoA desaturase ACSL/SCD network causing an epithelial-mesenchymal transition (EMT) program that promotes migration and invasion of colon cancer cells. The mesenchymal phenotype produced upon overexpression of these enzymes is reverted through ...

  12. FGFR signaling maintains a drug persistent cell population following epithelial-mesenchymal transition.

    Science.gov (United States)

    Brown, Wells S; Akhand, Saeed Salehin; Wendt, Michael K

    2016-12-13

    An emerging characteristic of drug resistance in cancer is the induction of epithelial-mesenchymal transition (EMT). However, the mechanisms of EMT-mediated drug resistance remain poorly defined. Therefore, we conducted long-term treatments of human epidermal growth factor receptor-2 (Her2)-transformed breast cancer cells with either the EGFR/Her2 kinase inhibitor, Lapatinib or TGF-β, a known physiological inducer of EMT. Both of these treatment regimes resulted in robust EMT phenotypes, but upon withdrawal a subpopulation of TGF-β induced cells readily underwent mesenchymal-epithelial transition, where as Lapatinib-induced cells failed to reestablish an epithelial population. The mesenchymal population that remained following TGF-β stimulation and withdrawal was quickly selected for during subsequent Lapatinib treatment, manifesting in inherent drug resistance. The Nanostring cancer progression gene panel revealed a dramatic upregulation of fibroblast growth factor receptor 1 (FGFR1) and its cognate ligand FGF2 in both acquired and inherent resistance. Mechanistically, FGF:Erk1/2 signaling functions to stabilize the EMT transcription factor Twist and thus maintain the mesenchymal and drug resistant phenotype. Finally, Lapatinib resistant cells could be readily eliminated using recently characterized covalent inhibitors of FGFR. Overall our data demonstrate that next-generation targeting of FGFR can be used in combination with Her2-targeted therapies to overcome resistance in this breast cancer subtype.

  13. Epithelioid peritoneal mesothelioma: a hybrid phenotype within a mesenchymal-epithelial/epithelial-mesenchymal transition framework.

    Science.gov (United States)

    Bozzi, Fabio; Brich, Silvia; Dagrada, Gian Paolo; Negri, Tiziana; Conca, Elena; Cortelazzi, Barbara; Belfiore, Antonino; Perrone, Federica; Gualeni, Ambra Vittoria; Gloghini, Annunziata; Cabras, Antonello; Brenca, Monica; Maestro, Roberta; Zaffaroni, Nadia; Casali, Paolo; Bertulli, Rossella; Deraco, Marcello; Pilotti, Silvana

    2016-11-15

    The aim of this study was to reconsider the biological characteristics of epithelioid malignant peritoneal mesothelioma (E-MpM) in the light of new concepts about epithelial mesenchymal transition and mesenchymal epithelial reverse transition (EMT/MErT) and the role of epigenetic reprogramming in this context. To this end we profiled surgical specimens and derived cells cultures by a number of complementary approaches i.e. immunohistochemistry, immunofluorescence, in situ hybridization, biochemistry, pluripotent stem cell arrays, treatments with cytokines, growth factors and specific inhibitors.The analyses of the surgical specimens showed that i) EZH2 is expressed throughout the spectrum of MpM, ii) that E-MpM (including the high-grade undifferentiated form) are characterised by c-MYC and miRNA 17-5p expression, and iii) that progression to sarcomatoid MpM is dictated by EMT regulators. They also showed that E-MpM expressed c-MET and are enriched in E- and P-cadherins- and VEGFR2-expressing CSCs, thus strongly supporting a role for MErT reprogramming in endowing E-MpM tumour cells with stemness and plasticity, and hence with a drug resistant phenotype. The cell culture-based experiments confirmed the stemness traits and plasticity of E-MpM, and support the view that EZH2 is a druggable target in this tumor.

  14. TGF-β regulates isoform switching of FGF receptors and epithelial-mesenchymal transition.

    Science.gov (United States)

    Shirakihara, Takuya; Horiguchi, Kana; Miyazawa, Keiji; Ehata, Shogo; Shibata, Tatsuhiro; Morita, Ikuo; Miyazono, Kohei; Saitoh, Masao

    2011-02-16

    The epithelial-mesenchymal transition (EMT) is a crucial event in wound healing, tissue repair, and cancer progression in adult tissues. Here, we demonstrate that transforming growth factor (TGF)-β induced EMT and that long-term exposure to TGF-β elicited the epithelial-myofibroblastic transition (EMyoT) by inactivating the MEK-Erk pathway. During the EMT process, TGF-β induced isoform switching of fibroblast growth factor (FGF) receptors, causing the cells to become sensitive to FGF-2. Addition of FGF-2 to TGF-β-treated cells perturbed EMyoT by reactivating the MEK-Erk pathway and subsequently enhanced EMT through the formation of MEK-Erk-dependent complexes of the transcription factor δEF1/ZEB1 with the transcriptional corepressor CtBP1. Consequently, normal epithelial cells that have undergone EMT as a result of combined TGF-β and FGF-2 stimulation promoted the invasion of cancer cells. Thus, TGF-β and FGF-2 may cooperate with each other and may regulate EMT of various kinds of cells in cancer microenvironment during cancer progression.

  15. Epithelial-mesenchymal transition and stemness features in circulating tumor cells from breast cancer patients.

    Science.gov (United States)

    Raimondi, Cristina; Gradilone, Angela; Naso, Giuseppe; Vincenzi, Bruno; Petracca, Arianna; Nicolazzo, Chiara; Palazzo, Antonella; Saltarelli, Rosa; Spremberg, Franco; Cortesi, Enrico; Gazzaniga, Paola

    2011-11-01

    Currently used methods to detect and enumerate circulating tumor cells (CTCs) rely on the expression of the epithelial cell adhesion molecule (EpCAM) and cytokeratins. This selection may exclude cells that have undergone intrinsic modifications of their phenotype, as epithelial-mesenchymal transition (EMT). Aim of the study was to investigate the expression of EMT and stemness markers in CTCs from breast cancer patients in all stages of disease. 92 female breast cancer patients were enrolled. CTCs were isolated by CELLection Dynabeads coated with the monoclonal antibody toward EpCam. Samples found positive for CTCs presence (CD45-/CK+) were evaluated for the expression of ER alpha, HER2, ALDH1, vimentin, and fibronectin. Samples negative for CTCs presence (CD45-/CK-) were also evaluated for the expression of vimentin and fibronectin, used as markers of EMT. CTCs were found in 66% of patients. The distribution of CTCs presence according to stage and grade of disease was found statistically significant. The expression of ALDH1 on CTCs was found to correlate to stage of disease and to the expression of vimentin and fibronectin. In 34% of patients, we detected cells with negative CK/CD45 expression but positive expression of vimentin and fibronectin. There is an urgent need for optimizing CTCs detection methods through the inclusion of EMT markers. The detection of cells in mesenchymal transition, retaining EMT and stemness features, may contribute to discover additional therapeutic targets useful to eradicate micrometastatic disease in breast cancer.

  16. Tangzhiqing Granules Alleviate Podocyte Epithelial-Mesenchymal Transition in Kidney of Diabetic Rats

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    Haiyan Xu

    2017-01-01

    Full Text Available This study discussed the effect of Tangzhiqing granules on podocyte epithelial-mesenchymal transition in kidney of diabetic rats. The diabetic rats were divided randomly into five groups: DM group treated with vehicle, Tangzhiqing granules low-dose treatment group, Tangzhiqing granules middle-dose treatment group, and Tangzhiqing granules high-dose treatment group. Eight Wistar rats used as control group were given saline solution. The intervention was all intragastric administration for 8 weeks. At the end of the 8 weeks, biochemical parameters and kidney weight/body weight ratio were measured. The kidney tissues were observed under light microscope and transmission electron microscopy. To search for the underlying mechanism, we examined the epithelial-to-mesenchymal transition (EMT related molecular markers and TGF-β/smad signaling pathway key proteins expression. The results showed that Tangzhiqing granules relieved the structural damage and functional changes of diabetic kidneys. Kidney podocyte EMT related molecular markers nephrin and CD2AP expression were increased, when desmin and α-SMA levels were decreased by Tangzhiqing granules in diabetic rats. Further TGF-β/smad signaling pathway key proteins TGF-β1 and p-smad2/3 levels were decreased in diabetic rats after treatment with Tangzhiqing granules. These findings suggest that Tangzhiqing granules may protect the podocytes of diabetic nephropathy rats via alleviating podocyte EMT and likely activating TGFβ/smad signaling pathway.

  17. Fluid shear stress induces epithelial-mesenchymal transition (EMT) in Hep-2 cells

    Science.gov (United States)

    Shen, Yang; Zhang, Yingying; Yin, Hongmei; Zeng, Ye; Liu, Jingxia; Yan, Zhiping; Liu, Xiaoheng

    2016-01-01

    Laryngeal squamous cell carcinoma (LSCC) is one of the most commonly diagnosed malignancies with high occurrence of tumor metastasis, which usually exposes to fluid shear stress (FSS) in lymphatic channel and blood vessel. Epithelial-mesenchymal transition (EMT) is an important mechanism that induces metastasis and invasion of tumors. We hypothesized that FSS induced a progression of EMT in laryngeal squamous carcinoma. Accordingly, the Hep-2 cells were exposed to 1.4 dyn/cm2 FSS for different durations. Our results showed that most of cells changed their morphology from polygon to elongated spindle with well-organized F-actin and abundant lamellipodia/filopodia in protrusions. After removing the FSS, cells gradually recovered their flat polygon morphology. FSS induced Hep-2 cells to enhance their migration capacity in a time-dependent manner. In addition, FSS down-regulated E-cadherin, and simultaneously up-regulated N-cadherin, translocated β-catenin into the nucleus. These results confirmed that FSS induced the EMT in Hep-2 cells, and revealed a reversible mesenchymal-epithelial transition (MET) process when FSS was removed. We further examined the time-expressions of signaling cascades, and demonstrated that FSS induces the EMT and enhances cell migration depending on integrin-ILK/PI3K-AKT-Snail signaling events. The current study suggests that FSS, an important biophysical factor in tumor microenvironment, is a potential determinant of cell behavior and function regulation. PMID:27096955

  18. Tumor progression, metastasis, and modulators of epithelial-mesenchymal transition in endometrioid endometrial carcinoma: an update.

    Science.gov (United States)

    Makker, Annu; Goel, Madhu Mati

    2016-02-01

    Endometrioid endometrial carcinoma (EEC), also known as type 1 endometrial cancer (EC), accounts for over 70-80% of all cases that are usually associated with estrogen stimulation and often develops in a background of atypical endometrial hyperplasia. The increased incidence of EC is mainly confined to this type of cancer. Most EEC patients present at an early stage and generally have a favorable prognosis; however, up to 30% of EEC present as high risk tumors, which have invaded deep into the myometrium at diagnosis and progressively lead to local or extra pelvic metastasis. The poor survival of advanced EC is related to the lack of effective therapies, which can be attributed to poor understanding of the molecular mechanisms underlying the progression of disease toward invasion and metastasis. Multiple lines of evidence illustrate that epithelial-mesenchymal transition (EMT)-like events are central to tumor progression and malignant transformation, endowing the incipient cancer cell with invasive and metastatic properties. The aim of this review is to summarize the current knowledge on molecular events associated with EMT in progression, invasion, and metastasis of EEC. Further, the role of epigenetic modifications and microRNA regulation, tumor microenvironment, and microcystic elongated and fragmented glands like invasion pattern have been discussed. We believe this article may perhaps stimulate further research in this field that may aid in identifying high risk patients within this clinically challenging patient group and also lead to the recognition of novel targets for the prevention of metastasis - the most fatal consequence of endometrial carcinogenesis.

  19. CCR7 regulates Twist to induce the epithelial-mesenchymal transition in pancreatic ductal adenocarcinoma.

    Science.gov (United States)

    Li, Kexin; Xu, Baofeng; Xu, Guangying; Liu, Rui

    2016-01-01

    As reported, the CC chemokine receptor 7 (CCR7) trigger a series of signaling cascades in the epithelial-mesenchymal transition (EMT) of some malignancies. Meanwhile, Twist promotes EMT in pancreatic ductal adenocarcinoma (PDAC) progression. Here, effects of Twist on CCR7-induced EMT in the PDAC were investigated in detail. The immunohistochemistry was used to detect the expression of Twist, and then, in vitro assays were applied. The expression rate of Twist was 72.0 % in PDAC samples and closely correlated with tumor-node-metastasis (TNM) stage and invasion. When PDAC cell line PANC1 was subjected to CCL19 stimulation, the expression of p-ERK, p-AKT, Twist, N-cadherin, MMP9, and α-smooth muscle actin (α-SMA) was induced, while the GSK1120212, BEZ235, and MK2206 prohibited the increase of Twist and EMT biomarkers. For another thing, the si-Twist treatment attenuated CCL19-stimulated EMT occurrence, migration, and invasion phenotypes of PANC1 cells. In conclusion, CCR7 pathway up-regulates Twist expression via ERK and PI3K/AKT signaling to manage the EMT of PDAC. Our work allows for clinical gene or protein-targeted regimen of PDAC patients in the near future.

  20. Chronic respiratory aeroallergen exposure in mice induces epithelial-mesenchymal transition in the large airways.

    Directory of Open Access Journals (Sweden)

    Jill R Johnson

    Full Text Available Chronic allergic asthma is characterized by Th2-polarized inflammation and leads to airway remodeling and fibrosis but the mechanisms involved are not clear. To determine whether epithelial-mesenchymal transition contributes to airway remodeling in asthma, we induced allergic airway inflammation in mice by intranasal administration of house dust mite (HDM extract for up to 15 consecutive weeks. We report that respiratory exposure to HDM led to significant airway inflammation and thickening of the smooth muscle layer in the wall of the large airways. Transforming growth factor beta-1 (TGF-β1 levels increased in mouse airways while epithelial cells lost expression of E-cadherin and occludin and gained expression of the mesenchymal proteins vimentin, alpha-smooth muscle actin (α-SMA and pro-collagen I. We also observed increased expression and nuclear translocation of Snail1, a transcriptional repressor of E-cadherin and a potent inducer of EMT, in the airway epithelial cells of HDM-exposed mice. Furthermore, fate-mapping studies revealed migration of airway epithelial cells into the sub-epithelial regions of the airway wall. These results show the contribution of EMT to airway remodeling in chronic asthma-like inflammation and suggest that Th2-polarized airway inflammation can trigger invasion of epithelial cells into the subepithelial regions of the airway wall where they contribute to fibrosis, demonstrating a previously unknown plasticity of the airway epithelium in allergic airway disease.

  1. miR-100 induces epithelial-mesenchymal transition but suppresses tumorigenesis, migration and invasion.

    Directory of Open Access Journals (Sweden)

    Dahu Chen

    2014-02-01

    Full Text Available Whether epithelial-mesenchymal transition (EMT is always linked to increased tumorigenicity is controversial. Through microRNA (miRNA expression profiling of mammary epithelial cells overexpressing Twist, Snail or ZEB1, we identified miR-100 as a novel EMT inducer. Surprisingly, miR-100 inhibits the tumorigenicity, motility and invasiveness of mammary tumor cells, and is commonly downregulated in human breast cancer due to hypermethylation of its host gene MIR100HG. The EMT-inducing and tumor-suppressing effects of miR-100 are mediated by distinct targets. While miR-100 downregulates E-cadherin by targeting SMARCA5, a regulator of CDH1 promoter methylation, this miRNA suppresses tumorigenesis, cell movement and invasion in vitro and in vivo through direct targeting of HOXA1, a gene that is both oncogenic and pro-invasive, leading to repression of multiple HOXA1 downstream targets involved in oncogenesis and invasiveness. These findings provide a proof-of-principle that EMT and tumorigenicity are not always associated and that certain EMT inducers can inhibit tumorigenesis, migration and invasion.

  2. Overexpression of metadherin mediates metastasis of osteosarcoma by regulating epithelial-mesenchymal transition.

    Science.gov (United States)

    Tang, J; Shen, L; Yang, Q; Zhang, C

    2014-10-01

    Osteosarcoma (OS) is one of the most common primary malignant bone tumours of childhood and adolescence, and is characterized by high propensity for metastasis (specially to the lung), which is the main cause of death. However, molecular mechanisms underlying metastasis of OS are still poorly understood. Metadherin (MTDH) was identified to be significantly upregulated in OS tissues that had metastasized compared to OS without metastasis, using a two-dimensional approach of electrophoresis, coupled with mass spectrometry. To understand the function of MTDH in OS, OS cell lines U2OS and SOSP-M were transfected with retroviral shRNA vector against MTDH. It was found that metastatic propensity as well as cell proliferation were significantly reduced in both U2OS and SOSP-M. Migration and invasion of U2OS and SOSP-M cells were significantly lower after knock-down of MTDH. In addition, epithelial-mesenchymal transition (EMT) was reduced after knock-down of MTDH. Clinicopathologically, overexpression of MTDH was significantly associated with metastasis and poor survival of patients with OS. Taken together, our results demonstrate that MTDH mediated metastasis of OS through regulating EMT. This could be an ideal therapeutic target against metastasis of OS. © 2014 John Wiley & Sons Ltd.

  3. CAMK1D amplification implicated in epithelial-mesenchymal transition in basal-like breast cancer.

    Science.gov (United States)

    Bergamaschi, Anna; Kim, Young H; Kwei, Kevin A; La Choi, Yoon; Bocanegra, Melanie; Langerød, Anita; Han, Wonshik; Noh, Dong-Young; Huntsman, David G; Jeffrey, Stefanie S; Børresen-Dale, Anne-Lise; Pollack, Jonathan R

    2008-12-01

    Breast cancer exhibits clinical and molecular heterogeneity, where expression profiling studies have identified five major molecular subtypes. The basal-like subtype, expressing basal epithelial markers and negative for estrogen receptor (ER), progesterone receptor (PR) and HER2, is associated with higher overall levels of DNA copy number alteration (CNA), specific CNAs (like gain on chromosome 10p), and poor prognosis. Discovering the molecular genetic basis of tumor subtypes may provide new opportunities for therapy. To identify the driver oncogene on 10p associated with basal-like tumors, we analyzed genomic profiles of 172 breast carcinomas. The smallest shared region of gain spanned just seven genes at 10p13, including calcium/calmodulin-dependent protein kinase ID (CAMK1D), functioning in intracellular signaling but not previously linked to cancer. By microarray, CAMK1D was overexpressed when amplified, and by immunohistochemistry exhibited elevated expression in invasive carcinomas compared to carcinoma in situ. Engineered overexpression of CAMK1D in non-tumorigenic breast epithelial cells led to increased cell proliferation, and molecular and phenotypic alterations indicative of epithelial-mesenchymal transition (EMT), including loss of cell-cell adhesions and increased cell migration and invasion. Our findings identify CAMK1D as a novel amplified oncogene linked to EMT in breast cancer, and as a potential therapeutic target with particular relevance to clinically unfavorable basal-like tumors.

  4. Andrographolide suppresses epithelial mesenchymal transition by inhibition of MAPK signalling pathway in lens epithelial cells

    Indian Academy of Sciences (India)

    Forum Kayastha; Kaid Johar; Devarshi Gajjar; Anshul Arora; Hardik Madhu; Darshini Ganatra; Abhay Vasavada

    2015-06-01

    Epithelial mesenchymal transition (EMT) of lens epithelial cells (LECs) may contribute to the development of posterior capsular opacification (PCO), which leads to visual impairment. Andrographolide has been shown to have therapeutic potential against various cancers. However, its effect on human LECs is still unknown. The purpose of this study is to evaluate the effect of andrographolide on EMT induced by growth factors in the fetal human lens epithelial cell line (FHL 124). Initially the LECs were treated with growth factors (TGF-2 and bFGF) to induce EMT. Subsequently these EMT-induced cells were treated with andrographolide at 100 and 500 nM concentrations for 24 h. Our results showed that FHL 124 cells treated with growth factors had a significant decrease in protein and m-RNA levels of epithelial markers pax6 and E-Cadherin. After administering andrographolide, these levels significantly increased. It was noticed that EMT markers -SMA, fibronectin and collagen IV significantly decreased after treatment with andrographolide when compared to the other group. Treatment with andrographolide significantly inhibited phosphorylation of ERK and JNK. Cell cycle analysis showed that andrographolide did not arrest cells at G0/G1 or G2/M at tested concentrations. Our findings suggest that andrographolide helps sustain epithelial characteristics by modulating EMT markers and inhibiting the mitogen-activated protein kinase (MAPK) signalling pathway in LECs. Hence it can prove to be useful in curbing EMT-mediated PCO.

  5. Role of epithelial-mesenchymal transition in the enrichment of colorectal cancer stem cells

    Directory of Open Access Journals (Sweden)

    Jia-ping CHENG

    2016-10-01

    Full Text Available Objective  To explore whether the enrichment of cancer stem cells (CSCs in colorectal cancer by suspension culture method is involved with epithelial-mesenchymal transition (EMT. Methods  3D microspheres were cultured by suspension culture method to human colorectal cancer SW620 cells. The 3D microspheres and SW620 cells were used as the research objects. To clarify whether 3D microspheres were enriched with CSCs, we made tumorigenicity experiments in NOD/SCID mice, soft agar cloning experiments, and detected the expression levels of cancer stem cells markers CD44 and Ep-CAM by flow cytometry or by Western blotting. The protein expression levels of EMT markers such as E-cadherin, N-cadherin and vimentin were detected by Western blotting. Results  Compared with the parental SW620 cells, colony formation in vitro (P<0.01 and tumorigenicity in NOD/SCID mice were significantly enhanced, the percentage of CD44-positive cells and Ep-CAM protein expression levels was significantly increased (P<0.01 in 3D microspheres. The protein expression level of epithelial marker E-cadherin was obviously increased (P<0.01, while the protein expression levels of mesenchymal markers N-cadherin and vimentin were significantly decreased (P<0.01. Conclusions  Colorectal cancer stem cells can be enriched by suspension culture method, and the process may be related to EMT. DOI: 10.11855/j.issn.0577-7402.2016.09.03

  6. Transcutaneous carbon dioxide suppresses epithelial-mesenchymal transition in oral squamous cell carcinoma.

    Science.gov (United States)

    Iwata, Eiji; Hasegawa, Takumi; Takeda, Daisuke; Ueha, Takeshi; Kawamoto, Teruya; Akisue, Toshihiro; Sakai, Yoshitada; Komori, Takahide

    2016-04-01

    Oral squamous cell carcinoma (OSCC) is the most common form of oral cancers. Recent studies have shown that the malignant transformation of various carcinomas, including OSCC, is associated with epithelial-mesenchymal transition (EMT), and that expression of the EMT factors are significantly associated with tumor invasion, tumor metastasis, and survival rates in OSCC patients. Hence, there is a possibility that EMT suppression may improve the prognosis of OSCC patients. Hypoxia inducible factor-1α (HIF-1α) is a crucial microenvironmental factor in tumor progression, which induces the expression of EMT factors. We previously reported that transcutaneous CO2 suppresses both human OSCC tumor growth and metastasis to the regional lymph nodes by improving hypoxia in treated tissue. According to this background, we hypothesized that increased EMT with HIF-1α expression may increase the progression and the metastatic potential of OSCC, and that decreased hypoxia by transcutaneous CO2 could suppress EMT. In the present study, in vitro studies showed that hypoxic conditions increased the expression of HIF-1α and EMT factors in OSCC cells. In addition, in vivo studies revealed that transcutaneous CO2 increased E-cadherin expression with the decreased expression of HIF-1α, Snail, Slug, N-cadherin, and Vimentin in tumor treatment. These results suggest that transcutaneous CO2 could suppress EMT by improving hypoxia, resulting in the reduction of metastatic potential of OSCC. The findings indicate that transcutaneous CO2 may be able to improve the prognosis of OSCC patients through the suppression of EMT.

  7. Dlx-2 and glutaminase upregulate epithelial-mesenchymal transition and glycolytic switch

    Science.gov (United States)

    Lee, Su Yeon; Jeon, Hyun Min; Ju, Min Kyung; Jeong, Eui Kyong; Kim, Cho Hee; Park, Hye Gyeong; Han, Song Iy; Kang, Ho Sung

    2016-01-01

    Most cancer cells depend on enhanced glucose and glutamine (Gln) metabolism for growth and survival. Oncogenic metabolism provides biosynthetic precursors for nucleotides, lipids, and amino acids; however, its specific roles in tumor progression are largely unknown. We previously showed that distal-less homeobox-2 (Dlx-2), a homeodomain transcription factor involved in embryonic and tumor development, induces glycolytic switch and epithelial-mesenchymal transition (EMT) by inducing Snail expression. Here we show that Dlx-2 also induces the expression of the crucial Gln metabolism enzyme glutaminase (GLS1), which converts Gln to glutamate. TGF-β and Wnt induced GLS1 expression in a Dlx-2-dependent manner. GLS1 shRNA (shGLS1) suppressed in vivo tumor metastasis and growth. Inhibition of Gln metabolism by shGLS1, Gln deprivation, and Gln metabolism inhibitors (DON, 968 and BPTES) prevented Dlx-2-, TGF-β-, Wnt-, and Snail-induced EMT and glycolytic switch. Finally, shDlx-2 and Gln metabolism inhibition decreased Snail mRNA levels through p53-dependent upregulation of Snail-targeting microRNAs. These results demonstrate that the Dlx-2/GLS1/Gln metabolism axis is an important regulator of TGF-β/Wnt-induced, Snail-dependent EMT, metastasis, and glycolytic switch. PMID:26771232

  8. Bisphenol A regulates Snail-mediated epithelial-mesenchymal transition in hemangioma cells.

    Science.gov (United States)

    Zhai, Denggao; He, Jiantai; Li, Xiaoli; Gong, Liansheng; Ouyang, Yang

    2016-08-01

    Hemangioma (HA) can be exposed to bisphenol A (BPA) through direct skin absorption. Although numerous studies indicated that BPA can trigger the progression of cancers, there is no study concerning the effects of BPA on development of HA. Our present study revealed that nanomolar BPA can significantly increase the in vitro migration and invasion of HA cells via induction of epithelial-mesenchymal transition (EMT), which was evidenced by the upregulation of vimentin and downregulation of E-cadherin. The BPA treatment also significantly increased the expression and nuclear localization of Snail and the key transcription factor of EMT, while it had no effect on the expression of other transcription factors such as Slug, Twist, or ZEB1. Silencing of Snail by small interfering RNAs attenuated BPA-induced downregulation of cadherin and upregulation of vimentin, suggesting that Snail is essential for BPA-induced EMT. Both estrogen receptor α (ERα) and G protein-coupled estrogen receptor (GPER) were expressed in HA cells; furthermore, BPA treatment can increase the expression of both ERα and GPER. However, only the inhibitor of ERα (ICI 182, 780), and not GPER (G15), can abolish BPA-induced upregulation of Snail. It suggested that ERα is involved in BPA-induced EMT of HA cells. Collectively, our data suggested that BPA can trigger the EMT of HA cells via ERα/Snail signals. It indicated that more attention should be paid to the skin exposure to BPA for HA patients. Copyright © 2016 John Wiley & Sons, Ltd.

  9. Hyperglycemia Promotes the Epithelial-Mesenchymal Transition of Pancreatic Cancer via Hydrogen Peroxide

    Directory of Open Access Journals (Sweden)

    Wei Li

    2016-01-01

    Full Text Available Diabetes mellitus (DM and pancreatic cancer are intimately related, as approximately 85% of patients diagnosed with pancreatic cancer have impaired glucose tolerance or even DM. Our previous studies have indicated that high glucose could promote the invasive and migratory abilities of pancreatic cancer cells. We therefore explored the underlying mechanism that hyperglycemia modulates the metastatic potential of pancreatic cancer. Our data showed that streptozotocin- (STZ- treated diabetic nude mice exhibit larger tumor size than that of the euglycemic mice. The number of nude mice that develop liver metastasis or ascites is much more in the STZ-treated group than that in the euglycemic group. Hyperglycemic mice contain a higher plasma H2O2-level than that from euglycemic mice. The injection of polyethylene glycol-conjugated catalase (PEG-CAT, an H2O2 scavenger, may reverse hyperglycemia-induced tumor metastasis. In addition, hyperglycemia could also modulate the expression of epithelial-mesenchymal transition- (EMT- related factors in pancreatic tumor tissues, as the E-cadherin level is decreased and the expression of mesenchymal markers N-cadherin and vimentin as well as transcription factor snail is strongly increased. The injection of PEG-CAT could also reverse hyperglycemia-induced EMT. These results suggest that the association between hyperglycemia and poor prognosis of pancreatic cancer can be attributed to the alterations of EMT through the production of hydrogen peroxide.

  10. Nrf2 inhibits epithelial-mesenchymal transition by suppressing snail expression during pulmonary fibrosis.

    Science.gov (United States)

    Zhou, Wencheng; Mo, Xiaoting; Cui, Wenhui; Zhang, Zhihui; Li, Delin; Li, Liucheng; Xu, Liang; Yao, Hongwei; Gao, Jian

    2016-12-16

    Epithelial-mesenchymal transition (EMT) is a phenotype conversion that plays a critical role in the development of pulmonary fibrosis (PF). It is known that snail could regulate the progression of EMT. Nuclear factor erythroid 2 related factor 2 (Nrf2), a key regulator of antioxidant defense system, protects cells against oxidative stress. However, it is not known whether Nrf2 regulates snail thereby modulating the development of PF. Here, bleomycin (BLM) was intratracheally injected into both Nrf2-knockout (Nrf2(-/-)) and wild-type mice to compare the development of PF. Rat type II alveolar epithelial cells (RLE-6TN) were treated with a specific Nrf2 activator sulforaphane, or transfected with Nrf2 and snail siRNAs to determine their effects on transforming growth factor β1 (TGF-β1)-induced EMT. We found that BLM-induced EMT and lung fibrosis were more severe in Nrf2(-/-) mice compared to wild-type mice. In vitro, sulforaphane treatment attenuated TGF-β1-induced EMT, accompanied by the down-regulation of snail. Inversely, silencing Nrf2 by siRNA enhanced TGF-β1-induced EMT along with increased expression of snail. Interestingly, when snail was silenced by siRNA, sulforaphane treatment was unable to reduce the progression of EMT in RLE-6TN cells. These findings suggest that Nrf2 attenuates EMT and fibrosis process by regulating the expression of snail in PF.

  11. Mechanisms of RON-mediated epithelial-mesenchymal transition in MDCK cells through the MAPK pathway

    Directory of Open Access Journals (Sweden)

    Xu Xiangming

    2011-07-01

    Full Text Available The epithelial-mesenchymal transition (EMT is involved in neoplastic metastasis, and the RON protein may be involved. In the present study, we determined the role and the mechanisms of action of RON in EMT in Madin-Darby canine kidney (MDCK cells by Western blot and cell migration analysis. Activation of RON by macrophage stimulating protein (MSP results in cell migration and initiates changes in the morphology of RON-cDNA-transfected MDCK cells. The absence of E-cadherin, the presence of vimentin and an increase in Snail were observed in RE7 cells, which were derived from MDCK cells transfected with wt-RON, compared with MDCK cells. Stimulation of RE7 cells with MSP resulted in increased migration (about 69% of the wounded areas were covered as well as increased activation of extracellular signal-regulated kinase 1/2 (Erk1/2 and glycogen synthase kinase-3β (GSK-3β; the percent of the activation ratio was 143.6/599.8% and 512.4%, respectively, which could be inhibited with an individual chemical inhibitor PD98059 (50 μM specific to MAPK/ERK kinase (the percent inhibition was 98.9 and 81.2%, respectively. Thus, the results indicated that RON protein could mediate EMT in MDCK cells via the Erk1/2 pathway. Furthermore, GSK-3β regulates the function of Snail in controlling EMT by this pathway.

  12. Fisetin inhibits migration, invasion and epithelial-mesenchymal transition of LMP1-positive nasopharyngeal carcinoma cells.

    Science.gov (United States)

    Li, Rong; Zhao, Yinhai; Chen, Jin; Shao, Songjun; Zhang, Xiujuan

    2014-02-01

    Fisetin (3,3',4',7-tetrahydroxyflavone) has been reported to possess certain anticancer properties. It may inhibit tumor cell proliferation, metastasis and induce apoptosis. However, the effects of fisetin in preventing the metastasis of nasopharyngeal carcinoma (NPC) cells remain to be determined. The epithelial-mesenchymal transition (EMT) is involved in several metastatic malignancies including NPC. It has been reported that the Epstein-Barr virus latent membrane protein-1 (LMP1) induced EMT and is associated with the metastasis of NPC. The aim of this study was to examine the effects of fisetin in preventing the migration and invasion of LMP1-expressing NPC cells (CNE1-LMP1 cells), as well as to investigate whether fisetin may inhibit the molecular changes associated with EMT induced by LMP1. The investigation demonstrated that fisetin suppressed the migration and invasion of CNE1-LMP1 cells under non-cytotoxic concentrations. Fisetin inhibited molecular changes associated with EMT induced by LMP1, upregulated the epithelial marker, E-cadherin protein, and downregulated the mesenchymal marker, vimentin protein, levels. Fisetin also significantly reduced the levels of Twist protein, an EMT regulator. The investigation suggested that fisetin inhibits the migration and invasion of LMP1-positive NPC cells, and the molecular mechanism involves fisetin reversing the EMT induced by LMP1 and downregulates the expression of Twist. This study indicated that fisetin serves as a potential candidate for the treatment of cancer metastasis.

  13. GATA3 inhibits breast cancer metastasis through the reversal of epithelial-mesenchymal transition.

    Science.gov (United States)

    Yan, Wei; Cao, Qing Jackie; Arenas, Richard B; Bentley, Brooke; Shao, Rong

    2010-04-30

    GATA3, a transcription factor that regulates T lymphocyte differentiation and maturation, is exclusively expressed in early stage well differentiated breast cancers but not in advanced invasive cancers. However, little is understood regarding its activity and the mechanisms underlying this differential expression in cancers. Here, we employed GATA3-positive, non-invasive (MCF-7) and GATA3-negative, invasive (MDA-MB-231) breast cancer cells to define its role in the transformation between these two distinct phenotypes. Ectopic expression of GATA3 in MDA-MB-231 cells led to a cuboidal-like epithelial phenotype and reduced cell invasive activity. These cells also increased E-cadherin expression but decreased levels of vimentin, N-cadherin, and MMP-9. Further, MDA-MB-231 cells expressing GATA3 grew smaller primary tumors without metastasis compared with larger metastatic tumors derived from control MDA-MB-231 cells in xenografted mice. GATA3 was found to induce E-cadherin expression through binding GATA-like motifs located in the E-cadherin promoter. Blockade of GATA3 using small interfering RNA gene knockdown in MCF-7 cells triggered fibroblastic transformation and cell invasion, resulting in distant metastasis. Studies of human breast cancer showed that GATA3 expression correlated with elevated E-cadherin levels, ER expression, and long disease-free survival. These data suggest that GATA3 drives invasive breast cancer cells to undergo the reversal of epithelial-mesenchymal transition, leading to the suppression of cancer metastasis.

  14. TP53 Mutation, Epithelial-Mesenchymal Transition, and Stemlike Features in Breast Cancer Subtypes

    Directory of Open Access Journals (Sweden)

    Danila Coradini

    2012-01-01

    Full Text Available Altered p53 protein is prevalently associated with the pathologic class of triple-negative breast cancers and loss of p53 function has recently been linked to the induction of an epithelial-mesenchymal transition (EMT and acquisition of stemness properties. We explored the association between TP53 mutational status and expression of some genes involved in the canonical TGF-β signaling pathway (the most potent EMT inducer and in two early EMT associated events: loss of cell polarity and acquisition of stemness-associated features. We used a publicly accessible microarray dataset consisting of 251 p53-sequenced primary breast cancers. Statistical analysis indicated that mutant p53 tumors (especially those harboring a severe mutation were consistent with the aggressive class of triple-negative cancers and that, differently from cell cultures, surgical tumors underexpressed some TGF-β related transcription factors known as involved in EMT (ID1, ID4, SMAD3, SMAD4, SMAD5, ZEB1. These unexpected findings suggest an interesting relationship between p53 mutation, mammary cell dedifferentiation, and the concomitant acquisition of stemlike properties (as indicated by the overexpression of PROM1 and NOTCH1 genes, which improve tumor cells aggressiveness as indicated by the overexpression of genes associated with cell proliferation (CDK4, CDK6, MKI67 and migration (CXCR4, MMP1.

  15. Systems biology approaches to understanding Epithelial Mesenchymal Transition (EMT) in mucosal remodeling and signaling in asthma

    Science.gov (United States)

    2014-01-01

    A pathological hallmark of asthma is chronic injury and repair, producing dysfunction of the epithelial barrier function. In this setting, increased oxidative stress, growth factor- and cytokine stimulation, together with extracellular matrix contact produces transcriptional reprogramming of the epithelial cell. This process results in epithelial-mesenchymal transition (EMT), a cellular state associated with loss of epithelial polarity, expression of mesenchymal markers, enhanced mobility and extracellular matrix remodeling. As a result, the cellular biology of the EMT state produces characteristic changes seen in severe, refractory asthma: myofibroblast expansion, epithelial trans-differentiation and subepithelial fibrosis. EMT also induces profound changes in epithelial responsiveness that affects innate immune signaling that may have impact on the adaptive immune response and effectiveness of glucocorticoid therapy in severe asthma. We discuss how this complex phenotype is beginning to be understood using systems biology-level approaches through perturbations coupled with high throughput profiling and computational modeling. Understanding the distinct changes induced by EMT at the systems level may provide translational strategies to reverse the altered signaling and physiology of refractory asthma. PMID:24982697

  16. Exosome-Mediated Metastasis: From Epithelial-Mesenchymal Transition to Escape from Immunosurveillance.

    Science.gov (United States)

    Syn, Nicholas; Wang, Lingzhi; Sethi, Gautam; Thiery, Jean-Paul; Goh, Boon-Cher

    2016-07-01

    Exosomes are extracellular signalosomes that facilitate eukaryotic intercellular communication under a wide range of normal physiological contexts. In malignancies, this regulatory circuit is co-opted to promote cancer cell survival and outgrowth. Tumour-derived exosomes (TDEs) carry a pro-EMT (epithelial-mesenchymal transition) programme including transforming growth factor beta (TGFβ), caveolin-1, hypoxia-inducible factor 1 alpha (HIF1α), and β-catenin that enhances the invasive and migratory capabilities of recipient cells, and contributes to stromal remodelling and premetastatic niche formation. The integrin expression patterns on TDEs appear to dictate their preferential uptake by organ-specific cells, implying a crucial role of this pathway in organotropic metastasis. Through the expression of immunomodulatory molecules such as CD39 and CD73, TDEs modify the immune contexture of the tumour microenvironment, which could have implications for immunotherapy. Hence, targeting TDE dysregulation pathways, such as the heparanase/syndecan-1 axis, could represent novel therapeutic strategies in the quest to conquer cancer. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Enhanced proliferation, invasion, and epithelial-mesenchymal transition of nicotine-promoted gastric cancer by periostin

    Institute of Scientific and Technical Information of China (English)

    Yu Liu; Bao-An Liu

    2011-01-01

    AIM: To investigate the contribution of periostin in nicotine-promoted gastric cancer cell proliferation, survival, invasion, drug resistance, and epithelial-mesenchymal transition (EMT). METHODS: Gastric cancer cells were treated with nicotine and periostin protein expression was determined by immunoblotting. Periostin mRNA in gastric cancer cells was silenced using small interfering RNA (siRNA) techniques and periostin gene expression was evaluated by quantitative reverse transcription-polymerase chain reaction. Gastric cancer cells transfected with control or periostin siRNA plasmid were compared in terms of cell proliferation using the methylthiazolyldiphenyl-tetrazolium bromide assay. Cell apoptosis was compared using annexin V-fluoresceine isothiocyanate and propidium iodine double staining. Tumor invasion was determined using the Boyden chamber invasion assay, and the EMT marker Snail expression was evaluated by immunoblotting. RESULTS: Nicotine upregulated periostin in gastric cancer cells through a COX-2 dependent pathway, which was blocked by the COX-2-specific inhibitor NS398. Periostin mRNA expression was decreased by ~87.2% by siRNA in gastric cancer cells, and stable periostinsilenced cells were obtained by G418 screening. Periostin- silenced gastric cancer cells exhibited reduced cell proliferation, elevated sensitivity to chemotherapy with 5-fluorouracil, and decreased cell invasion and Snail expression (P < 0.05). CONCLUSION: Periostin is a nicotine target gene in gastric cancer and plays a role in gastric cancer cell growth, invasion, drug resistance, and EMT facilitated by nicotine.

  18. Curcumin inhibits invasive capabilities through epithelial mesenchymal transition in breast cancer cell lines.

    Science.gov (United States)

    Gallardo, Marcela; Calaf, Gloria M

    2016-09-01

    Curcumin (diferuloyl methane) is an antioxidant that exerts antiproliferative and apoptotic effects and has anti-invasive and anti-metastatic properties. Evidence strongly implicates that epithelial-mesenchymal transition (EMT) is involved in malignant progression affecting genes such as Slug, AXL and Twist1. These genes are abnormally expressed in many tumors and favor metastasis. The purpose of this study was to determine the potential effect of curcumin on EMT, migration and invasion. Triple-positive and triple-negative breast cancer cell lines for estrogen receptor (ER), progesterone receptor (PgR) and HER/neu were used: i) MCF-10F, a normal immortalized breast epithelial cell line (negative), ii) Tumor2, a malignant and tumorigenic cell line (positive) derived from Alpha5 cell line injected into the immunologically depressed mice and transformed by 60/60 cGy doses of high LET (linear energy transfer) α particles (150 keV/µm) of radiation and estrogen, and iii) a commercially available MDA-MB‑231 (negative). The effect of curcumin (30 µM for 48 h) was evaluated on expression of EMT-related genes by RT-qPCR. Results showed that curcumin decreased E-cadherin, N-cadherin, β-catenin, Slug, AXL, Twist1, Vimentin and Fibronectin protein expression, independently of the positivity of the markers in the cell lines. Curcumin also decreased migration and invasive capabilities in comparison to their own controls. It can be concluded that curcumin influenced biochemical changes associated with EMT-related genes that seems to promote such transition and are at the core of several signaling pathways that mediate the transition. Thus, it can be suggested that curcumin is able to prevent or delay cancer progression through the interruption of this process.

  19. The biological and clinical importance of epithelial-mesenchymal transition in circulating tumor cells.

    Science.gov (United States)

    Liu, Huiying; Zhang, Xiaofeng; Li, Jun; Sun, Bin; Qian, Haihua; Yin, Zhengfeng

    2015-02-01

    Movement of tumor cells from a primary tumor to a nonadjacent or distant site is a contiguous and complex process. Among the multiple natural cellular programs that promote initiation and progression of tumor metastasis, epithelial-mesenchymal transition (EMT) may play a key role in the ultimate generation of a metastatic foci. Acquisition of the EMT phenotype by tumor cells not only increases their migration and invasion potentials, thereby facilitating their ability to infiltrate blood vessels and to produce circulating tumor cells (CTCs), but also promotes survival of CTCs in the bloodstream and their ability to extravasate out of the circulatory system and invade proximal tissues. In organs distal to the primary tumor, the phenotypic switching mechanism of mesenchymal-epithelial transition (MET) enables CTCs to grow and colonize, enhancing the likelihood of establishing metastasis. In addition, CTCs that have undergone EMT attain increased resistance to chemotherapy and targeted therapy. CTCs with the EMT phenotype have become recognized as an active source of metastases, and targeting EMT/MET processes during the individual steps of tumor metastasis represents a promising new approach for alleviating cancer metastasis and recurrence. In this article, we focus on the biological and clinical importance of EMT and/or MET in CTCs during the individual steps of tumor metastasis, summarizing the recent findings of the regulatory roles played by EMT and/or MET in the generation, survival, and recolonization of CTCs and discussing the EMT-targeting strategies developed for tumor diagnosis as well as their potential for management of metastatic malignant diseases.

  20. Epithelial Mesenchymal Transition (EMT) in Metastatic Breast Cancer in Omani Women.

    Science.gov (United States)

    Lakhtakia, Ritu; Aljarrah, Adil; Furrukh, Muhammad; Ganguly, Shyam S

    2017-05-19

    Breast cancer (BC) in Oman affects younger women and has a more aggressive course. Clinical and biological variables like age, pregnancy, tumor size, type, grade, receptor expression and proliferation predict disease aggression but there is no direct predictor of metastasis except lymphovascular invasion. Epithelial-mesenchymal transition (EMT) is characterized by epithelial cells losing epithelial and acquiring mesenchymal morpho-immunophenotypic characteristics. In tumors, EMT-like transitions may signify a metastatic phenotype and have features in common with cancer stem cells (CSC) which show resistance to chemotherapy. This study aimed to identify EMT and CSC phenotypes in metastatic and non-metastatic breast cancer in Omani women and their association with conventional clinico-pathological predictors of BC. In a retrospective study of ninety-six Omani women with breast cancer, the association of age, pregnancy/lactation, tumor size, type, grade, ductal carcinoma insitu (DCIS), lymphovascular invasion, hormone/ HER2 receptor expression and Ki67 proliferation index (Ki67 PI) was tested with EMT/ CSC phenotype and metastasis. Young age ≤ 40 years, lymphovascular invasion and EMT had a strong association with metastasis; CSC approached significance. Vimentin expression in tumor cells, fibronectin and MMP-11 in stroma were reliable markers of EMT; dual EMT and CSC phenotype (Vim+/ CD44+/ CD 24-/low) had a strong association with apocrine variant, basal-like tumors and triple negative cancers. EMT had a strong association with Ki67 proliferation index (PI) and CSC with HER2-like tumors and distant metastasis. These select markers may be useful in metastasis-prediction in pre-treatment biopsies.

  1. Chidamide alleviates TGF-β-induced epithelial-mesenchymal transition in lung cancer cell lines.

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    Lin, Sheng-Hao; Wang, Bing-Yen; Lin, Ching-Hsiung; Chien, Peng-Ju; Wu, Yueh-Feng; Ko, Jiunn-Liang; Chen, Jeremy J W

    2016-07-01

    Transforming growth factor-β (TGF-β)-induced epithelial-mesenchymal transition is a critical process in the initiation of metastasis of various types of cancer. Chidamide is a class I histone deacetylase inhibitor with anti-tumor activity. This study investigated the effects of chidamide on TGF-β-mediated suppression of E-cadherin expression in adenocarcinomic lung epithelial cells and the molecular mechanisms involved in these effects. Western blot analysis, confocal microscopy, Quantitative methyl-specific PCR and bisulfite sequencing were used to evaluate the effects of different treatments on chidamide ameliorating TGF-β induced-E-cadherin loss. H3 acetylation binding to the promoter of E-cadherin was detected by chromatin immunoprecipitations (CHIP). We found that chidamide reduced the level of lung cancer cell migration observed using a Boyden chamber assay (as an indicator of metastatic potential). Chidamide inhibited TGF-β-induced SMAD2 phosphorylation and attenuated TGF-β-induced loss of E-cadherin expression in lung cancer cells by Western blotting and confocal microscopy, respectively. Quantitative methyl-specific PCR and bisulfite sequencing revealed that TGF-β-enhanced E-cadherin promoter methylation was ameliorated in cells treated with chidamide. We demonstrated that histone H3 deacetylation within the E-cadherin promoter was required for TGF-β-induced E-cadherin loss; cell treatment with chidamide increased the H3 acetylation detected by CHIP. Taken together, our results demonstrate that TGF-β suppressed E-cadherin expression by regulating promoter methylation and histone H3 acetylation. Chidamide significantly enhanced E-cadherin expression in TGF-β-treated cells and inhibited lung cancer cell migration. These findings indicate that chidamide has a potential therapeutic use due to its capacity to prevent cancer cell metastasis.

  2. Epithelial-Mesenchymal Transition Is a Critical Step in Tumorgenesis of Pancreatic Neuroendocrine Tumors

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    Fendrich, Volker, E-mail: fendrich@med.uni-marburg.de; Maschuw, Katja; Waldmann, Jens [Department of Surgery, Philipps University Marburg, Baldingerstraße, Marburg D-35043 (Germany); Buchholz, Malte [Department of Gastroenterology and Endocrinology, Philipps-University Marburg, Baldingerstraße, Marburg D-35043 (Germany); Rehm, Johannes [Department of Surgery, Philipps University Marburg, Baldingerstraße, Marburg D-35043 (Germany); Gress, Thomas M. [Department of Gastroenterology and Endocrinology, Philipps-University Marburg, Baldingerstraße, Marburg D-35043 (Germany); Bartsch, Detlef K. [Department of Surgery, Philipps University Marburg, Baldingerstraße, Marburg D-35043 (Germany); König, Alexander [Department of Gastroenterology and Endocrinology, Philipps-University Marburg, Baldingerstraße, Marburg D-35043 (Germany)

    2012-03-08

    The transcription factors Snail, Slug and Twist repress E-cadherin and induce epithelial-mesenchymal transition (EMT), a process exploited by invasive cancer cells. In this study, we evaluated the role of EMT in the tumorgenesis of neuroendocrine tumors of the pancreas (PNETs) in vitro, in vivo and human tumor specimen. Expression of EMT markers was analyzed using immunohistochemistry and real-time PCR. For in vitro studies, BON-1 cells were analyzed regarding expression of EMT markers before and after transfection with siRNA against Slug or Snail, and cell aggregation assays were performed. To asses in vivo effects, Rip1Tag2 mice were treated with vehicle or the snail-inhibitor polythlylenglykol from week 5-10 of age. The resected pancreata were evaluated by weight, tumor cell proliferation and apoptosis. Snail and Twist was expressed in 61 % and 64% of PNETs. This was associated with loss of E-cadherin. RT-PCR revealed conservation of the EMT markers Slug and Snail in BON-1 cells. Transfection with siRNA against Slug was associated with upregulation of E-cadherin, enhanced cell-cell adhesion and inhibition of cell proliferation. Snail-inhibition in vivo by PEG was associated with increased apoptosis, decreased tumor cell proliferation and dramatic reduced tumor volume in Rip1Tag2 mice. The presented data show that EMT plays a key role in tumorgenesis of PNETs. The activation of Snail in a considerable subset of human PNETs and the successful effect of Snail inhibition by PEG in islet cell tumors of transgenic mice provides first evidence of Snail as a drug target in PNETs.

  3. Dynamic transcription factor networks in epithelial-mesenchymal transition in breast cancer models.

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    Siletz, Anaar; Schnabel, Michael; Kniazeva, Ekaterina; Schumacher, Andrew J; Shin, Seungjin; Jeruss, Jacqueline S; Shea, Lonnie D

    2013-01-01

    The epithelial-mesenchymal transition (EMT) is a complex change in cell differentiation that allows breast carcinoma cells to acquire invasive properties. EMT involves a cascade of regulatory changes that destabilize the epithelial phenotype and allow mesenchymal features to manifest. As transcription factors (TFs) are upstream effectors of the genome-wide expression changes that result in phenotypic change, understanding the sequential changes in TF activity during EMT provides rich information on the mechanism of this process. Because molecular interactions will vary as cells progress from an epithelial to a mesenchymal differentiation program, dynamic networks are needed to capture the changing context of molecular processes. In this study we applied an emerging high-throughput, dynamic TF activity array to define TF activity network changes in three cell-based models of EMT in breast cancer based on HMLE Twist ER and MCF-7 mammary epithelial cells. The TF array distinguished conserved from model-specific TF activity changes in the three models. Time-dependent data was used to identify pairs of TF activities with significant positive or negative correlation, indicative of interdependent TF activity throughout the six-day study period. Dynamic TF activity patterns were clustered into groups of TFs that change along a time course of gene expression changes and acquisition of invasive capacity. Time-dependent TF activity data was combined with prior knowledge of TF interactions to construct dynamic models of TF activity networks as epithelial cells acquire invasive characteristics. These analyses show EMT from a unique and targetable vantage and may ultimately contribute to diagnosis and therapy.

  4. Dynamic transcription factor networks in epithelial-mesenchymal transition in breast cancer models.

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    Anaar Siletz

    Full Text Available The epithelial-mesenchymal transition (EMT is a complex change in cell differentiation that allows breast carcinoma cells to acquire invasive properties. EMT involves a cascade of regulatory changes that destabilize the epithelial phenotype and allow mesenchymal features to manifest. As transcription factors (TFs are upstream effectors of the genome-wide expression changes that result in phenotypic change, understanding the sequential changes in TF activity during EMT provides rich information on the mechanism of this process. Because molecular interactions will vary as cells progress from an epithelial to a mesenchymal differentiation program, dynamic networks are needed to capture the changing context of molecular processes. In this study we applied an emerging high-throughput, dynamic TF activity array to define TF activity network changes in three cell-based models of EMT in breast cancer based on HMLE Twist ER and MCF-7 mammary epithelial cells. The TF array distinguished conserved from model-specific TF activity changes in the three models. Time-dependent data was used to identify pairs of TF activities with significant positive or negative correlation, indicative of interdependent TF activity throughout the six-day study period. Dynamic TF activity patterns were clustered into groups of TFs that change along a time course of gene expression changes and acquisition of invasive capacity. Time-dependent TF activity data was combined with prior knowledge of TF interactions to construct dynamic models of TF activity networks as epithelial cells acquire invasive characteristics. These analyses show EMT from a unique and targetable vantage and may ultimately contribute to diagnosis and therapy.

  5. BMPER Promotes Epithelial-Mesenchymal Transition in the Developing Cardiac Cushions.

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    Laura Dyer

    Full Text Available Formation of the cardiac valves is an essential component of cardiovascular development. Consistent with the role of the bone morphogenetic protein (BMP signaling pathway in cardiac valve formation, embryos that are deficient for the BMP regulator BMPER (BMP-binding endothelial regulator display the cardiac valve anomaly mitral valve prolapse. However, how BMPER deficiency leads to this defect is unknown. Based on its expression pattern in the developing cardiac cushions, we hypothesized that BMPER regulates BMP2-mediated signaling, leading to fine-tuned epithelial-mesenchymal transition (EMT and extracellular matrix deposition. In the BMPER-/- embryo, EMT is dysregulated in the atrioventricular and outflow tract cushions compared with their wild-type counterparts, as indicated by a significant increase of Sox9-positive cells during cushion formation. However, proliferation is not impaired in the developing BMPER-/- valves. In vitro data show that BMPER directly binds BMP2. In cultured endothelial cells, BMPER blocks BMP2-induced Smad activation in a dose-dependent manner. In addition, BMP2 increases the Sox9 protein level, and this increase is inhibited by co-treatment with BMPER. Consistently, in the BMPER-/- embryos, semi-quantitative analysis of Smad activation shows that the canonical BMP pathway is significantly more active in the atrioventricular cushions during EMT. These results indicate that BMPER negatively regulates BMP-induced Smad and Sox9 activity during valve development. Together, these results identify BMPER as a regulator of BMP2-induced cardiac valve development and will contribute to our understanding of valvular defects.

  6. Overexpression of c-myc induces epithelial mesenchymal transition in mammary epithelial cells.

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    Cho, Kyoung Bin; Cho, Min Kyong; Lee, Won Young; Kang, Keon Wook

    2010-07-28

    The c-myc gene is frequently overexpressed in human breast cancer and its target genes are involved in tumorigenesis. Epithelial mesenchymal transitions (EMT), where cells undergo a developmental switch from a polarized epithelial phenotype to a highly motile mesenchymal phenotype, are associated with invasion and motility of cancer cells. Basal E-cadherin expression was down-regulated in c-myc overexpressing MCF10A (c-myc-MCF10A) cells compared to GFP-overexpressing MCF10A (GFP-MCF10A) cells, while N-cadherin was distinctly increased in c-myc-MCF10A cells. Given that glycogen synthase kinase-3beta (GSK-3beta) and the snail axis have key roles in E-cadherin deregulation during EMT, we investigated the role of GSK-3beta/snail signaling pathways in the induction of EMT by c-myc overexpression. In contrast to GFP-MCF10A cells, both the transcriptional activity and the ubiquitination-dependent protein stability of snail were enhanced in c-myc-MCF10A cells, and this was reversed by GSK-3beta overexpression. We also found that c-myc overexpression inhibits GSK-3beta activity through activation of extracellular signal-regulated kinase (ERK). Inhibition of ERK by dominant negative mutant transfection or chemical inhibitor significantly suppressed snail gene transcription. These results suggest that c-myc overexpression during transformation of mammary epithelial cells (MEC) is involved in EMTs via ERK-dependent GSK-3beta inactivation and subsequent snail activation.

  7. A novel function of HPV16-E6/E7 in epithelial-mesenchymal transition.

    Science.gov (United States)

    Jung, Young-Suk; Kato, Ikuko; Kim, Hyeong-Reh Choi

    2013-06-07

    Human papillomavirus (HPV) 16 is among the most important etiological factors in many human cancers, including head and neck squamous cell carcinomas (HNSCCs) not associated with alcohol or tobacco use. HPV16-E6 and E7 oncoproteins target intracellular signaling networks, altering key molecular and cellular events during tumor progression. The present study investigates the role of HPV16-E6 and E7 oncogenes on the epithelial-mesenchymal transition (EMT), a cellular process thought to be critical for tumor cell invasion and metastasis. Using the epithelial MDCK cell line as an in vitro model, we show that the stable expression of HPV16-E6 or E7 induces morphological conversion from cobblestone-shaped epithelium to spindle-shaped mesenchyme-like phenotype. Consistent with these morphological changes, both E6 and E7 induce expression of the EMT-activating transcriptional factors Slug, Twist, ZEB1 and ZEB2, especially ZEBs, accompanied with switch from epithelial to mesenchymal markers. Importantly, E6 and E7 expression results in induction of the migratory and invasive potential, a functional hallmark of EMT. When we examined the association between HPV16 and the EMT signature in HNSCC cell lines derived from head and neck cancer patients, we found a correlation between HPV16 positivity and the expression of EMT transcription factor ZEB1. Taken together, our findings suggest HPV16 induces EMT-like processes via induction of the EMT transcription factors which may contribute to tumor progression and metastasis. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. Foxn1 Transcription Factor Regulates Wound Healing of Skin through Promoting Epithelial-Mesenchymal Transition.

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    Barbara Gawronska-Kozak

    Full Text Available Transcription factors are key molecules that finely tune gene expression in response to injury. We focused on the role of a transcription factor, Foxn1, whose expression is limited to the skin and thymus epithelium. Our previous studies showed that Foxn1 inactivity in nude mice creates a pro-regenerative environment during skin wound healing. To explore the mechanistic role of Foxn1 in the skin wound healing process, we analyzed post-injured skin tissues from Foxn1::Egfp transgenic and C57BL/6 mice with Western Blotting, qRT-PCR, immunofluorescence and flow cytometric assays. Foxn1 expression in non-injured skin localized to the epidermis and hair follicles. Post-injured skin tissues showed an intense Foxn1-eGFP signal at the wound margin and in leading epithelial tongue, where it co-localized with keratin 16, a marker of activated keratinocytes. This data support the concept that suprabasal keratinocytes, expressing Foxn1, are key cells in the process of re-epithelialization. The occurrence of an epithelial-mesenchymal transition (EMT was confirmed by high levels of Snail1 and Mmp-9 expression as well as through co-localization of vimentin/E-cadherin-positive cells in dermis tissue at four days post-wounding. Involvement of Foxn1 in the EMT process was verified by co-localization of Foxn1-eGFP cells with Snail1 in histological sections. Flow cytometric analysis showed the increase of double positive E-cadherin/N-cadherin cells within Foxn1-eGFP population of post-wounded skin cells isolates, which corroborated histological and gene expression analyses. Together, our findings indicate that Foxn1 acts as regulator of the skin wound healing process through engagement in re-epithelization and possible involvement in scar formation due to Foxn1 activity during the EMT process.

  9. Simvastatin Attenuates TGF-β1-Induced Epithelial-Mesenchymal Transition in Human Alveolar Epithelial Cells

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    Tuo Yang

    2013-06-01

    Full Text Available Background: Transforming growth factor-β1 (TGF-β1-induced epithelial-mesenchymal transition (EMT of alveolar epithelial cells (AEC may contribute to idiopathic pulmonary fibrosis (IPF. TGF-β1-induced EMT in A549 cells (a human AEC cell line resulted in the adoption of mesenchymal responses that were predominantly mediated via the TGF-β1-Smad2/3 signaling pathway. Simvastatin (Sim, a 3-hydroxy-3-methylglutaryl CoA (HMG-CoA reductase inhibitor, has been previously reported to inhibit EMT in human proximal tubular epithelial cells and porcine lens epithelial cells and to suppress Smad2/3 phosphorylation in animal models. However, whether Sim can attenuate TGF-β1-induced EMT in A549 cells and its underlying mechanisms remains unknown. Methods: Cells were incubated with TGF-β1 in the presence or absence of Sim. The epithelial marker E-cadherin (E-Cad and the mesenchymal markers, α-smooth muscle actin (α-SMA, vimentin (Vi and fibronectin (FN, were detected using western blotting analyses and immunofluorescence. Phosphorylated Smad2 and Smad3 levels and connective tissue growth factor (CTGF were analyzed using western blotting. In addition, a cell migration assay was performed. Moreover, the levels of matrix metalloproteinase (MMP-2 and -9 in the culture medium were examined using ELISA. Results: Sim significantly attenuated the TGF-β1-induced decrease in E-Cad levels and elevated the levels of α-SMA, Vi and FN via the suppression of Smad2 and Smad3 phosphorylation. Furthermore, Sim inhibited the mesenchymal-like responses in A549 cells, including cell migration, CTGF expression and secretion of MMP-2 and -9. However, Sim failed to reverse the cell morphologial changes induced by TGF-β1 in A549 cells. Conclusion: Sim attenuated TGF-β1-induced EMT in A549 cells and might be a promising therapeutic agent for treating IPF.

  10. Schisandrin B attenuates cancer invasion and metastasis via inhibiting epithelial-mesenchymal transition.

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    Zhen Liu

    Full Text Available BACKGROUND: Metastasis is the major cause of cancer related death and targeting the process of metastasis has been proposed as a strategy to combat cancer. Therefore, to develop candidate drugs that target the process of metastasis is very important. In the preliminary studies, we found that schisandrin B (Sch B, a naturally-occurring dibenzocyclooctadiene lignan with very low toxicity, could suppress cancer metastasis. METHODOLOGY: BALB/c mice were inoculated subcutaneously or injected via tail vein with murine breast cancer 4T1 cells. Mice were divided into Sch B-treated and control groups. The primary tumor growth, local invasion, lung and bone metastasis, and survival time were monitored. Tumor biopsies were examined immuno- and histo-pathologically. The inhibitory activity of Sch B on TGF-β induced epithelial-mesenchymal transition (EMT of 4T1 and primary human breast cancer cells was assayed. PRINCIPAL FINDINGS: Sch B significantly suppressed the spontaneous lung and bone metastasis of 4T1 cells inoculated s.c. without significant effect on primary tumor growth and significantly extended the survival time of these mice. Sch B did not inhibit lung metastasis of 4T1 cells that were injected via tail vein. Delayed start of treatment with Sch B in mice with pre-existing tumors did not reduce lung metastasis. These results suggested that Sch B acted at the step of local invasion. Histopathological evidences demonstrated that the primary tumors in Sch B group were significantly less locally invasive than control tumors. In vitro assays demonstrated that Sch B could inhibit TGF-β induced EMT of 4T1 cells and of primary human breast cancer cells. CONCLUSIONS: Sch B significantly suppresses the lung and bone metastasis of 4T1 cells via inhibiting EMT, suggesting its potential application in targeting the process of cancer metastasis.

  11. Induction of epithelial-mesenchymal transition (EMT) in breast cancer cells is calcium signal dependent.

    Science.gov (United States)

    Davis, F M; Azimi, I; Faville, R A; Peters, A A; Jalink, K; Putney, J W; Goodhill, G J; Thompson, E W; Roberts-Thomson, S J; Monteith, G R

    2014-05-01

    Signals from the tumor microenvironment trigger cancer cells to adopt an invasive phenotype through epithelial-mesenchymal transition (EMT). Relatively little is known regarding key signal transduction pathways that serve as cytosolic bridges between cell surface receptors and nuclear transcription factors to induce EMT. A better understanding of these early EMT events may identify potential targets for the control of metastasis. One rapid intracellular signaling pathway that has not yet been explored during EMT induction is calcium. Here we show that stimuli used to induce EMT produce a transient increase in cytosolic calcium levels in human breast cancer cells. Attenuation of the calcium signal by intracellular calcium chelation significantly reduced epidermal growth factor (EGF)- and hypoxia-induced EMT. Intracellular calcium chelation also inhibited EGF-induced activation of signal transducer and activator of transcription 3 (STAT3), while preserving other signal transduction pathways such as Akt and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation. To identify calcium-permeable channels that may regulate EMT induction in breast cancer cells, we performed a targeted siRNA-based screen. We found that transient receptor potential-melastatin-like 7 (TRPM7) channel expression regulated EGF-induced STAT3 phosphorylation and expression of the EMT marker vimentin. Although intracellular calcium chelation almost completely blocked the induction of many EMT markers, including vimentin, Twist and N-cadherin, the effect of TRPM7 silencing was specific for vimentin protein expression and STAT3 phosphorylation. These results indicate that TRPM7 is a partial regulator of EMT in breast cancer cells, and that other calcium-permeable ion channels are also involved in calcium-dependent EMT induction. In summary, this work establishes an important role for the intracellular calcium signal in the induction of EMT in human breast cancer cells. Manipulation of

  12. Sub-circuits of a gene regulatory network control a developmental epithelial-mesenchymal transition.

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    Saunders, Lindsay R; McClay, David R

    2014-04-01

    Epithelial-mesenchymal transition (EMT) is a fundamental cell state change that transforms epithelial to mesenchymal cells during embryonic development, adult tissue repair and cancer metastasis. EMT includes a complex series of intermediate cell state changes including remodeling of the basement membrane, apical constriction, epithelial de-adhesion, directed motility, loss of apical-basal polarity, and acquisition of mesenchymal adhesion and polarity. Transcriptional regulatory state changes must ultimately coordinate the timing and execution of these cell biological processes. A well-characterized gene regulatory network (GRN) in the sea urchin embryo was used to identify the transcription factors that control five distinct cell changes during EMT. Single transcription factors were perturbed and the consequences followed with in vivo time-lapse imaging or immunostaining assays. The data show that five different sub-circuits of the GRN control five distinct cell biological activities, each part of the complex EMT process. Thirteen transcription factors (TFs) expressed specifically in pre-EMT cells were required for EMT. Three TFs highest in the GRN specified and activated EMT (alx1, ets1, tbr) and the 10 TFs downstream of those (tel, erg, hex, tgif, snail, twist, foxn2/3, dri, foxb, foxo) were also required for EMT. No single TF functioned in all five sub-circuits, indicating that there is no EMT master regulator. Instead, the resulting sub-circuit topologies suggest EMT requires multiple simultaneous regulatory mechanisms: forward cascades, parallel inputs and positive-feedback lock downs. The interconnected and overlapping nature of the sub-circuits provides one explanation for the seamless orchestration by the embryo of cell state changes leading to successful EMT.

  13. HDAC inhibitors induce epithelial-mesenchymal transition in colon carcinoma cells.

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    Ji, Meiying; Lee, Eun Jeoung; Kim, Ki Bae; Kim, Yangmi; Sung, Rohyun; Lee, Sang-Jeon; Kim, Don Soo; Park, Seon Mee

    2015-05-01

    The effects of histone deacetylase (HDAC) inhibitors on epithelial-mesenchymal transition (EMT) differ in various types of cancers. We investigated the EMT phenotype in four colon cancer cell lines when challenged with HDAC inhibitors trichostatin A (TSA) and valproic acid (VPA) with or without transforming growth factor-β1 (TGF-β1) treatment. Four colon cancer cell lines with different phenotypes in regards to tumorigenicity, microsatellite stability and DNA mutation were used. EMT phenotypes were assessed by the expression of E-cadherin and vimentin using western blot analysis, immunofluorescence, quantitative real-time RT-PCR following treatment with TSA (100 or 200 nM) or VPA (0.5 mM) with or without TGF-β1 (5 ng/ml) for 24 h. Biological EMT phenotypes were also evaluated by cell morphology, migration and invasion assays. TSA or VPA induced mesenchymal features in the colon carcinoma cells by a decrease in E-cadherin and an increase in vimentin expression at the mRNA and protein levels. Confocal microscopy revealed membranous attenuation or nuclear translocation of E-cadherin and enhanced expression of vimentin. These responses occurred after 6 h and increased until 24 h. Colon cancer cells changed from a round or rectangular shape to a spindle shape with increased migration and invasion ability following TSA or VPA treatment. The susceptibility to EMT changes induced by TSA or VPA was comparable in microsatellite stable (SW480 and HT29) and microsatellite unstable cells (DLD1 and HCT116). TSA or VPA induced a mesenchymal phenotype in the colon carcinoma cells and these effects were augmented in the presence of TGF-β1. HDAC inhibitors require careful caution before their application as new anticancer drugs for colon cancers.

  14. Role of PRL-3, Snail, Cytokeratin and Vimentin expression in epithelial mesenchymal transition in breast carcinoma.

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    Patel, Nupur A; Patel, Prabhudas S; Vora, Hemangini H

    2015-01-01

    To study epithelial mesenchymal transition (EMT) in breast cancer, molecules such as PRL-3, Snail, Cytokeratin and Vimentin involved in EMT were evaluated. In this study, m-RNA expression of PRL3 and Snail by RT PCR, protein expression of PRL-3, Snail, Cytokeratin and Vimentin by immunohistochemistry were evaluated on paraffin-embedded tissue sections of 100 patients with breast cancer. PRL3 m-RNA expression (above cut off level > 2487301.00) and PRL-3 protein expression was noted in 52% and 70% of breast carcinoma patients, respectively. The higher incidence of PRL3 protein than m-RNA expression could be due to post translation modification. Further, Snail m-RNA expression (above cut off level > 1285142.00) and Snail protein expression was noted in 53% and 54% of breast cancer patients respectively and Snail protein expression was found significantly higher in patients with pre-menopausal status. The loss of cytokeratin expression in 32% and gain of vimentin expression in 17% was noted in these patients. Vimentin expression was found significantly higher in patients with stage IV disease, BR score 4 and PR negativity. In multivariate survival analysis, Vimentin expression found as strong indicator of biologically aggressive breast cancer predicting reduced disease free survival (DFS) and overall survival (OS). In our study reveals that Vimentin expression emerged as significant biomarker for predicting reduced DFS and OS in breast cancer. The study proposes routine evaluation of Vimentin with other predictive parameters can allow use of EMT inhibitors with conventional therapy to revert EMT in breast cancer.

  15. TRPP2 Enhances Metastasis by Regulating Epithelial-Mesenchymal Transition in Laryngeal Squamous Cell Carcinoma

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    Kaile Wu

    2016-11-01

    Full Text Available Background/Aim: Surgery and chemotherapy treatments of human laryngeal squamous cell carcinoma (HLSCC may fail due to metastasis, in which epithelial-mesenchymal transition (EMT plays an important role. TRPP2, a nonselective cation channel, is expressed in various cell types and participates in many biological processes. Here, we show that TRPP2 enhanced metastasis by regulating EMT. Methods: We used immunohistochemistry, western blotting, Ca2+ imaging, transwell and wound healing assays to investigate TRPP2 expression levels in HLSCC tissue, and the role of TRPP2 in invasion and metastasis of a human laryngocarcinoma cell line (Hep2 cell. Results: We found that TRPP2 protein expression levels were significantly increased in HLSCC tissue; higher TRPP2 levels were associated with decreased patient survival time and degree of differentiation and advanced clinical stage. Knockdown of TRPP2 by transfection with TRPP2 siRNA markedly suppressed ATP-induced Ca2+ release, wound healing, and cell invasion in Hep2 cells. Moreover, TRPP2 siRNA significantly decreased vimentin expression but increased E-cadherin expression in Hep2 cells. In the EMT signalling pathway, TRPP2 siRNA significantly decreased Smad4, STAT3, SNAIL, SLUG and TWIST expression in Hep2 cells. Conclusion: We revealed a previously unknown function of TRPP2 in cancer development and a TRPP2-dependent mechanism underlying laryngocarcinoma cell invasion and metastasis. Our results suggest that TRPP2 may be used as a biomarker for evaluating patient prognosis and as a novel therapeutic target in HLSCC.

  16. Mechanisms of RhoGDI2 Mediated Lung Cancer Epithelial-Mesenchymal Transition Suppression

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    Huiyan Niu

    2014-11-01

    Full Text Available Background: The aim of this study was to evaluate the function of RhoGDI2 in lung cancer epithelial-mesenchymal transition (EMT process and to illustrate the underlying mechanisms that will lead to improvement of lung cancer treatment. Methods: The RhoGDI2 knock-down and overexpressing A549 cell lines were first constructed. The influence of RhoGDI2 on cytoskeleton in A549 cells was studied using two approaches: G-LISA-based Rac1 activity measurement and immunostaining-based F-actin distribution. The expression levels of key EMT genes were analyzed using real time quantitative polymerase chain reaction (RT-qPCR, western blot and immunostaining in untreated and RhoGDI2 knock-down or overexpressing A549 cells in both in vivo and in vitro experimental settings. Results: Our study showed that the activity of Rac1, a key gene that is crucial for the initiation and metastasis of human lung adenocarcinoma, causing the redistribution of F-actin with partial loss of cell-cell adhesions and stress fibers, was significantly suppressed by RhoGDI2. RhoGDI2 promoted the expression of EMT marker gene E-cadherin and repressed EMT promoting genes Slug, Snail, α-SMA in both A549 cells and lung and liver organs derived from the mouse models. Knocking-down RhoGDI2 induced abnormal morphology for lung organs. Conclusion: These findings indicate that RhoGDI2 repressed the activity of Rac1 and may be involved in the rearrangement of cytoskeleton in lung cancer cells. RhoGDI2 suppresses the metastasis of lung cancer mediated through EMT by regulating the expression of key genes such as E-cadherin, Slug, Snail and α-SMA in both in vivo and in vitro models.

  17. Roles of Dietary Phytoestrogens on the Regulation of Epithelial-Mesenchymal Transition in Diverse Cancer Metastasis

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    Geum-A. Lee

    2016-05-01

    Full Text Available Epithelial-mesenchymal transition (EMT plays a key role in tumor progression. The cells undergoing EMT upregulate the expression of cell motility-related proteins and show enhanced migration and invasion. The hallmarks of EMT in cancer cells include changed cell morphology and increased metastatic capabilities in cell migration and invasion. Therefore, prevention of EMT is an important tool for the inhibition of tumor metastasis. A novel preventive therapy is needed, such as treatment of natural dietary substances that are nontoxic to normal human cells, but effective in inhibiting cancer cells. Phytoestrogens, such as genistein, resveratrol, kaempferol and 3,3′-diindolylmethane (DIM, can be raised as possible candidates. They are plant-derived dietary estrogens, which are found in tea, vegetables and fruits, and are known to have various biological efficacies, including chemopreventive activity against cancers. Specifically, these phytoestrogens may induce not only anti-proliferation, apoptosis and cell cycle arrest, but also anti-metastasis by inhibiting the EMT process in various cancer cells. There have been several signaling pathways found to be associated with the induction of the EMT process in cancer cells. Phytoestrogens were demonstrated to have chemopreventive effects on cancer metastasis by inhibiting EMT-associated pathways, such as Notch-1 and TGF-beta signaling. As a result, phytoestrogens can inhibit or reverse the EMT process by upregulating the expression of epithelial phenotypes, including E-cadherin, and downregulating the expression of mesenchymal phenotypes, including N-cadherin, Snail, Slug, and vimentin. In this review, we focused on the important roles of phytoestrogens in inhibiting EMT in many types of cancer and suggested phytoestrogens as prominent alternative compounds to chemotherapy.

  18. Epithelial-Mesenchymal Transition Is a Critical Step in Tumorgenesis of Pancreatic Neuroendocrine Tumors

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    Alexander König

    2012-03-01

    Full Text Available The transcription factors Snail, Slug and Twist repress E-cadherin and induce epithelial-mesenchymal transition (EMT, a process exploited by invasive cancer cells. In this study, we evaluated the role of EMT in the tumorgenesis of neuroendocrine tumors of the pancreas (PNETs in vitro, in vivo and human tumor specimen. Expression of EMT markers was analyzed using immunohistochemistry and real-time PCR. For in vitro studies, BON-1 cells were analyzed regarding expression of EMT markers before and after transfection with siRNA against Slug or Snail, and cell aggregation assays were performed. To asses in vivo effects, Rip1Tag2 mice were treated with vehicle or the snail-inhibitor polythlylenglykol from week 5-10 of age. The resected pancreata were evaluated by weight, tumor cell proliferation and apoptosis. Snail and Twist was expressed in 61 % and 64% of PNETs. This was associated with loss of E-cadherin. RT-PCR revealed conservation of the EMT markers Slug and Snail in BON-1 cells. Transfection with siRNA against Slug was associated with upregulation of E-cadherin, enhanced cell-cell adhesion and inhibition of cell proliferation. Snail-inhibition in vivo by PEG was associated with increased apoptosis, decreased tumor cell proliferation and dramatic reduced tumor volume in Rip1Tag2 mice. The presented data show that EMT plays a key role in tumorgenesis of PNETs. The activation of Snail in a considerable subset of human PNETs and the successful effect of Snail inhibition by PEG in islet cell tumors of transgenic mice provides first evidence of Snail as a drug target in PNETs.

  19. The ectopic expression of Snail in MDBK cells does not induce epithelial-mesenchymal transition.

    Science.gov (United States)

    Izawa, Genya; Kobayashi, Wakako; Haraguchi, Misako; Sudo, Akiharu; Ozawa, Masayuki

    2015-07-01

    Epithelial-mesenchymal transition (EMT), a key process in the tumor metastatic cascade, is characterized by the loss of cell-cell junctions and cell polarity, as well as by the acquisition of migratory and invasive properties. However, the precise molecular events that initiate this complex EMT process are poorly understood. Snail expression induces EMT in Madin-Darby canine kidney (MDCK) cells and the human epidermoid carcinoma cell line, A431. Snail is a zinc finger transcription factor and triggers EMT by suppressing E-cadherin expression. In the present study, to broaden our knowledge of Snail‑induced EMT, we generated stable Snail transfectants using Madin-Darby bovine kidney (MDBK) cells. Contrary to the MDCK or A431 cells examined in our previous studies, the MDBK cells transfected with the Snail construct maintained an epithelial morphology and showed no sign of reduced cell-cell adhesiveness compared to the control cells. Consistent with these observations, the downregulation of epithelial marker proteins, e.g. E-cadherin and desmoglein, and the upregulation of mesenchymal marker proteins, e.g., N-cadherin and fibronectin, were not detected. Furthermore, the E-cadherin promoter was not methylated. Therefore, in the MDBK cells, the ectopic expression of Snail failed to induce EMT. As previously demonstrated, in MDCK cells, Snail expression is accompanied by the increased expression of other EMT-inducing transcription factors, e.g., Slug and zinc finger E-box-binding homeobox 1 (ZEB1). However, the MDBK cells transfected with the Snail construct did not exhibit an increased expression of these factors. Thus, it is possible that the failure to upregulate other EMT-related transcription factors may explain the lack of Snail-mediated induction of EMT in MDBK cells.

  20. Transposon mutagenesis identifies genes and cellular processes driving epithelial-mesenchymal transition in hepatocellular carcinoma

    Science.gov (United States)

    Kodama, Takahiro; Newberg, Justin Y.; Kodama, Michiko; Rangel, Roberto; Yoshihara, Kosuke; Tien, Jean C.; Parsons, Pamela H.; Wu, Hao; Finegold, Milton J.; Copeland, Neal G.; Jenkins, Nancy A.

    2016-01-01

    Epithelial-mesenchymal transition (EMT) is thought to contribute to metastasis and chemoresistance in patients with hepatocellular carcinoma (HCC), leading to their poor prognosis. The genes driving EMT in HCC are not yet fully understood, however. Here, we show that mobilization of Sleeping Beauty (SB) transposons in immortalized mouse hepatoblasts induces mesenchymal liver tumors on transplantation to nude mice. These tumors show significant down-regulation of epithelial markers, along with up-regulation of mesenchymal markers and EMT-related transcription factors (EMT-TFs). Sequencing of transposon insertion sites from tumors identified 233 candidate cancer genes (CCGs) that were enriched for genes and cellular processes driving EMT. Subsequent trunk driver analysis identified 23 CCGs that are predicted to function early in tumorigenesis and whose mutation or alteration in patients with HCC is correlated with poor patient survival. Validation of the top trunk drivers identified in the screen, including MET (MET proto-oncogene, receptor tyrosine kinase), GRB2-associated binding protein 1 (GAB1), HECT, UBA, and WWE domain containing 1 (HUWE1), lysine-specific demethylase 6A (KDM6A), and protein-tyrosine phosphatase, nonreceptor-type 12 (PTPN12), showed that deregulation of these genes activates an EMT program in human HCC cells that enhances tumor cell migration. Finally, deregulation of these genes in human HCC was found to confer sorafenib resistance through apoptotic tolerance and reduced proliferation, consistent with recent studies showing that EMT contributes to the chemoresistance of tumor cells. Our unique cell-based transposon mutagenesis screen appears to be an excellent resource for discovering genes involved in EMT in human HCC and potentially for identifying new drug targets. PMID:27247392

  1. Benzyl isothiocyanate inhibits epithelial-mesenchymal transition in cultured and xenografted human breast cancer cells.

    Science.gov (United States)

    Sehrawat, Anuradha; Singh, Shivendra V

    2011-07-01

    We showed previously that cruciferous vegetable constituent benzyl isothiocyanate (BITC) inhibits growth of cultured and xenografted human breast cancer cells and suppresses mammary cancer development in a transgenic mouse model. We now show, for the first time, that BITC inhibits epithelial-mesenchymal transition (EMT) in human breast cancer cells. Exposure of estrogen-independent MDA-MB-231 and estrogen-responsive MCF-7 human breast cancer cell lines and a pancreatic cancer cell line (PL-45) to BITC resulted in upregulation of epithelial markers (e.g., E-cadherin and/or occludin) with a concomitant decrease in protein levels of mesenchymal markers, including vimentin, fibronectin, snail, and/or c-Met. The BITC-mediated induction of E-cadherin protein was accompanied by an increase in its transcription, whereas BITC-treated MDA-MB-231 cells exhibited suppression of vimentin, snail, and slug mRNA levels. Experimental EMT induced by exposure to TGFβ and TNFα or Rb knockdown in a spontaneously immortalized nontumorigenic human mammary epithelial cell line (MCF-10A) was also partially reversed by BITC treatment. The TGFβ-/TNFα-induced migration of MCF-10A cells was inhibited in the presence of BITC, which was partially attenuated by RNA interference of E-cadherin. Inhibition of MDA-MB-231 xenograft growth in vivo in female athymic mice by BITC administration was associated with an increase in protein level of E-cadherin and suppression of vimentin and fibronectin protein expression. In conclusion, this study reports a novel anticancer effect of BITC involving inhibition of EMT, a process triggered during progression of cancer to invasive state.

  2. Human NUMB6 Induces Epithelial-Mesenchymal Transition and Enhances Breast Cancer Cells Migration and Invasion.

    Science.gov (United States)

    Karaczyn, Aldona A; Adams, Tamara L; Cheng, Robert Y S; Matluk, Nicholas N; Verdi, Joseph M

    2017-02-01

    Mammalian NUMB is alternatively spliced generating four isoforms NUMB1-NUMB4 that can function as tumor suppressors. NUMB1-NUMB4 proteins, which normally determine how different cell types develop, are reduced in 21% of primary breast tumors. Our previous work has, however, indicated that two novel NUMB isoforms, NUMB5 and NUMB6 have the pro-oncogenic functions. Herein, we address a novel function of human NUMB isoform 6 (NUMB6) in promoting cancer cell migration and invasion. We found that NUMB6 induced expression of embryonic transcription factor Slug, which in turn actively repressed E-cadherin, prompting cells to undergo epithelial-mesenchymal transition (EMT). Low-metastatic breast cancer cells DB-7 stably expressing NUMB6, lost their epithelial phenotype, exhibited migratory and pro-invasive behavior, and ultimately elevated expression of mesenchymal markers. Among these markers, increased vimentin, β-catenin, and fibronectin expression elicited metalloproteinase 9 (MMP9) production. Our results revealed that NUMB6-DB-7 cells have significantly increased level of Akt1 and Akt2 phosphorylation. Therefore, antagonizing Akt signaling using a chemical inhibitor LY294002, we found that NUMB6-induced Slug expression was reduced, and ultimately accompanied with decreased cell migration and invasion. In summary, this study identified a novel molecular determinant of breast cancer progression, uncovering a potential oncogenic role for the NUMB6 protein in cancer cell migration and invasion, coupled to the maintenance of mesenchymal-like cells. J. Cell. Biochem. 118: 237-251, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  3. Fourier transform infra-red spectroscopic signatures for lung cells' epithelial mesenchymal transition: A preliminary report

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    Sarkar, Atasi; Sengupta, Sanghamitra; Mukherjee, Anirban; Chatterjee, Jyotirmoy

    2017-02-01

    Infra red (IR) spectral characterization can provide label-free cellular metabolic signatures of normal and diseased circumstances in a rapid and non-invasive manner. Present study endeavoured to enlist Fourier transform infra red (FTIR) spectroscopic signatures for lung normal and cancer cells during chemically induced epithelial mesenchymal transition (EMT) for which global metabolic dimension is not well reported yet. Occurrence of EMT was validated with morphological and immunocytochemical confirmation. Pre-processed spectral data was analyzed using ANOVA and principal component analysis-linear discriminant analysis (PCA-LDA). Significant differences observed in peak area corresponding to biochemical fingerprint (900-1800 cm- 1) and high wave-number (2800-3800 cm- 1) regions contributed to adequate PCA-LDA segregation of cells undergoing EMT. The findings were validated by re-analysis of data using another in-house built binary classifier namely vector valued regularized kernel approximation (VVRKFA), in order to understand EMT progression. To improve the classification accuracy, forward feature selection (FFS) tool was employed in extracting potent spectral signatures by eliminating undesirable noise. Gradual increase in classification accuracy with EMT progression of both cell types indicated prominence of the biochemical alterations. Rapid changes in cellular metabolome noted in cancer cells within first 24 h of EMT induction along with higher classification accuracy for cancer cell groups in comparison to normal cells might be attributed to inherent differences between them. Spectral features were suggestive of EMT triggered changes in nucleic acid, protein, lipid and bound water contents which can emerge as the useful markers to capture EMT related cellular characteristics.

  4. Kaempferol modulates the metastasis of human non-small cell lung cancer cells by inhibiting epithelial-mesenchymal transition

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    Meng Hang

    2015-06-01

    Full Text Available The present study was done to determine whether kaempferol, a natural polyphenol of the flavonoid family, affects Epithelial-Mesenchymal Transition (EMT in non-small cell lung cancer cells. Kaempferol not only inhibited cancer cell proliferation and migration in a dose-dependent manner but also modulated the expression of EMT-related proteins E-cadherin and vimentin which are indispensible to cellular motility, invasiveness and metastasis. These results indicate that kaempferol suppresses non-small cell lung cancer migration by modulating the expression of EMT proteins. Therefore, kaempferol may be useful as a potential anticancer agent for non-small cell lung cancer.

  5. Heterogeneity of expression of epithelial-mesenchymal transition (EMT markers in melanocytes and melanoma cell lines

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    Ji Eun eKim

    2013-05-01

    Full Text Available The epithelial-mesenchymal transition (EMT describes a reversible switch from an epithelial-like to a mesenchymal-like phenotype. It is essential for the development of the normal epithelium and also contributes to the invasive properties of carcinomas. At the molecular level, the EMT transition is characterised by a series of coordinated changes including downregulation of the junctional protein E-cadherin (CDH1, up-regulation of transcriptional repressors of E-cadherin such as Snail (SNAI1 and Slug (SNAI2, and up-regulation of N-cadherin. We wished to determine whether cultured normal melanocytes and melanoma cell lines, which are derived from the neural crest, showed signs of a similarly coordinated phenotypic switch. We investigated normal melanocytes and 25 cell lines derived from New Zealand patients with metastatic melanoma. Most lines had been previously genotyped for common mutations such as BRAF, NRAS, PIK3CA, TP53 and CDKN2A. Expression of E-cadherin, N-cadherin, MITF, Snail, Slug, Axl, p53 and Hdm2 was compared by western blotting. Normal melanocytes expressed each of these proteins except for Snail, while normal melanocytes and almost every melanoma line expressed Slug. Expression of individual markers among different melanoma lines varied from high to low or undetectable. Quantitation of western blots showed that expression of MITF-M, the melanocyte-specific isoform of MITF, was positively related to that of E-cadherin but inversely related to that of N-cadherin and Axl. There was also no apparent relationship between expression of any particular marker and the presence of BRAF, NRAS, PIK3CA, TP53 or CDKN2A mutations. The results suggest that melanomas do not show the classical epithelial and mesenchymal phenotypes but rather display either high E-cadherin/high MITF-M expression on one hand, or high N-cadherin/high Axl expression on the other. These may correspond to differentiated and invasive phenotypes in vivo.

  6. Actin cytoskeleton regulation of epithelial mesenchymal transition in metastatic cancer cells.

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    Jay Shankar

    Full Text Available Epithelial-mesenchymal transition (EMT is associated with loss of the cell-cell adhesion molecule E-cadherin and disruption of cell-cell junctions as well as with acquisition of migratory properties including reorganization of the actin cytoskeleton and activation of the RhoA GTPase. Here we show that depolymerization of the actin cytoskeleton of various metastatic cancer cell lines with Cytochalasin D (Cyt D reduces cell size and F-actin levels and induces E-cadherin expression at both the protein and mRNA level. Induction of E-cadherin was dose dependent and paralleled loss of the mesenchymal markers N-cadherin and vimentin. E-cadherin levels increased 2 hours after addition of Cyt D in cells showing an E-cadherin mRNA response but only after 10-12 hours in HT-1080 fibrosarcoma and MDA-MB-231 cells in which E-cadherin mRNA level were only minimally affected by Cyt D. Cyt D treatment induced the nuclear-cytoplasmic translocation of EMT-associated SNAI 1 and SMAD1/2/3 transcription factors. In non-metastatic MCF-7 breast cancer cells, that express E-cadherin and represent a cancer cell model for EMT, actin depolymerization with Cyt D induced elevated E-cadherin while actin stabilization with Jasplakinolide reduced E-cadherin levels. Elevated E-cadherin levels due to Cyt D were associated with reduced activation of Rho A. Expression of dominant-negative Rho A mutant increased and dominant-active Rho A mutant decreased E-cadherin levels and also prevented Cyt D induction of E-cadherin. Reduced Rho A activation downstream of actin remodelling therefore induces E-cadherin and reverses EMT in cancer cells. Cyt D treatment inhibited migration and, at higher concentrations, induced cytotoxicity of both HT-1080 fibrosarcoma cells and normal Hs27 fibroblasts, but only induced mesenchymal-epithelial transition in HT-1080 cancer cells. Our studies suggest that actin remodelling is an upstream regulator of EMT in metastatic cancer cells.

  7. Inhibition of SDF-1/CXCR4-induced epithelial-mesenchymal transition by kisspeptin-10.

    Science.gov (United States)

    Gründker, Carsten; Bauerschmitz, Gerd; Knapp, Juliane; Schmidt, Elena; Olbrich, Theresa; Emons, Günter

    2015-07-01

    Recently we have shown that breast cancer cell invasion was dramatically increased when co-cultured with MG63 cells. In addition we have generated mesenchymal transformed MCF-7 breast cancer cells (MCF-7-EMT), showing significantly increased invasion in contrast to wild type MCF-7 cells (MCF-7 WT). In this study we have analyzed whether stromal derived factor-1 (SDF-1) is responsible for MCF-7 and T-47-D breast cancer cell invasion and epithelial-mesenchymal-transition (EMT). In addition we have analyzed whether kisspeptin-10 (KP-10) treatment affects SDF-1-induced invasion and EMT. Invasion was quantified by assessment of MCF-7 and T-47-D breast cancer cell migration rate through an artificial basement membrane in a modified Boyden chamber during co-culture with MG63 cells or after treatment with SDF-1α, SDF-1β or the combination of both isoforms. Induction of EMT was verified by analysis of protein expression of epithelial marker E-cadherin (CDH1) and mesenchymal markers N-cadherin (CDH2) and Vimentin (VIM). The role of SDF-1 for invasion and induction of EMT in breast cancer cells was analyzed by blocking SDF-1 secretion during co-culture with MG63 cells. In addition effects of KP-10 treatment on SDF-1-induced invasion and EMT were analyzed. Breast cancer cell invasion was significantly increased when co-cultured with MG63 cells. During co-culture SDF-1 protein expression of MG63 cells was significantly induced. The increased breast cancer cell invasion could be blocked by anti-SDF-1 antibodies. Treatment of breast cancer cells in monoculture (without MG63) with SDF-1α, SDF-1β or the combination of both isoforms resulted in a significant escalation of breast cancer cell invasion and induction of EMT. Protein expression of mesenchymal markers CDH2 and VIM was clearly elevated, whereas protein expression of epithelial marker CDH1 was clearly decreased. The SDF-1-induced increase of cell invasion was significantly reduced after treatment with KP-10. In addition

  8. HNF1α inhibition triggers epithelial-mesenchymal transition in human liver cancer cell lines

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    Vignjevic Danijela

    2011-10-01

    Full Text Available Abstract Background Hepatocyte Nuclear Factor 1α (HNF1α is an atypical homeodomain-containing transcription factor that transactivates liver-specific genes including albumin, α-1-antitrypsin and α- and β-fibrinogen. Biallelic inactivating mutations of HNF1A have been frequently identified in hepatocellular adenomas (HCA, rare benign liver tumors usually developed in women under oral contraceptives, and in rare cases of hepatocellular carcinomas developed in non-cirrhotic liver. HNF1α-mutated HCA (H-HCA are characterized by a marked steatosis and show activation of glycolysis, lipogenesis, translational machinery and mTOR pathway. We studied the consequences of HNF1α silencing in hepatic cell lines, HepG2 and Hep3B and we reproduced most of the deregulations identified in H-HCA. Methods We transfected hepatoma cell lines HepG2 and Hep3B with siRNA targeting HNF1α and obtained a strong inhibition of HNF1α expression. We then looked at the phenotypic changes by microscopy and studied changes in gene expression using qRT-PCR and Western Blot. Results Hepatocytes transfected with HNF1α siRNA underwent severe phenotypic changes with loss of cell-cell contacts and development of migration structures. In HNF1α-inhibited cells, hepatocyte and epithelial markers were diminished and mesenchymal markers were over-expressed. This epithelial-mesenchymal transition (EMT was related to the up regulation of several EMT transcription factors, in particular SNAIL and SLUG. We also found an overexpression of TGFβ1, an EMT initiator, in both cells transfected with HNF1α siRNA and H-HCA. Moreover, TGFβ1 expression is strongly correlated to HNF1α expression in cell models, suggesting regulation of TGFβ1 expression by HNF1α. Conclusion Our results suggest that HNF1α is not only important for hepatocyte differentiation, but has also a role in the maintenance of epithelial phenotype in hepatocytes.

  9. Leptin promotes metastasis by inducing an epithelial-mesenchymal transition in A549 lung cancer cells.

    Science.gov (United States)

    Feng, Helin; Liu, Qingyi; Zhang, Ning; Zheng, Lihua; Sang, Meixiang; Feng, Jiangang; Zhang, Jinming; Wu, Xiangyun; Shan, Baoen

    2013-01-01

    Leptin, an adipocyte-derived cytokine associated with obesity, has been reported to participate in carcinogenesis. Epithelial-mesenchymal transition (EMT) is also considered as a key event in tumor metastasis. The aim of this study is to investigate the mechanism of leptin in the promotion of EMT leading to metastasis in A549 lung cancer cells. We investigated the effect of leptin on migration of A549 cells using wound healing and transwell assays. The incidence of EMT in A549 cells was examined by real-time PCR and immunofluorescence staining. The expression of TGF-β in A549 cells was detected by real-time PCR, and blocking of TGF-β in A549 cells was achieved by siRNA techniques. Additional work was performed using 100 patient samples, which included samples from 50 patients diagnosed with lung cancer and an additional 50 patients diagnosed with lung cancer with metastatic bone lesions. Leptin expression was measured using immunohistochemistry techniques. We demonstrated that leptin can effectively enhance the metastasis of human lung cancer A549 cell line using both wound healing and transwell assays. We also found the incidence of EMT in A549 cells after leptin exposure. Furthermore, we detected the expression of TGF-β in A549 cells, which had been reported to play an important role in inducing EMT. We showed that leptin can significantly upregulate TGF-β at both the mRNA and protein levels in A549 cells. Using siRNA to block the expression of TGF-β in A549 cells, we confirmed the role of TGF-β in the promotion of metastasis and induction of EMT. Furthermore, we found that in patient samples leptin was present at higher levels in samples associated with diagnosis of lung cancer bone metastases tissue than lung cancer tissue. Our results indicated that leptin promoted the metastasis of A549 human lung cancer cell lines by inducing EMT in a TGF-β-dependent manner.

  10. Cell surface glycan alterations in epithelial mesenchymal transition process of Huh7 hepatocellular carcinoma cell.

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    Shan Li

    Full Text Available BACKGROUND AND OBJECTIVE: Due to recurrence and metastasis, the mortality of Hepatocellular carcinoma (HCC is high. It is well known that the epithelial mesenchymal transition (EMT and glycan of cell surface glycoproteins play pivotal roles in tumor metastasis. The goal of this study was to identify HCC metastasis related differential glycan pattern and their enzymatic basis using a HGF induced EMT model. METHODOLOGY: HGF was used to induce HCC EMT model. Lectin microarray was used to detect the expression of cell surface glycan and the difference was validated by lectin blot and fluorescence cell lectin-immunochemistry. The mRNA expression levels of glycotransferases were determined by qRT-PCR. RESULTS: After HGF treatment, the Huh7 cell lost epithelial characteristics and obtained mesenchymal markers. These changes demonstrated that HGF could induce a typical cell model of EMT. Lectin microarray analysis identified a decreased affinity in seven lectins ACL, BPL, JAC, MPL, PHA-E, SNA, and SBA to the glycan of cell surface glycoproteins. This implied that glycan containing T/Tn-antigen, NA2 and bisecting GlcNAc, Siaα2-6Gal/GalNAc, terminal α or βGalNAc structures were reduced. The binding ability of thirteen lectins, AAL, LCA, LTL, ConA, NML, NPL, DBA, HAL, PTL II, WFL, ECL, GSL II and PHA-L to glycan were elevated, and a definite indication that glycan containing terminal αFuc and ± Sia-Le, core fucose, α-man, gal-β(α GalNAc, β1,6 GlcNAc branching and tetraantennary complex oligosaccharides structures were increased. These results were further validated by lectin blot and fluorescence cell lectin-immunochemistry. Furthermore, the mRNA expression level of Mgat3 decreased while that of Mgat5, FucT8 and β3GalT5 increased. Therefore, cell surface glycan alterations in the EMT process may coincide with the expression of glycosyltransferase. CONCLUSIONS: The findings of this study systematically clarify the alterations of cell surface

  11. 14-3-3ε overexpression contributes to epithelial-mesenchymal transition of hepatocellular carcinoma.

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    Tzu-An Liu

    Full Text Available BACKGROUND: 14-3-3ε is implicated in regulating tumor progression, including hepatocellular carcinoma (HCC. Our earlier study indicated that elevated 14-3-3ε expression is significantly associated with higher risk of metastasis and lower survival rates of HCC patients. However, the molecular mechanisms of how 14-3-3ε regulates HCC tumor metastasis are still unclear. METHODOLOGY AND PRINCIPAL FINDINGS: In this study, we show that increased 14-3-3ε expression induces HCC cell migration and promotes epithelial-mesenchymal transition (EMT, which is determined by the reduction of E-cadherin expression and induction of N-cadherin and vimentin expression. Knockdown with specific siRNA abolished 14-3-3ε-induced cell migration and EMT. Furthermore, 14-3-3ε selectively induced Zeb-1 and Snail expression, and 14-3-3ε-induced cell migration was abrogated by Zeb-1 or Snail siRNA. In addition, the effect of 14-3-3ε-reduced E-cadherin was specifically restored by Zeb-1 siRNA. Positive 14-3-3ε expression was significantly correlated with negative E-cadherin expression, as determined by immunohistochemistry analysis in HCC tumors. Analysis of 14-3-3ε/E-cadherin expression associated with clinicopathological characteristics revealed that the combination of positive 14-3-3ε and negative E-cadherin expression is significantly correlated with higher incidence of HCC metastasis and poor 5-year overall survival. In contrast, patients with positive 14-3-3ε and positive E-cadherin expression had better prognostic outcomes than did those with negative E-cadherin expression. SIGNIFICANCE: Our findings show for the first time that E-cadherin is one of the downstream targets of 14-3-3ε in modulating HCC tumor progression. Thus, 14-3-3ε may act as an important regulator in modulating tumor metastasis by promoting EMT as well as cell migration, and it may serve as a novel prognostic biomarker or therapeutic target for HCC.

  12. Expression of epithelial-mesenchymal transition markers at the invasive front of oral squamous cell carcinoma

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    Liana Cristina Melo Carneiro COSTA

    2015-04-01

    Full Text Available Oral squamous cell carcinoma (OSCC is one of the most common malignances. In epithelial-mesenchymal transition (EMT, epithelial cells switch to mesenchymal-like cells exhibiting high mobility. This migratory phenotype is significant during tumor invasion and metastasis. Objective : The aim of this study is to evaluate the expression of the EMT markers E-cadherin, N-cadherin and vimentin in OSCC. Material and Methods : Immunohistochemical detection of E-cadherin, N-cadherin and vimentin was performed on 20 OSCC samples. Differences in the expression of each protein at the invasive front (IF and in the central/superficial areas (CSA of the tumor were assessed. Differences in the expression of each protein at the IF of both histologically high- and low-invasive OSCCs were evaluated. Associations among expression of proteins at the IF were assessed. Correlations between the expression levels of each protein at the IF and the tumor stage and clinical nodal status were also evaluated. Results : Reduced expression of E-cadherin was detected in 15 samples (75%. E-cadherin expression was reduced at the IF when compared to the CSA and in high-invasive tumors when compared to low-invasive tumors. All samples were negative for N-cadherin, even though one sample showed an inconspicuous expression. Positive expression of vimentin was observed in 6 samples (30%. Nevertheless, there was no difference in vimentin expression between the IF and the CSA regions or between the low- and high-invasive tumors. Furthermore, no association was observed among protein expression levels at the IF. Finally, no correlations were observed between each protein’s expression levels and tumor stage or clinical nodal status. Conclusions : Reduced E-cadherin expression at the IF and its association with histological invasiveness suggest that this protein is a noteworthy EMT marker in OSCC. Although vimentin was also detected as an EMT marker, its expression was neither limited to

  13. The effect of statin on epithelial-mesenchymal transition in peritoneal mesothelial cells.

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    Tae Ik Chang

    Full Text Available Statins have recently been highlighted for their pleiotropic actions distinct from cholesterol-lowering effects. Despite this interest, it is currently unknown whether statin therapy inhibits peritoneal dialysis (PD-related epithelial-mesenchymal transition (EMT.In vitro, human peritoneal mesothelial cells (HPMCs were exposed to 5.6 mM glucose (NG or 100 mM glucose (HG with or without simvastatin (1 µM. In vivo, PD catheters were inserted into 32 Sprague-Dawley rats, and saline (C, n = 16 or 4.25% peritoneal dialysis fluid (PDF (PD, n = 16 was infused for 4 weeks. Eight rats from each group were treated with 5 mg/kg/day of simvastatin intraperitoneally. Changes in the protein expression of EMT markers such as E-cadherin, α-SMA, Snail, and fibronectin in HPMCs and the peritoneum were evaluated by Western blot analysis and immunofluorescence or immunohistochemical staining. We also explored whether activation of the mevalonate pathway and its downstream small GTPases were involved in dialysis-related peritoneal EMT and could be inhibited by statin treatment.Compared to NG cells, E-cadherin expression was significantly decreased, while α-SMA, Snail, and fibronectin expression were significantly increased in HPMCs exposed to HG, and these changes were abrogated by simvastatin (p<0.05. In addition, the cobblestone-like appearance of normal HPMCs was converted into a fibroblast-like morphology after HG treatment, which was reversed by simvastatin. These EMT-like changes were also observed in HPMCs treated with geranyl-geranyl pyrophosphate (5 µM. HG significantly increased the protein expression of RhoA and Rac1 in the membrane fractions, and these increases were ameliorated by simvastatin (p<0.05. In PD rats, E-cadherin in the peritoneum was significantly decreased, whereas α-SMA, Snail, and fibronectin expression were significantly increased (p<0.05 compared to C rats. The thickness of the mesothelial layer in the peritoneum were also

  14. Role of epithelial mesenchymal transition (EMT in pancreatic ductal adenocarcinoma (PDAC: is tumor budding the missing link?

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    Eva eKaramitopoulou

    2013-09-01

    Full Text Available Pancreatic ductal adenocarcinoma (PDAC ranks as the fourth commonest cause of cancer death while its incidence is increasing worldwide. For all stages, survival at 5 years is <5%. The lethal nature of pancreatic cancer is attributed to its high metastatic potential to the lymphatic system and distant organs. Lack of effective therapeutic options contributes to the high mortality rates of PDAC. Recent evidence suggests that epithelial-mesenchymal transition (EMT plays an important role to the disease progression and development of drug resistance in PDAC. Tumor budding is thought to reflect the process of epithelial-mesenchymal transition (EMT which allows neoplastic epithelial cells to acquire a mesenchymal phenotype thus increasing their capacity for migration and invasion and help them become resistant to apoptotic signals. In a recent study by our own group the presence and prognostic significance of tumor budding in PDAC were investigated and an association between high-grade budding and aggressive clinicopathological features of the tumors as well as worse outcome of the patients was found. The identification of EMT phenotypic targets may help identifying new molecules so that future therapeutic strategies directed specifically against them could potentially have an impact on drug resistance and invasiveness and hence improve the prognosis of PDAC patients. The aim of this short review is to present an insight on the morphological and molecular aspects of EMT and on the factors that are involved in the induction of EMT in PDAC.

  15. BMP4 Signaling Is Able to Induce an Epithelial-Mesenchymal Transition-Like Phenotype in Barrett's Esophagus and Esophageal Adenocarcinoma through Induction of SNAIL2

    NARCIS (Netherlands)

    Kestens, C.; Siersema, P.D.; Offerhaus, G.J.; Baal, J.W.P.M. van

    2016-01-01

    BACKGROUND: Bone morphogenetic protein 4 (BMP4) signaling is involved in the development of Barrett's esophagus (BE), a precursor of esophageal adenocarcinoma (EAC). In various cancers, BMP4 has been found to induce epithelial-mesenchymal transition (EMT) but its function in the development of EAC

  16. BMP4 signaling is able to induce an epithelial-mesenchymal transition-like phenotype in Barrett's esophagus and esophageal adenocarcinoma through induction of SNAIL2

    NARCIS (Netherlands)

    Kestens, Christine; Siersema, Peter D; Offerhaus, G Johan A; van Baal, Jantine W P M

    2016-01-01

    Background: Bone morphogenetic protein 4 (BMP4) signaling is involved in the development of Barrett's esophagus (BE), a precursor of esophageal adenocarcinoma (EAC). In various cancers, BMP4 has been found to induce epithelial-mesenchymal transition (EMT) but its function in the development of EAC

  17. Elucidation of epithelial-mesenchymal transition-related pathways in a triple-negative breast cancer cell line model by multi-omics interactome analysis

    DEFF Research Database (Denmark)

    Pauling, Josch K; Christensen, Anne G; Batra, Richa

    2014-01-01

    obtained from a triple-negative breast cancer cell line model, combining data sets of gene and protein expression as well as protein phosphorylation. We focus on alterations associated with the phenotypical differences arising from epithelial-mesenchymal transition in two breast cancer cell lines...

  18. Comprehensive study of gene and microRNA expression related to epithelial-mesenchymal transition in prostate cancer.

    Directory of Open Access Journals (Sweden)

    Betina Katz

    Full Text Available Prostate cancer is the most common cancer in men, and most patients have localized disease at the time of diagnosis. However, 4% already present with metastatic disease. Epithelial-mesenchymal transition is a fundamental process in carcinogenesis that has been shown to be involved in prostate cancer progression. The main event in epithelial-mesenchymal transition is the repression of E-cadherin by transcription factors, but the process is also regulated by microRNAs. The aim of this study was to analyze gene and microRNA expression involved in epithelial-mesenchymal transition in localized prostate cancer and metastatic prostate cancer cell lines and correlate with clinicopathological findings. We studied 51 fresh frozen tissue samples from patients with localized prostate cancer (PCa treated by radical prostatectomy and three metastatic prostate cancer cell lines (LNCaP, DU145, PC3. The expression of 10 genes and 18 miRNAs were assessed by real-time PCR. The patients were divided into groups according to Gleason score, pathological stage, preoperative PSA, biochemical recurrence, and risk group for correlation with clinicopathological findings. The majority of localized PCa cases showed an epithelial phenotype, with overexpression of E-cadherin and underexpression of the mesenchymal markers. MiRNA-200 family members and miRNAs 203, 205, 183, 373, and 21 were overexpressed, while miRNAs 9, 495, 29b, and 1 were underexpressed. Low-expression levels of miRNAs 200b, 30a, and 1 were significantly associated with pathological stage. Lower expression of miR-200b was also associated with a Gleason score ≥ 8 and shorter biochemical recurrence-free survival. Furthermore, low-expression levels of miR-30a and high-expression levels of Vimentin and Twist1 were observed in the high-risk group. Compared with the primary tumor, the metastatic cell lines showed significantly higher expression levels of miR-183 and Twist1. In summary, miRNAs 200b, 30a, 1, and

  19. MicroRNA在丹酚酸B逆转肾小管上皮细胞转分化时的表达变化%Alteration of miRNA Expression in Salvianolic Acid B-reversed Epithelial-mesenchymal Transition of Renal Tubular Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    潘荣华; 芮国华; 柏小辉

    2012-01-01

    目的:研究丹酚酸B逆转转化生长因子(TGF)-β1诱导的肾小管上皮细胞-间充质细胞转分化时microRNA(miRNA)表达谱的变化.方法:将体外培养的人近端肾小管上皮细胞系(HK-2)细胞分为3组:①对照组:未加入丹酚酸B或者TGF-β1;②TGF-β1组:在细胞培养基中加入TGF-β1(浓度为5ng/mL);③丹酚酸B逆转组:在细胞培养基中先TGF-β1(浓度为5ng/mL),48h后HK-2细胞成功诱导EMT后形成的间充质细胞,再更换为TGF-β1(浓度为5ng/mL)和丹酚酸B(50μmol/L)继续培养72h,采用倒置相差显微镜观察细胞形态学变化;免疫细胞化学染色检测细胞E-cadherin的表达情况;基因芯片检测各组细胞miRNA的表达变化.结果:①丹酚酸B具有逆转HK-2细胞EMT的作用;②与正常HK-2细胞比较,TGF-β1组有25种miRNA的表达出现显著改变,其中15种表达下调2倍以上,10种表达上调2倍以上.③与TGF-β1组比较,丹酚酸B逆转组,有44种miRNA的表达出现显著改变,其中24种表达下调2倍以上,20种表达上调2倍以上.④有10种miRNA表达在两组同时出现显著改变,其中8种在TGF-β1组明显下调,而在丹酚酸B逆转组明显上调,1种在TGF-β1组明显上调,而在丹酚酸B逆转组则明显下调.结论:丹酚酸B具有阻止慢性肾脏疾病进行性发展的潜能,而这一作用与其能有效调控miRNA的表达有关.%Objective: To investigate the alteration of microRNA ( miRNA) expression patterns during the reversal effects of Salvianolic acid B ( Sal B)on epithelial-mesenchymal Transition ( EMT) induced by transforming growth factor-β1 ( TGF-β1 ) . Methods;In this study,Human kidney proximal tubular cell line(HK-2)was cultured in vitro as the proximal tubular cell model. Cells were divided into three groups as follows: control group, transforming growth factor-pl ( TGF-β1) group, Salvianolic acid B (Sal B) reversal group. Epithelial-mesenchymal transition (EMT) was induced by human TGF-β1 for 48h

  20. The Roles of Mitogen-Activated Protein Kinase Pathways in TGF-β-Induced Epithelial-Mesenchymal Transition.

    Science.gov (United States)

    Gui, Ting; Sun, Yujing; Shimokado, Aiko; Muragaki, Yasuteru

    2012-01-01

    The mitogen-activated protein kinase (MAPK) pathway allows cells to interpret external signals and respond appropriately, especially during the epithelial-mesenchymal transition (EMT). EMT is an important process during embryonic development, fibrosis, and tumor progression in which epithelial cells acquire mesenchymal, fibroblast-like properties and show reduced intercellular adhesion and increased motility. TGF-β signaling is the first pathway to be described as an inducer of EMT, and its relationship with the Smad family is already well characterized. Studies of four members of the MAPK family in different biological systems have shown that the MAPK and TGF-β signaling pathways interact with each other and have a synergistic effect on the secretion of additional growth factors and cytokines that in turn promote EMT. In this paper, we present background on the regulation and function of MAPKs and their cascades, highlight the mechanisms of MAPK crosstalk with TGF-β signaling, and discuss the roles of MAPKs in EMT.

  1. Prognostic significance of epithelial-mesenchymal transition proteins Twist and Foxc2 in phyllodes tumours of the breast.

    Science.gov (United States)

    Lim, Jeffrey Chun Tatt; Koh, Valerie Cui Yun; Tan, Jane Sie Yong; Tan, Wai Jin; Thike, Aye Aye; Tan, Puay Hoon

    2015-02-01

    Epithelial-mesenchymal transition (EMT), an important process during embryonic development, is reportedly exploited during tumour progression. Deregulation of EMT-related molecules has been shown in many malignancies, including breast carcinoma. We aim to investigate the clinical relevance and prognostic significance of EMT proteins, Twist and Foxc2, in breast phyllodes tumours (PTs). The study cohort comprised 271 PTs diagnosed from 2003 to 2010. Of these, 188 (69.4 %) were benign, 60 (22.1 %) borderline, and 23 (8.5 %) malignant. Immunohistochemistry for Twist and Foxc2 was performed on tissue microarray sections. Percentage of tumour cells stained was evaluated and correlated with clinicopathological parameters and clinical outcome. Twist and Foxc2 stromal nuclear expression was associated with tumour grade (P = 0.038 and 0.012). Foxc2 stromal nuclear expression was positively correlated with epithelial expression (P EMT-related molecules may be worthy of further investigation in PTs.

  2. PKD1 phosphorylation-dependent degradation of SNAIL by SCF-FBXO11 regulates epithelial-mesenchymal transition and metastasis

    Science.gov (United States)

    Zheng, Hanqiu; Shen, Minhong; Cha, Yin-Lian; Li, Wenyang; Wei, Yong; Blanco, Mario Andres; Ren, Guangwen; Zhou, Tianhua; Storz, Peter; Wang, Hui-Yun; Kang, Yibin

    2014-01-01

    SUMMARY Metastatic dissemination is often initiated by the reactivation of an embryonic development program referred to as epithelial-mesenchymal transition (EMT). The transcription factor SNAIL promotes EMT and elicits associated pathological characteristics, such as invasion, metastasis and stemness. To better understand the post-translational regulation of SNAIL, we performed a luciferase-based genome-wide E3 ligase siRNA library screen and identified SCF-FBXO11 as an important E3 which targets SNAIL for ubiquitylation and degradation. Furthermore, we discovered that SNAIL degradation by FBXO11 is dependent on Serine-11 phosphorylation of SNAIL by protein kinase D1 (PKD1). FBXO11 blocks SNAIL-induced EMT, tumor initiation and metastasis in multiple breast cancer models. These findings establish the PKD1-FBXO11-SNAIL axis as a mechanism of post-translational regulation of EMT and cancer metastasis. PMID:25203322

  3. S100A4: a common mediator of epithelial-mesenchymal transition, fibrosis and regeneration in diseases?

    DEFF Research Database (Denmark)

    Schneider, Mikael; Hansen, Jakob L; Sheikh, Søren P

    2008-01-01

    Multiple reports have focused on S100A4's role in cancer progression, specifically its ability to enhance metastasis. However, recent studies have linked S100A4 to several diseases besides cancer, including kidney fibrosis, cirrhosis, pulmonary disease, cardiac hypertrophy and fibrosis, arthritis...... and neuronal injuries. Common to all these diseases is the involvement of fibrotic and inflammatory processes, i.e. processes greatly dependent on tissue remodelling, cell motility and epithelial-mesenchymal transition. Therefore, the basic biological mechanisms behind S100A4's effects are emerging. S100A4...... belongs to the S100 family of proteins that contain two Ca(2+)-binding sites including a canonical EF-hand motif. S100A4 is involved in the regulation of a wide range of biological effects including cell motility, survival, differentiation and contractility. S100A4 has both intracellular and extracellular...

  4. S100A4: a common mediator of epithelial-mesenchymal transition, fibrosis and regeneration in diseases?

    DEFF Research Database (Denmark)

    Schneider, M.; Sheikh, S.P.; Hansen, Jakob Lerche

    2008-01-01

    Multiple reports have focused on S100A4's role in cancer progression, specifically its ability to enhance metastasis. However, recent studies have linked S100A4 to several diseases besides cancer, including kidney fibrosis, cirrhosis, pulmonary disease, cardiac hypertrophy and fibrosis, arthritis...... and neuronal injuries. Common to all these diseases is the involvement of fibrotic and inflammatory processes, i.e. processes greatly dependent on tissue remodelling, cell motility and epithelial-mesenchymal transition. Therefore, the basic biological mechanisms behind S100A4's effects are emerging. S100A4...... belongs to the S100 family of proteins that contain two Ca2+-binding sites including a canonical EF-hand motif. S100A4 is involved in the regulation of a wide range of biological effects including cell motility, survival, differentiation and contractility. S100A4 has both intracellular and extracellular...

  5. AURKA promotes cancer metastasis by regulating epithelial-mesenchymal transition and cancer stem cell properties in hepatocellular carcinoma.

    Science.gov (United States)

    Chen, Chenlin; Song, Guangyuan; Xiang, Jue; Zhang, Hongcheng; Zhao, Shaoyun; Zhan, Yinchu

    2017-04-29

    AURKA (aurora kinase A) has been confirmed as an oncogene in cancer development; however, its role and underlying mechanisms in the metastasis of hepatocellular carcinoma (HCC) remain unknown. In this study, We found that AURKA was up-regulated in HCC tissues and correlated with pathological stage and distant metastasis. Further found that AURKA was involved in the cancer metastases after radiation in HCC. While overexpression of AURKA induced epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) behaviors though PI3K/AKT pathway, silencing AURKA suppressed radiation-enhanced cell invasiveness of HCC. Taken together, our results suggested that AURKA contributed in metastasis of irradiated residul HCC though facilitating EMT and CSC properties, suggesting the potential clinical application of AURKA inhibitors in radiotherapy for patients with HCC.

  6. AMP-activated protein kinase inhibits TGF-β-, angiotensin II-, aldosterone-, high glucose-, and albumin-induced epithelial-mesenchymal transition.

    Science.gov (United States)

    Lee, Jang Han; Kim, Ji Hyun; Kim, Ja Seon; Chang, Jai Won; Kim, Soon Bae; Park, Jung Sik; Lee, Sang Koo

    2013-03-15

    The epithelial-mesenchymal transition (EMT) is a novel mechanism that promotes renal fibrosis. Transforming growth factor-β (TGF-β), angiotensin II, aldosterone, high glucose, and urinary albumin are well-known causes of EMT and renal fibrosis. We examined whether and how activation of AMP-activated protein kinase (AMPK) suppressed EMT induced by the above agents in tubular epithelial cells. All experiments were performed using HK-2 cells. Protein expression was measured by Western blot analysis. Intracellular reactive oxygen species (ROS) were analyzed by flow cytometry. Exposure of tubular cells to TGF-β (10 ng/ml), angiotensin II (1 μM), aldosterone (100 nM), high glucose (30 mM), and albumin (5 mg/ml) for 5 days induced EMT, as shown by upregulation of α-smooth muscle actin and downregulation of E-cadherin. ROS and NADPH oxidase 4 (Nox4) expression were increased, and antioxidants such as tiron and N-acetylcysteine inhibited EMT induction. Metformin (the best known clinical activator of AMPK) suppressed EMT induction through inhibition of ROS via induction of heme oxygenase-1 and endogenous antioxidant thioredoxin. An AMPK inhibitor (compound C) and AMPK small interfering RNA blocked the effect of metformin, and another AMPK activator [5-aminoimidazole-4-carboxamide-1β riboside (AICAR)] exerted the same effects as metformin. In conclusion, AMPK activation might be beneficial in attenuating the tubulointerstitial fibrosis induced by TGF-β, angiotensin II, aldosterone, high glucose, and urinary albumin.

  7. Caveolin-1 deficiency induces a MEK-ERK1/2-Snail-1-dependent epithelial-mesenchymal transition and fibrosis during peritoneal dialysis.

    Science.gov (United States)

    Strippoli, Raffaele; Loureiro, Jesús; Moreno, Vanessa; Benedicto, Ignacio; Pérez Lozano, María Luisa; Barreiro, Olga; Pellinen, Teijo; Minguet, Susana; Foronda, Miguel; Osteso, Maria Teresa; Calvo, Enrique; Vázquez, Jesús; López Cabrera, Manuel; del Pozo, Miguel Angel

    2015-01-01

    Peritoneal dialysis (PD) is a form of renal replacement therapy whose repeated use can alter dialytic function through induction of epithelial-mesenchymal transition (EMT) and fibrosis, eventually leading to PD discontinuation. The peritoneum from Cav1-/- mice showed increased EMT, thickness, and fibrosis. Exposure of Cav1-/- mice to PD fluids further increased peritoneal membrane thickness, altered permeability, and increased the number of FSP-1/cytokeratin-positive cells invading the sub-mesothelial stroma. High-throughput quantitative proteomics revealed increased abundance of collagens, FN, and laminin, as well as proteins related to TGF-β activity in matrices derived from Cav1-/- cells. Lack of Cav1 was associated with hyperactivation of a MEK-ERK1/2-Snail-1 pathway that regulated the Smad2-3/Smad1-5-8 balance. Pharmacological blockade of MEK rescued E-cadherin and ZO-1 inter-cellular junction localization, reduced fibrosis, and restored peritoneal function in Cav1-/- mice. Moreover, treatment of human PD-patient-derived MCs with drugs increasing Cav1 levels, as well as ectopic Cav1 expression, induced re-acquisition of epithelial features. This study demonstrates a pivotal role of Cav1 in the balance of epithelial versus mesenchymal state and suggests targets for the prevention of fibrosis during PD. © 2014 The Authors. Published under the terms of the CC BY 4.0 license.

  8. A Twist-Snail Axis Critical for TrkB-Induced Epithelial-Mesenchymal Transition-Like Transformation, Anoikis Resistance, and Metastasis▿ §

    OpenAIRE

    Smit, Marjon A.; Thomas R Geiger; Song, Ji-Ying; Gitelman, Inna; Peeper, Daniel S.

    2009-01-01

    In a genomewide anoikis suppression screen for metastasis genes, we previously identified the neurotrophic receptor tyrosine kinase TrkB. In mouse xenografts, activated TrkB caused highly invasive and metastatic tumors. Here, we describe that TrkB also induces a strong morphological transformation, resembling epithelial-mesenchymal transition (EMT). This required TrkB kinase activity, a functional mitogen-activated protein kinase pathway, suppression of E-cadherin, and induction of Twist, a t...

  9. Withaferin A Inhibits Experimental Epithelial-Mesenchymal Transition in MCF-10A Cells and Suppresses Vimentin Protein Level in Vivo in Breast Tumors

    OpenAIRE

    Lee, Joomin; Hahm, Eun-Ryeong; Marcus, Adam I.; Singh, Shivendra V.

    2013-01-01

    We have shown previously that withaferin A (WA), a bioactive component of the medicinal plant Withania somnifera, inhibits growth of cultured and xenografted human breast cancer cells and prevents breast cancer development and pulmonary metastasis incidence in a transgenic mouse model. The present study was undertaken to determine if the anticancer effect of WA involved inhibition of epithelial-mesenchymal transition (EMT). Experimental EMT induced by exposure of MCF-10A cells to tumor necros...

  10. SIRT1 promotes epithelial-mesenchymal transition and metastasis in colorectal cancer by regulating Fra-1 expression.

    Science.gov (United States)

    Cheng, Feifei; Su, Li; Yao, Chao; Liu, Limei; Shen, Junjie; Liu, Chungang; Chen, Xuejiao; Luo, Yongli; Jiang, Lupin; Shan, Juanjuan; Chen, Jun; Zhu, Wei; Shao, Jimin; Qian, Cheng

    2016-06-01

    Understanding molecular mechanisms of colorectal cancer (CRC) metastasis is urgently required for targeted therapy and prognosis of metastatic CRC. In this study, we explored potential effects of silent mating type information regulation 2 homolog 1 (SIRT1) on CRC metastasis. Our data showed that ectopic expression of SIRT1 markedly increased the migration and invasion of CRC cells. In contrast, silencing SIRT1 repressed this behavior in aggressive CRC cells. Tumor xenograft experiments revealed that knockdown of SIRT1 impaired CRC metastasis in vivo. Silencing SIRT1 in CRC cells induced mesenchymal-epithelial transition (MET), which is the reverse process of epithelial-mesenchymal transition (EMT) and characterized by a gain of epithelial and loss of mesenchymal markers. We provided a mechanistic insight toward regulation of Fra-1 by SIRT1 and demonstrated a direct link between the SIRT1-Fra-1 axis and EMT. Moreover, SIRT1 expression correlated positively with Fra-1 expression, metastasis and overall survival in patients with CRC. Taken together, our data provide a novel mechanistic role of SIRT1 in CRC metastasis, suggesting that SIRT1 may serve as a potential therapeutic target for metastatic CRC.

  11. Differential expression of the epithelial mesenchymal transition factors Snail, Slug, Twist, TGF-β, and E-cadherin in ameloblastoma.

    Science.gov (United States)

    Kurioka, Kagami; Wato, Masahiro; Iseki, Tomio; Tanaka, Akio; Morita, Shosuke

    2017-06-01

    Epithelial mesenchymal transition (EMT), the transition of epithelial cells into motile mesenchymal cells, plays an important role in embryogenesis, cancer invasion, and metastasis. Ameloblastomas are common epithelial odontogenic tumors, occurring exclusively in the mandible with locally invasive growth. Thirty-seven ameloblastoma cases were evaluated for the involvement of EMT by immunohistochemical staining and western blotting using antibodies against Slug, Snail, Twist, TGF-β, and E-cadherin. Double immunostaining was also performed. Slug and TGF-β were expressed in the nuclei of peripheral and stellate reticulum cells of ameloblastoma nests. Twenty cases of Snail, 36 of Slug, 8 of Twist, and 19 of TGF-β showed strong expression in tumor cells in follicular and plexiform patterns. Expression of Slug and TGF-β increased in regions where the expression of E-cadherin was reduced. EMT was found to be associated with the local invasive growth of ameloblastoma. These data suggest that reduced expression of E-cadherin and over-expression of Slug, Snail, and TGF-β induce EMT. Given that ameloblastomas are characterized by local invasiveness, EMT might be related to their development. Thus, strong expression of Slug and TGF-β and reduced expression of E-cadherin might be related to the local invasiveness of ameloblastoma.

  12. Human cancer cells express Slug-based epithelial-mesenchymal transition gene expression signature obtained in vivo

    Directory of Open Access Journals (Sweden)

    Anastassiou Dimitris

    2011-12-01

    Full Text Available Abstract Background The biological mechanisms underlying cancer cell motility and invasiveness remain unclear, although it has been hypothesized that they involve some type of epithelial-mesenchymal transition (EMT. Methods We used xenograft models of human cancer cells in immunocompromised mice, profiling the harvested tumors separately with species-specific probes and computationally analyzing the results. Results Here we show that human cancer cells express in vivo a precise multi-cancer invasion-associated gene expression signature that prominently includes many EMT markers, among them the transcription factor Slug, fibronectin, and α-SMA. We found that human, but not mouse, cells express the signature and Slug is the only upregulated EMT-inducing transcription factor. The signature is also present in samples from many publicly available cancer gene expression datasets, suggesting that it is produced by the cancer cells themselves in multiple cancer types, including nonepithelial cancers such as neuroblastoma. Furthermore, we found that the presence of the signature in human xenografted cells was associated with a downregulation of adipocyte markers in the mouse tissue adjacent to the invasive tumor, suggesting that the signature is triggered by contextual microenvironmental interactions when the cancer cells encounter adipocytes, as previously reported. Conclusions The known, precise and consistent gene composition of this cancer mesenchymal transition signature, particularly when combined with simultaneous analysis of the adjacent microenvironment, provides unique opportunities for shedding light on the underlying mechanisms of cancer invasiveness as well as identifying potential diagnostic markers and targets for metastasis-inhibiting therapeutics.

  13. Expression of transcription factors Twist and Snail in cervical intraepithelial neoplasia tissues and their relationship with epithelial-mesenchymal transition and cell proliferation

    Institute of Scientific and Technical Information of China (English)

    Hai-Dan Fu

    2016-01-01

    Objective:To study the expression of transcription factors Twist and Snail in cervical intraepithelial neoplasia tissues and their relationship with epithelial-mesenchymal transition and cell proliferation.Methods: cervical intraepithelial neoplasia tissues (n=67) and normal cervical tissues (n=85) were collected, and the contents of Twist and Snail as well as epithelial-mesenchymal transition-related molecules and proliferation-related molecules in the tissues were detected.Results:Twist and Snail contents in CIN cervical tissues were lower than those in normal cervical tissues; Twist, Snail, PI3K, Akt, STAT3, Vimentin, N-cadherin, Prdx4, EZH2 and STOML-2 contents in CIN cervical tissues were higher than those in normal cervical tissues, and E-cadherin content was significantly lower than that in normal cervical tissues; E-cadherin content in CIN tissues with high expression of Twist and Snail was significantly lower than that in CIN tissues with low expression of Twist and Snail, and Vimentin, N-cadherin, Prdx4, EZH2 and STOML-2 contents were higher than those in CIN tissues with low expression of Twist and Snail.Conclusions:Transcription factors Twist and Snail expression increase and downstream signaling pathway function is enhanced in cervical intraepithelial neoplasia tissues; Twist and Snail can regulate epithelial-mesenchymal transition and cell proliferation.

  14. The tumor microenvironment: An irreplaceable element of tumor budding and epithelial-mesenchymal transition-mediated cancer metastasis.

    Science.gov (United States)

    Li, Hui; Xu, Fangying; Li, Si; Zhong, Anjing; Meng, Xianwen; Lai, Maode

    2016-07-03

    Tumor budding occurs at the invasive front of cancer; the tumor cells involved have metastatic and stemness features, indicating a poor prognosis. Tumor budding is partly responsible for cancer metastasis, and its initiation is based on the epithelial-mesenchymal transition (EMT) process. The EMT process involves the conversion of epithelial cells into migratory and invasive cells, and is a profound event in tumorigenesis. The EMT, associated with the formation of cancer stem cells (CSCs) and resistance to therapy, results from a combination of gene mutation, epigenetic regulation, and microenvironmental control. Tumor budding can be taken to represent the EMT in vivo. The EMT process is under the influence of the tumor microenvironment as well as tumor cells themselves. Here, we demonstrate that the tumor microenvironment dominates EMT development and impacts cancer metastasis, as well as promotes CSC formation and mediates drug resistance. In this review, we mainly discuss components of the microenvironment, such as the extracellular matrix (ECM), inflammatory cytokines, metabolic products, and hypoxia, that are involved in and impact on the acquisition of tumor-cell motility and dissemination, the EMT, metastatic tumor-cell formation, tumor budding and CSCs, and cancer metastasis, including subsequent chemo-resistance. From our point of view, the tumor microenvironment now constitutes a promising target for cancer therapy.

  15. Blocking TGF-β expression inhibits silica particle-induced epithelial-mesenchymal transition in human lung epithelial cells.

    Science.gov (United States)

    Rong, Yi; Shen, Yan; Zhang, Zhihong; Cui, Xiuqing; Xiao, Lili; Liu, Yuewei; Luo, Xin; Chen, Weihong

    2015-11-01

    The main characteristic of silicosis is irreversible fibrosis. Certain studies have shown that epithelial-mesenchymal transition (EMT) regulated by transforming growth factor-β (TGF-β) is involved in fibrosis. Thus, we suggest that TGF-β regulated EMT may play an important role in silicosis. In this study, we determined the expression of TGF-β-Smad2/3, EMT- and ECM-related markers in lung epithelial cells treated with silica particle by RT-PCR, western-blot and ELISA. In order to explore the role of TGF-β, we used TGF-β inhibitor in the cell model. We found that the cells lost the expression of epithelial phenotypic markers and acquired increased expression of mesenchymal cells markers with ECM deposition after treatment with silica particle. Moreover, the changes of EMT-related event was restricted in response to TGF-β inhibitor. These findings suggest that EMT is essentially involved in the pathogenesis of fibrosis induced by silica particles and down-regulating the TGF-β expression can inhibit the process of EMT.

  16. Transcriptional silencing of ETS-1 abrogates epithelial-mesenchymal transition resulting in reduced motility of pancreatic cancer cells.

    Science.gov (United States)

    Li, Chunyan; Wang, Zhonghan; Chen, Yan; Zhou, Min; Zhang, Haijun; Chen, Rong; Shi, Fangfang; Wang, Cailian; Rui, Zongdao

    2015-02-01

    v-ets erythroblastosis virus E26 oncogene homolog 1 (ETS-1) plays crucial roles in a spectrum of malignancies. ETS-1 has gained attention in cancer research for its importance in cell migration, invasion and proliferation. In the present study, we focused on the effect of ETS-1 on epithelial-mesenchymal transition (EMT), which is characterized by reduced E-cadherin expression and increased N-cadherin expression. We found that ETS-1 mRNA expression was positively correlated with N-cadherin and negatively correlated with E-cadherin mRNA expression in five pancreatic cancer cell lines. To elucidate the functionality of ETS-1 on EMT in pancreatic cancer cells, we constructed a green fluorescent protein (GFP)-expressing plasmid carrying ETS-1 short hairpin RNA (shRNA), and transfected Panc-1 cells with the plasmid. We detected reduced N-cadherin and vascular endothelial growth factor yet higher E-cadherin expression in the ETS-1-silenced cells compared with the control group. In addition, we observed reduced cell migration and increased adhesion in these cells. Our data showed that ETS-1 actively functioned as a regulator of EMT in Panc-1 cells, and provide additional evidence supporting a fundamental role for ETS-1 in metastatic pancreatic cancer cells. These results suggest that analysis of ETS-1 expression levels may provide an avenue for evaluating prognosis in pancreatic cancer.

  17. Hyperthermia inhibits hypoxia-induced epithelial-mesenchymal transition in HepG2 hepatocellular carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Guang-Jin Yuan; Qian-wen Li; Shun-Lin Shan; Wu-Ming Wang; Sen Jiang; Xi-Ming Xu

    2012-01-01

    AIM:TO investigate the effect of hyperthermia on hypoxia-induced epithelial-mesenchymal transition (EMT)in HepG2 hepatocellular carcinoma (HCC) cells,and its mechanism.METHODS:Cells were treated with hyperthermia at 43 ℃ for 0.5 h,followed by incubation under hypoxic or normoxic conditions for 72 h.Cell morphology was observed.Expressions of E-cadherin and vimentin were determined by immunofluorescence assay or Western blot.The protein and mRNA expressions of Snail were also determined by Western blot and reverse transcription-polymerase chain reaction.Cell migratory capacity was evaluated.RESULTS:Hypoxia induced EMT in HepG2 cells,which was evidenced by morphological,molecular and functional changes,including the formation of a spindle shape and the loss of cell contact.The expression of E-cadherin was decreased but the expression of vimentin was increased; also,the migratory capability was increased by 2.2 ± 0.20-fold as compared with normoxia.However,those effects were inhibited by hyperthermia pretreatment.Furthermore,protein synthesis and mRNA expression of Snail in the cells were enhanced by hypoxia as compared with normoxia,and also significantly inhibited by hyperthermia pretreatment.CONCLUSION:Hyperthermia may inhibit hypoxiainduced EMT in HepG2 HCC cells,and the mechanism may involve inhibition of induced expression of Snail.

  18. Expression of epithelial-mesenchymal transition-related genes increases with copy number in multiple cancer types.

    Science.gov (United States)

    Zhao, Min; Liu, Yining; Qu, Hong

    2016-04-26

    Epithelial-mesenchymal transition (EMT) is a cellular process through which epithelial cells transform into mesenchymal cells. EMT-implicated genes initiate and promote cancer metastasis because mesenchymal cells have greater invasive and migration capacities than epithelial cells. In this pan-cancer analysis, we explored the relationship between gene expression changes and copy number variations (CNVs) for EMT-implicated genes. Based on curated 377 EMT-implicated genes from the literature, we identified 212 EMT-implicated genes associated with more frequent copy number gains (CNGs) than copy number losses (CNLs) using data from The Cancer Genome Atlas (TCGA). Then by correlating these CNV data with TCGA gene expression data, we identified 71 EMT-implicated genes with concordant CNGs and gene up-regulation in 20 or more tumor samples. Of those, 14 exhibited such concordance in over 110 tumor samples. These 14 genes were predominantly apoptosis regulators, which may implies that apoptosis is critical during EMT. Moreover, the 71 genes with concordant CNG and up-regulation were largely involved in cellular functions such as phosphorylation cascade signaling. This is the first observation of concordance between CNG and up-regulation of specific genes in hundreds of samples, which may indicate that somatic CNGs activate gene expression by increasing the gene dosage.

  19. CCR7 pathway induces epithelial-mesenchymal transition through up-regulation of Snail signaling in gastric cancer.

    Science.gov (United States)

    Zhang, Jianping; Zhou, Yunzhe; Yang, Yonggang

    2015-02-01

    The chemokine receptor 7 (CCR7) and Snail signaling have been linked to various types of cancers. The associations between these signalings and the epithelial-mesenchymal transition (EMT) are not clear in gastric cancer. Here, the expression of CCR7 and Snail was detected in gastric cancer by immunohistochemistry and Western blot. Meanwhile, gastric cancer cells were subjected to CCL19, si-control, and si-Snail treatment. Cell cycle, migration, and invasion were also analyzed. The expression patterns of CCR7 and Snail were similar in either gastric cancer tissues or cells. The increased expression of CCR7 was closely associated with the increased Snail expression, which both were closely correlated with metastasis, stage and differentiation, and poor prognosis. The increased p-ERK, p-AKT, Snail, and MMP9 expression and the decreased E-cadherin were confirmed in MGC803 cells in a dose-dependent manner in response to CCL19 treatment. However, the blockade of Snail abrogated the up-regulation of MMP9 and down-regulation of E-cadherin. CCR7-induced ERK and PI3K pathway regulated Snail signaling. Besides si-Snail treatment led to MGC803 cell cycle arrest and affected the migration and invasion. In conclusion, our study suggested that CCR7 promotes Snail expression to induce the EMT, resulting in cell cycle progression, migration, and invasion in gastric cancer. CCR7-Snail pathway provided more potential regimens for cancer therapy.

  20. Inhibition of RAB1A suppresses epithelial-mesenchymal transition and proliferation of triple-negative breast cancer cells.

    Science.gov (United States)

    Xu, Hui; Qian, Mingping; Zhao, Bingkun; Wu, Chenyang; Maskey, Niraj; Song, Hongming; Li, Dengfeng; Song, Jialu; Hua, Kaiyao; Fang, Lin

    2017-03-01

    RAB1A acts as an oncogene in various cancers, and emerging evidence has verified that RAB1A is an mTORC1 activator in hepatocellular and colorectal cancer, but the role of RAB1A in breast cancer remains unclear. In this investigation, RAB1A siRNA was successfully transfected in MDA-MB-231 and BT-549 human triple-negative breast cancer cells, and verified by real‑time quantitative polymerase chain reaction and western blotting. Then, MTT cell proliferation, colony formation, cell invasion and wound healing assays were performed to characterize the function of RAB1A in the breast cancer cell lines. Downregulation of RAB1A inhibited cellular growth, cell migration, cell invasion and cell epithelial-mesenchymal transition. Furthermore, compared with NC siRNA transfected cells, RAB1A siRNA transfected breast cancer cells inhibited the phosphorylation of S6K1, the effector molecular of mTORC1. Collectively, our data suggested that RAB1A acts as an oncogene by regulating cellular proliferation, growth, invasion and metastasis via activation of mTORC1 pathway in triple-negative breast cancer.

  1. The Roles of Mitogen-Activated Protein Kinase Pathways in TGF-β-Induced Epithelial-Mesenchymal Transition

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    Ting Gui

    2012-01-01

    Full Text Available The mitogen-activated protein kinase (MAPK pathway allows cells to interpret external signals and respond appropriately, especially during the epithelial-mesenchymal transition (EMT. EMT is an important process during embryonic development, fibrosis, and tumor progression in which epithelial cells acquire mesenchymal, fibroblast-like properties and show reduced intercellular adhesion and increased motility. TGF-β signaling is the first pathway to be described as an inducer of EMT, and its relationship with the Smad family is already well characterized. Studies of four members of the MAPK family in different biological systems have shown that the MAPK and TGF-β signaling pathways interact with each other and have a synergistic effect on the secretion of additional growth factors and cytokines that in turn promote EMT. In this paper, we present background on the regulation and function of MAPKs and their cascades, highlight the mechanisms of MAPK crosstalk with TGF-β signaling, and discuss the roles of MAPKs in EMT.

  2. Vimentin contributes to epithelial-mesenchymal transition cancer cell mechanics by mediating cytoskeletal organization and focal adhesion maturation.

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    Liu, Ching-Yi; Lin, Hsi-Hui; Tang, Ming-Jer; Wang, Yang-Kao

    2015-06-30

    Modulations of cytoskeletal organization and focal adhesion turnover correlate to tumorigenesis and epithelial-mesenchymal transition (EMT), the latter process accompanied by the loss of epithelial markers and the gain of mesenchymal markers (e.g., vimentin). Clinical microarray results demonstrated that increased levels of vimentin mRNA after chemotherapy correlated to a poor prognosis of breast cancer patients. We hypothesized that vimentin mediated the reorganization of cytoskeletons to maintain the mechanical integrity in EMT cancer cells. By using knockdown strategy, the results showed reduced cell proliferation, impaired wound healing, loss of directional migration, and increased large membrane extension in MDA-MB 231 cells. Vimentin depletion also induced reorganization of cytoskeletons and reduced focal adhesions, which resulted in impaired mechanical strength because of reduced cell stiffness and contractile force. In addition, overexpressing vimentin in MCF7 cells increased cell stiffness, elevated cell motility and directional migration, reoriented microtubule polarity, and increased EMT phenotypes due to the increased β1-integrin and the loss of junction protein E-cadherin. The EMT-related transcription factor slug was also mediated by vimentin. The current study demonstrated that vimentin serves as a regulator to maintain intracellular mechanical homeostasis by mediating cytoskeleton architecture and the balance of cell force generation in EMT cancer cells.

  3. Matrix stiffness drives epithelial-mesenchymal transition and tumour metastasis through a TWIST1-G3BP2 mechanotransduction pathway.

    Science.gov (United States)

    Wei, Spencer C; Fattet, Laurent; Tsai, Jeff H; Guo, Yurong; Pai, Vincent H; Majeski, Hannah E; Chen, Albert C; Sah, Robert L; Taylor, Susan S; Engler, Adam J; Yang, Jing

    2015-05-01

    Matrix stiffness potently regulates cellular behaviour in various biological contexts. In breast tumours, the presence of dense clusters of collagen fibrils indicates increased matrix stiffness and correlates with poor survival. It is unclear how mechanical inputs are transduced into transcriptional outputs to drive tumour progression. Here we report that TWIST1 is an essential mechanomediator that promotes epithelial-mesenchymal transition (EMT) in response to increasing matrix stiffness. High matrix stiffness promotes nuclear translocation of TWIST1 by releasing TWIST1 from its cytoplasmic binding partner G3BP2. Loss of G3BP2 leads to constitutive TWIST1 nuclear localization and synergizes with increasing matrix stiffness to induce EMT and promote tumour invasion and metastasis. In human breast tumours, collagen fibre alignment, a marker of increasing matrix stiffness, and reduced expression of G3BP2 together predict poor survival. Our findings reveal a TWIST1-G3BP2 mechanotransduction pathway that responds to biomechanical signals from the tumour microenvironment to drive EMT, invasion and metastasis.

  4. Siva1 suppresses epithelial-mesenchymal transition and metastasis of tumor cells by inhibiting stathmin and stabilizing microtubules.

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    Li, Nan; Jiang, Peng; Du, Wenjing; Wu, Zhengsheng; Li, Cong; Qiao, Mengran; Yang, Xiaolu; Wu, Mian

    2011-08-02

    Epithelial-mesenchymal transition (EMT) enables epithelial cells to acquire motility and invasiveness that are characteristic of mesenchymal cells. It plays an important role in development and tumor cell metastasis. However, the mechanisms of EMT and their dysfunction in cancer cells are still not well understood. Here we report that Siva1 interacts with stathmin, a microtubule destabilizer. Siva1 inhibits stathmin's activity directly as well as indirectly through Ca(2+)/calmodulin-dependent protein kinase II-mediated phosphorylation of stathmin at Ser16. Via the inhibition of stathmin, Siva1 enhances the formation of microtubules and impedes focal adhesion assembly, cell migration, and EMT. Low levels of Siva1 and Ser16-phosphorylated stathmin correlate with high metastatic states of human breast cancer cells. In mouse models, knockdown of Siva1 promotes cancer dissemination, whereas overexpression of Siva1 inhibits it. These results suggest that microtubule dynamics are critical for EMT. Furthermore, they reveal an important role for Siva1 in suppressing cell migration and EMT and indicate that down-regulation of Siva1 may contribute to tumor cell metastasis.

  5. A cell-based small molecule screening method for identifying inhibitors of epithelial-mesenchymal transition in carcinoma.

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    Kian-Ngiap Chua

    Full Text Available Epithelial Mesenchymal Transition (EMT is a crucial mechanism for carcinoma progression, as it provides routes for in situ carcinoma cells to dissociate and become motile, leading to localized invasion and metastatic spread. Targeting EMT therefore represents an important therapeutic strategy for cancer treatment. The discovery of oncogene addiction in sustaining tumor growth has led to the rapid development of targeted therapeutics. Whilst initially optimized as anti-proliferative agents, it is likely that some of these compounds may inhibit EMT initiation or sustenance, since EMT is also modulated by similar signaling pathways that these compounds were designed to target. We have developed a novel screening assay that can lead to the identification of compounds that can inhibit EMT initiated by growth factor signaling. This assay is designed as a high-content screening assay where both cell growth and cell migration can be analyzed simultaneously via time-course imaging in multi-well plates. Using this assay, we have validated several compounds as viable EMT inhibitors. In particular, we have identified compounds targeting ALK5, MEK, and SRC as potent inhibitors that can interfere with EGF, HGF, and IGF-1 induced EMT signaling. Overall, this EMT screening method provides a foundation for improving the therapeutic value of recently developed compounds in advanced stage carcinoma.

  6. Palbociclib inhibits epithelial-mesenchymal transition and metastasis in breast cancer via c-Jun/COX-2 signaling pathway.

    Science.gov (United States)

    Qin, Ge; Xu, Fei; Qin, Tao; Zheng, Qiufan; Shi, Dingbo; Xia, Wen; Tian, Yun; Tang, Yanlai; Wang, Jingshu; Xiao, Xiangshen; Deng, Wuguo; Wang, Shusen

    2015-12-08

    Palbociclib, a highly selective CDK4/6 inhibitor, has been shown to be a novel anti-tumor agent that suppresses breast cancer cell proliferation. However, its anti-metastasis activity remains controversial. In the present study, we evaluated whether palbociclib prevented breast cancer cell metastasis and revealed its regulatory mechanism. We found that palbociclib inhibited migration and invasion in the breast cancer cells MDA-MB-231 and T47D. The epithelial-mesenchymal transition (EMT) markers, vimentin and Snail, were down-regulated with palbociclib treatment. Moreover, we revealed that this inhibition was mediated by the c-Jun/COX-2 pathway. COX-2 was decreased after palbociclib treatment. The production of PGE2 was also reduced along with COX-2. Additionally, our data showed that c-Jun, a crucial transcriptional regulator of COX-2, was down-regulated by palbociclib. We found that palbociclib weakened the COX-2 promoter binding activity of c-Jun and prevented its translocation from the cytoplasm to cell nuclei. Bioluminescence imaging and tail intravenous injection were used to evaluate the anti-metastasis effect of palbociclib in vivo. The data demonstrated that palbociclib reduced breast cancer metastasis to the lung. These results therefore demonstrated that the anti-metastasis activity of palbociclib is mediated via the c-Jun/COX-2 signaling pathway by inhibiting EMT in breast cancer cells.

  7. SPRY4 Intronic Transcript 1 Promotes Epithelial-Mesenchymal Transition Through Association with Snail1 in Osteosarcoma.

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    Ru, Neng; Liang, Jie; Zhang, Fan; Wu, Weifei; Wang, Feifan; Liu, Xinzong; Du, Yuanli

    2016-06-01

    Osteosarcoma is an aggressive tumor and the most common malignancy of the skeleton. Due to pulmonary metastasis, the 5-year survival rate is still unsatisfactory. It has been reported that SPRY4 intronic transcript 1 (SPRY4-IT1) promotes cell growth, invasion, and inhibits apoptosis in several cancers. However, the role of SPRY4-IT1 in osteosarcoma remains unclear. In the present study, we investigated the role of SPRY4-IT1 in osteosarcoma cells. Loss- and gain-of-function assays demonstrated that SPRY4-IT1 promoted cell proliferation, migration, and invasion in osteosarcoma. Moreover, SPRY4-IT1 induced epithelial-mesenchymal transition phenotype in osteosarcoma cells. Subsequent investigations revealed that SPRY4-IT1 promoted migration and invasion through association with Snail1 and regulating its stability. Based on these findings, the SPRY4-IT1/Snail1/E-cadherin pathway may play a crucial role in promoting osteosarcoma metastasis. Thus, SPRY4-IT1 may be a potential target for new therapies of osteosarcoma.

  8. Up-Regulated FASN Expression Promotes Transcoelomic Metastasis of Ovarian Cancer Cell through Epithelial-Mesenchymal Transition

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    Li Jiang

    2014-06-01

    Full Text Available Fatty acid synthase (FASN, responsible for the de novo synthesis of fatty acids, has been shown to act as an oncogene in various human cancers. However, the mechanisms by which FASN favors the progression of ovarian carcinoma remain unknown. In this study, we evaluated FASN expression in ovarian cancer and investigated how FASN regulates the aggressiveness of ovarian cancer cells. Our results show that increased FASN is associated with the peritoneal metastasis of ovarian cancers. Over-expression of FASN results in a significant increase of tumor burden in peritoneal dissemination, accompanied by augment in cellular colony formation and metastatic ability. Correspondingly, FASN knockdown using RNA interference in ovarian cancer cells inhibits the migration in vitro and experimental peritoneal dissemination in vivo. Mechanistic studies reveal that FASN promotes Epithelial-mesenchymal Transition (EMT via a transcriptional regulation of E-cadherin and N-cadherin, which is also confirmed by luciferase promoter activity analysis. Taken together, our work demonstrates that FASN promotes the peritoneal dissemination of ovarian cancer cells, at least in part through the induction of EMT. These findings suggest that FASN plays a critical role in the peritoneal metastasis of ovarian cancer. Targeting de novo lipogenesis may have a therapeutic potential for advanced ovarian cancer.

  9. The expression of SIRT1 regulates the metastaticplasticity of chondrosarcoma cells by inducing epithelial-mesenchymal transition

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    Feng, Helin; Wang, Jin; Xu, Jianfa; Xie, Congcong; Gao, Fulu; Li, Zhiyong

    2017-01-01

    SIRT1 belongs to the mammalian sirtuin family and plays an important role in deacetylating histone and nonhistone proteins. It is reported that SIRT1 is associated with tumor metastasis in several kinds of tumors. However, the effect of SIRT1 on the metastasis of chondrosarcoma cells is still unknown. In this study, we demonstrated that up and down-regulation of SIRT1 expression could significantly change the invasive and metastatic potential in chondrosarcoma cell line. Besides that, the result from the nude mice confirmed the effect of SIRT1 on metastasis of chondrosarcoma cells. Furthermore, we also found that SIRT1 effectively enhanced the metastasis by inducing epithelial-mesenchymal transition (EMT) in chondrosarcoma cells. Inhibition the expression of SIRT1 could block the incidence of metastasis and EMT in chondrosarcoma cells. In addition, we also observed that SIRT1 could enhance the expression of Twist which is a key transcriptional factor of EMT. A clinicopathological analysis showed that SIRT1 expression was significantly correlated with the poor prognosis of pelvis chondrosarcoma. Kaplan-Meier survival curves revealed that positive SIRT1 expression was associated with poor prognosis in patients with pelvis chondrosarcoma. Taken together, these results indicate that SIRT1 may promote the metastasis of chondrosarcoma by inducing EMT and can be a potential molecular target for chondrosarcoma therapy. PMID:28112277

  10. Change in Cell Shape Is Required for Matrix Metalloproteinase-Induced Epithelial-Mesenchymal Transition of Mammary Epithelial Cells

    Science.gov (United States)

    Nelson, Celeste M.; Khauv, Davitte; Bissell, Mina J.; Radisky, Derek C.

    2010-01-01

    Cell morphology dictates response to a wide variety of stimuli, controlling cell metabolism, differentiation, proliferation, and death. Epithelial-mesenchymal transition (EMT) is a developmental process in which epithelial cells acquire migratory characteristics, and in the process convert from a “cuboidal” epithelial structure into an elongated mesenchymal shape. We had shown previously that matrix metalloproteinase-3 (MMP3) can stimulate EMT of cultured mouse mammary epithelial cells through a process that involves increased expression of Rac1b, a protein that stimulates alterations in cytoskeletal structure. We show here that cells treated with MMP-3 or induced to express Rac1b spread to cover a larger surface, and that this induction of cell spreading is a requirement of MMP-3/Rac1b-induced EMT. We find that limiting cell spreading, either by increasing cell density or by culturing cells on precisely defined micropatterned substrata, blocks expression of characteristic markers of EMT in cells treated with MMP-3. These effects are not caused by general disruptions in cell signaling pathways, as TGF-β-induced EMT is not affected by similar limitations on cell spreading. Our data reveal a previously unanticipated cell shape-dependent mechanism that controls this key phenotypic alteration and provide insight into the distinct mechanisms activated by different EMT-inducing agents. PMID:18506791

  11. Change in cell shape is required for matrix metalloproteinase-induced epithelial-mesenchymal transition of mammary epithelial cells

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    Nelson, Celeste M.; Khauv, Davitte; Bissell, Mina J.; Radisky, Derek C.

    2008-06-26

    Cell morphology dictates response to a wide variety of stimuli, controlling cell metabolism, differentiation, proliferation, and death. Epithelial-mesenchymal transition (EMT) is a developmental process in which epithelial cells acquire migratory characteristics, and in the process convert from a 'cuboidal' epithelial structure into an elongated mesenchymal shape. We had shown previously that matrix metalloproteinase-3 (MMP3) can stimulate EMT of cultured mouse mammary epithelial cells through a process that involves increased expression of Rac1b, a protein that stimulates alterations in cytoskeletal structure. We show here that cells treated with MMP-3 or induced to express Rac1b spread to cover a larger surface, and that this induction of cell spreading is a requirement of MMP-3/Rac1b-induced EMT. We find that limiting cell spreading, either by increasing cell density or by culturing cells on precisely defined micropatterned substrata, blocks expression of characteristic markers of EMT in cells treated with MMP-3. These effects are not caused by general disruptions in cell signaling pathways, as TGF-{beta}-induced EMT is not affected by similar limitations on cell spreading. Our data reveal a previously unanticipated cell shape-dependent mechanism that controls this key phenotypic alteration and provide insight into the distinct mechanisms activated by different EMT-inducing agents.

  12. NANOG regulates epithelial-mesenchymal transition and chemoresistance through activation of the STAT3 pathway in epithelial ovarian cancer.

    Science.gov (United States)

    Liu, Suqing; Sun, Jing; Cai, Bin; Xi, Xiaowei; Yang, Liu; Zhang, Zhenbo; Feng, Youji; Sun, Yunyan

    2016-07-01

    NANOG is a key transcription factor that is overexpressed and plays an important role in various cancers. Its overexpression is associated with highly tumorigenic, drug-resistant, and poor prognosis. However, the underlying mechanism of action of NANOG in ovarian cancer remains unclear. Epithelial-mesenchymal transition (EMT), which is a critical process in cancer invasion and metastasis, is also associated with drug resistance. We determined whether NANOG is associated with EMT and chemoresistance in epithelial ovarian cancer cells. NANOG expression was increased in epithelial ovarian cancer cells (HEY and SKOV3) compared with normal epithelial ovarian cells (Moody). Low expression of NANOG increased the expression of E-cadherin and decreased the expression of vimentin, β-catenin, and Snail. Furthermore, the cell migration and invasion abilities were decreased. The multidrug resistance genes MDR-1 and GST-π were also downregulated when NANOG was lowly expressed. The cells that were transfected with the si-NANOG plasmid were more sensitive to cisplatin compared with the cells that were transfected with empty vector. The data demonstrated that Stat3 was correlated with NANOG-mediated EMT and drug resistance. The silencing of Stat3 expression abrogated NANOG-mediated EMT changes and increased the sensitivity of the cells to chemotherapy. These results suggest that NANOG mediates EMT and drug resistance through activation of the Stat3 pathway in epithelial ovarian cancer.

  13. Wheatgrass extract inhibits hypoxia-inducible factor-1-mediated epithelial-mesenchymal transition in A549 cells

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    Do, Nam Yong; Shin, Hyun-Jae

    2017-01-01

    BACKGROUND/OBJECTIVES Epithelial-mesenchymal transition (EMT) is involved in not only cancer development and metastasis but also non-cancerous conditions. Hypoxia is one of the proposed critical factors contributing to formation of chronic rhinosinusitis or nasal polyposis. Wheatgrass (Triticum aestivum) has antioxidant, anti-aging, and anti-inflammatory effects. In this study, we analyzed whether wheatgrass has an inhibitory effect on the EMT process in airway epithelial cells. MATERIALS/METHODS A549 human lung adenocarcinoma cells were incubated in hypoxic conditions (CO2 5%/O2 1%) for 24 h in the presence of different concentrations of wheatgrass extract (50, 75, 100, and 150 µg/mL) and changes in expression of epithelial or mesenchymal markers were evaluated by immunoblotting and immunofluorescence. Accordingly, associated EMT-related transcriptional factors, Snail and Smad, were also evaluated. RESULTS Hypoxia increased expression of N-cadherin and reduced expression of E-cadherin. Mechanistically, E-cadherin levels were recovered during hypoxia by silencing hypoxia inducible factor (HIF)-1α or administering wheatgrass extract. Wheatgrass inhibited the hypoxia-mediated EMT by reducing the expression of phosphorylated Smad3 (pSmad3) and Snail. It suppressed the hypoxia-mediated EMT processes of airway epithelial cells via HIF-1α and the pSmad3 signaling pathway. CONCLUSION These results suggest that wheatgrass has potential as a therapeutic or supplementary agent for HIF-1-related diseases.

  14. Epithelial-mesenchymal transition associates with maintenance of stemness in spheroid-derived stem-like colon cancer cells.

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    Xiao-Yan Han

    Full Text Available Despite earlier studies demonstrating characteristics of colon cancer stem cells (CCSCs and the role of epithelial-mesenchymal transition (EMT in tumor development, it remains controversial as to the relationship between CCSCs and EMT. In this study, in order to present an insight into this relationship in colon cancer, we developed HCT116 and HT29 sphere models, which are known to be the cells enriching cancer stem cells. Compared to their parental counterparts, spheroid cells displayed lower homotypic/heterotypic adhesion but higher in vitro migratory/invasive capacity, as well as higher tumorigenic and metastatic potential in vivo. The spheroid cells also demonstrated down-regulated E-cadherin and up-regulated α-SMA and Vimentin expression, which is the typical phenotype of EMT. In order to explore whether this phenomenon is associated to activation of Wnt/β-catenin pathway, we detected several key signaling molecules. Compared with their parental cells, HCT116 and HT29 spheroid cells demonstrated down-regulated expression of GSK3β, but up-regulated expression of Slug and Snail. And also, the up-regulation of nucleus β-catenin in spheroid cells indicated that the free β-catenin transferred from cytoplasm to cell nucleus. Our findings indicate that spheroid cells have the characteristics of colon cancer stem cells, and EMT may account for their stemness and malignancy. And persistent activation of Wnt/β-catenin pathway may play an important role in the EMT of CCSCs.

  15. HIF-1α induces the epithelial-mesenchymal transition in gastric cancer stem cells through the Snail pathway.

    Science.gov (United States)

    Yang, Shi-Wei; Zhang, Zhi-Gang; Hao, Ying-Xue; Zhao, Yong-Liang; Qian, Feng; Shi, Yan; Li, Ping-Ang; Liu, Chun-Yang; Yu, Pei-Wu

    2017-02-07

    Substantial evidence suggests that the epithelial-mesenchymal transition (EMT) phenotype is associated with the invasive characteristics of cancer stem cells (CSCs),which possess an EMT phenotype that may predominate in tumor invasion and metastasis. However, the mechanisms for the generation and regulation of these CSCs have not been clearly defined. As hypoxia and EMT-related factors may have important functions in EMT-like CSCs, the aim of this study was to investigate the effects of hypoxia on these cells. CSCs were established from the gastric cancer cell lines MGC-803 and SGC7901, and the relationship between hypoxia and EMT-like CSCs was investigated in gastric cancer. After hypoxia treatment, some gastric CSCs exhibited a marked increase in hypoxia-inducible factor-1α (HIF-1α)expression and increased migration and invasion capabilities compared with the normoxic control. These CSCs were defined by activation of the mesenchymal cell marker Vimentin and by inhibition of the epithelial cell marker E-cadherin. Our analyses also show that HIF-1α was responsible for activating EMT via increased expression of the transcription factor Snail in gastric CSCs. Moreover, inhibition of Snail by shRNA reduced HIF-1α-induced EMT in gastric CSCs. The results demonstrated that hypoxia-induced EMT-like CSCs rely on HIF-1αto activate Snail, which may result in recurrence and metastasis of gastric cancer.

  16. Snail Enhances Glycolysis in the Epithelial-Mesenchymal Transition Process by Targeting FBP1 in Gastric Cancer.

    Science.gov (United States)

    Yu, Jie; Li, Jing; Chen, Yong; Cao, Wenmiao; Lu, Yuanyuan; Yang, Jianqi; Xing, Enmin

    2017-08-24

    Snail is a key regulator of epithelial-mesenchymal transition (EMT) in cancer. However, the regulatory role and underlying mechanisms of Snail in gastric cancer metabolism are unknown. In this study, we characterized the regulation of aerobic glycolysis by Snail in gastric cancer. The impact of Snail on glucose metabolism was studied in vitro. Combining maximum standardized uptake value (SUVmax), which was obtained preoperatively via a PET/CT scan, with immunohistochemistry staining, we further analyzed the correlation between SUVmax and Snail expression in gastric cancer tissues. Increased expression of Snail promoted lactate production, glucose utilization, and decreased FBP1 expression at both mRNA and protein level. The expression level of Snail was positively associated with SUVmax in gastric cancer patients (P=0.022). Snail and FBP1 expression were inversely correlated at both mRNA and protein level (P=0.002 and P=0.015 respectively) in gastric cancer tissues. Further studies demonstrated that Snail inhibited the FBP1 gene expression at the transcriptional level. Restoring FBP1 expression reversed the effects of glycolysis and EMT induced by Snail in gastric cancer cells. Our results thus reveal that Snail serves as a positive regulator of glucose metabolism through regulation of the FBP1 in gastric cancer. Disrupting the Snail-FBP1 signaling axis may be effective to prevent primary tumor EMT and glycolysis process. © 2017 The Author(s). Published by S. Karger AG, Basel.

  17. Reversibility of the Snail-induced epithelial-mesenchymal transition revealed by the Cre-loxP system.

    Science.gov (United States)

    Ozawa, Masayuki; Kobayashi, Wakako

    2015-03-13

    The epithelial-mesenchymal transition (EMT), a key process in the tumor metastatic cascade, is characterized by the loss of cell-cell junctions and cell polarity, as well as the acquisition of migratory and invasive properties. Snail is an EMT-inducer whose expression in several different epithelial cells, e.g., Madin-Darby canine kidney (MDCK), leads to EMT. To further understand EMT induced by Snail expression, the Cre-loxP site-specific recombination system was used to investigate its reversibility. Transfection of MDCK cells with loxP-flanked Snail (Snail-loxP) resulted in EMT induction, which included the acquisition of a spindle-shaped fibroblastic morphology, the downregulation of epithelial markers, and the upregulation of mesenchymal markers. DNA methylation of the E-cadherin promoter, which often occurs during E-cadherin downregulation, was not observed in Snail+ cells. After Cre-mediated excision of Snail-loxP, the cells reacquired an epithelial morphology, upregulated epithelial markers, and downregulated mesenchymal markers. Thus, EMT induced by Snail expression was reversible. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Histone deacetylase inhibitor valproic acid (VPA) promotes the epithelial mesenchymal transition of colorectal cancer cells via up regulation of Snail.

    Science.gov (United States)

    Feng, Jutao; Cen, Junhua; Li, Jun; Zhao, Rujin; Zhu, Canhua; Wang, Zongxin; Xie, Jiafen; Tang, Wei

    2015-01-01

    Histone deacetylase inhibitors (HDACIs) have been shown to have antiproliferative activity through cell-cycle arrest, differentiation, and apoptosis in colorectal cancer (CRC) cells. Our present study revealed that one HDAC inhibitor, valproic acid (VPA), can obviously promote in vitro motility of HCT-116 and SW480 cells. VPA treatment significantly down regulates the expression of epithelial markers E-Cadherin (E-Cad) and Zona occludin-1(ZO-1) while up regulates the mesenchymal markers Vimentin (Vim) and N-cadherin (N-Cad), suggesting that VPA can trigger the epithelial-mesenchymal transition (EMT) of CRC cells. VPA treatment significantly increases the expression and nuclear localization of Snail, the key transcription factors of EMT. Snail knockdown by siRNAs obviously reverses VPA induced EMT of HCT-116 and SW480 cells. Further, VPA can decrease the ubiquitination, increase the acetylation, and then elevate the stabilization of Snail. VPA also increases the phosphorylation of Akt/GSK-3β. The inhibitor of PI3K/Akt, LY2994002, significantly attenuates VPA induced phosphorylation of Akt and GSK-3β and up regulation of Snail and Vim. Collectively, our data reveal that VPA can trigger the EMT of CRC cells via up regulation of Snail through AKT/GSK-3β signals and post-transcriptional modification. It suggests that more attention should be paid when VPA used as a new anticancer drug for CRC patients.

  19. Celastrol inhibits TGF-β1-induced epithelial-mesenchymal transition by inhibiting Snail and regulating E-cadherin expression.

    Science.gov (United States)

    Kang, Hyereen; Lee, Minjae; Jang, Sung-Wuk

    2013-08-09

    The epithelial-mesenchymal transition (EMT) is a pivotal event in the invasive and metastatic potentials of cancer progression. Celastrol inhibits the proliferation of a variety of tumor cells including leukemia, glioma, prostate, and breast cancer; however, the possible role of celastrol in the EMT is unclear. We investigated the effect of celastrol on the EMT. Transforming growth factor-beta 1 (TGF-β1) induced EMT-like morphologic changes and upregulation of Snail expression. The downregulation of E-cadherin expression and upregulation of Snail in Madin-Darby Canine Kidney (MDCK) and A549 cell lines show that TGF-β1-mediated the EMT in epithelial cells; however, celastrol markedly inhibited TGF-β1-induced morphologic changes, Snail upregulation, and E-cadherin expression. Migration and invasion assays revealed that celastrol completely inhibited TGF-β1-mediated cellular migration in both cell lines. These findings indicate that celastrol downregulates Snail expression, thereby inhibiting TGF-β1-induced EMT in MDCK and A549 cells. Thus, our findings provide new evidence that celastrol suppresses lung cancer invasion and migration by inhibiting TGF-β1-induced EMT. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. RMP promotes epithelial-mesenchymal transition through NF-κB/CSN2/Snail pathway in hepatocellular carcinoma.

    Science.gov (United States)

    Zhou, Wei; Wang, Qi; Xu, Yi; Jiang, Jingting; Guo, Jingchun; Yu, Huijun; Wei, Wenxiang

    2017-06-20

    Epithelial-mesenchymal transition (EMT) is a significant risk factor for metastasis in hepatocellular carcinoma (HCC) patients and with poor prognosis. In this study, we demonstrate the key role of RPB5-mediating protein (RMP) in EMT of HCC cells and the mechanism by which RMP promote EMT. RMP increases migration, invasion, and the progress of EMT of HCC cells, which facilitates the accumulation of Snail, a transcriptional repressor involved in EMT initiation. NF-κB is activated by RMP, which directly promotes the expression of COP9 signalosome 2 (CSN2) to repress the degradation of Snail. Pulmonary metastases mouse model demonstrates that RMP induces metastasis in vivo. Immunohistochemical analysis of human HCC tissues confirms the correlation of RMP with the expression of E-cadherin, p65, CSN2 and Snail in vivo. Collectively, these findings indicate that RMP promotes EMT and HCC metastasis through NF-κB/CSN2/Snail pathway. These results suggest that RMP and p65 may serve as potential candidates of the targets in the treatment of metastatic HCC.

  1. Regulation of matrix stiffness on the epithelial-mesenchymal transition of breast cancer cells under hypoxia environment

    Science.gov (United States)

    Lv, Yonggang; Chen, Can; Zhao, Boyuan; Zhang, Xiaomei

    2017-06-01

    Substrate stiffness and hypoxia are associated with tumor development and progression, respectively. However, the synergy of them on the biological behavior of human breast cancer cell is still largely unknown. This study explored how substrate stiffness regulates the cell phenotype, viability, and epithelial-mesenchymal transition (EMT) of human breast cancer cells MCF-7 under hypoxia (1% O2). TRITC-phalloidin staining showed that MCF-7 cells transformed from round to irregular polygon with stiffness increase either in normoxia or hypoxia. While being accompanied with the upward tendency from a 0.5- to a 20-kPa substrate, the percentage of cell apoptosis was significantly higher in hypoxia than that in normoxia, especially on the 20-kPa substrate. Additionally, it was hypoxia, but not normoxia, that promoted the EMT of MCF-7 by upregulating hypoxia-inducible factor-1α (HIF-1α), vimentin, Snail 1, and matrix metalloproteinase 2 (MMP 2) and 9 (MMP 9), and downregulating E-cadherin simultaneously regardless of the change of substrate stiffness. In summary, this study discovered that hypoxia and stiffer substrate (20 kPa) could synergistically induce phenotype change, apoptosis, and EMT of MCF-7 cells. Results of this study have an important significance on further exploring the synergistic effect of stiffness and hypoxia on the EMT of breast cancer cells and its molecular mechanism.

  2. Tead2 expression levels control the subcellular distribution of Yap and Taz, zyxin expression and epithelial-mesenchymal transition.

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    Diepenbruck, Maren; Waldmeier, Lorenz; Ivanek, Robert; Berninger, Philipp; Arnold, Phil; van Nimwegen, Erik; Christofori, Gerhard

    2014-04-01

    The cellular changes during an epithelial-mesenchymal transition (EMT) largely rely on global changes in gene expression orchestrated by transcription factors. Tead transcription factors and their transcriptional co-activators Yap and Taz have been previously implicated in promoting an EMT; however, their direct transcriptional target genes and their functional role during EMT have remained elusive. We have uncovered a previously unanticipated role of the transcription factor Tead2 during EMT. During EMT in mammary gland epithelial cells and breast cancer cells, levels of Tead2 increase in the nucleus of cells, thereby directing a predominant nuclear localization of its co-factors Yap and Taz via the formation of Tead2-Yap-Taz complexes. Genome-wide chromatin immunoprecipitation and next generation sequencing in combination with gene expression profiling revealed the transcriptional targets of Tead2 during EMT. Among these, zyxin contributes to the migratory and invasive phenotype evoked by Tead2. The results demonstrate that Tead transcription factors are crucial regulators of the cellular distribution of Yap and Taz, and together they control the expression of genes critical for EMT and metastasis.

  3. Suppression of FOXQ1 in benzyl isothiocyanate-mediated inhibition of epithelial-mesenchymal transition in human breast cancer cells.

    Science.gov (United States)

    Sehrawat, Anuradha; Kim, Su-Hyeong; Vogt, Andreas; Singh, Shivendra V

    2013-04-01

    We showed previously that breast cancer chemoprevention with benzyl isothiocyanate (BITC) in MMTV-neu mice was associated with induction of E-cadherin protein in vivo. Loss of E-cadherin expression and induction of mesenchymal markers (e.g. vimentin) are biochemical hallmarks of epithelial-mesenchymal transition (EMT), a developmental process implicated in progression of cancer to aggressive state. This study offers novel insights into the mechanism by which BITC inhibits EMT. Exposure of MDA-MB-231, SUM159 and MDA-MB-468 human breast cancer cells to BITC (2.5 and 5 µM) resulted in transcriptional repression of urokinase-type plasminogen activator (uPA) as well as its receptor (uPAR). However, ectopic expression of uPAR in MDA-MB-468 cells failed to confer protection against induction of E-cadherin and inhibition of cell invasion/migration resulting from BITC treatment. The BITC-mediated induction of E-cadherin and inhibition of cell migration was sustained in MDA-MB-231 and SUM159 cells transiently transfected with an uPAR-targeted small interfering RNA. Overexpression of Forkhead Box Q1 (FOXQ1), whose protein and messenger RNA levels were decreased by BITC treatment in cells and MDA-MB-231 xenografts, conferred marked protection against BITC-mediated inhibition of EMT and cell migration. In conclusion, this study implicates FOXQ1 suppression in BITC-mediated inhibition of EMT in human breast cancer cells.

  4. Ginsenoside Rg3 inhibits epithelial-mesenchymal transition (EMT) and invasion of lung cancer by down-regulating FUT4.

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    Tian, Lili; Shen, Dachuan; Li, Xiaodong; Shan, Xiu; Wang, Xiaoqi; Yan, Qiu; Liu, Jiwei

    2016-01-12

    The epithelial-mesenchymal transition (EMT) is an important factor in lung cancer metastasis, and targeting EMT is a potential therapeutic strategy. Fucosyltransferase IV (FUT4) and its synthetic cancer sugar antigen Lewis Y (LeY) was abnormally elevated in many cancers. In this study, a traditional Chinese medicine ginsenoside Rg3 was used to investigate whether its inhibition to EMT and invasion of lung cancer is by the glycobiology mechanism. We found that Rg3 treatment (25, 50, 100 μg/ml) inhibited cell migration and invasion by wound-healing and transwell assays. Rg3 could significantly alter EMT marker proteins with increased E-cadherin, but decreased Snail, N-cadherin and Vimentin expression. Rg3 also down-regulated FUT4 gene and protein expression in lung cancer cells by qPCR, Western blot and immunofluorescence. After FUT4 down-regulated with shFUT4, EMT was obviously inhibited. Furthermore, the activation of EGFR through decreased LeY biosynthesis was inhibited, which blocked the downstream MAPK and NF-κB signal pathways. In addition, Rg3 reduced tumor volume and weight in xenograft mouse model, and significantly decreased tumor metastasis nodules in lung tissues by tail vein injection. In conclusion, Rg3 inhibits EMT and invasion of lung cancer by down-regulating FUT4 mediated EGFR inactivation and blocking MAPK and NF-κB signal pathways. Rg3 may be a potentially effective agent for the treatment of lung cancer.

  5. Long Non-Coding RNAs: Key Regulators of Epithelial-Mesenchymal Transition, Tumour Drug Resistance and Cancer Stem Cells

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    Heery, Richard; Finn, Stephen P.; Cuffe, Sinead; Gray, Steven G.

    2017-01-01

    Epithelial mesenchymal transition (EMT), the adoption by epithelial cells of a mesenchymal-like phenotype, is a process co-opted by carcinoma cells in order to initiate invasion and metastasis. In addition, it is becoming clear that is instrumental to both the development of drug resistance by tumour cells and in the generation and maintenance of cancer stem cells. EMT is thus a pivotal process during tumour progression and poses a major barrier to the successful treatment of cancer. Non-coding RNAs (ncRNA) often utilize epigenetic programs to regulate both gene expression and chromatin structure. One type of ncRNA, called long non-coding RNAs (lncRNAs), has become increasingly recognized as being both highly dysregulated in cancer and to play a variety of different roles in tumourigenesis. Indeed, over the last few years, lncRNAs have rapidly emerged as key regulators of EMT in cancer. In this review, we discuss the lncRNAs that have been associated with the EMT process in cancer and the variety of molecular mechanisms and signalling pathways through which they regulate EMT, and finally discuss how these EMT-regulating lncRNAs impact on both anti-cancer drug resistance and the cancer stem cell phenotype. PMID:28430163

  6. Ionizing Radiation Promotes Migration and Invasion of Cancer Cells Through Transforming Growth Factor-Beta-Mediated Epithelial-Mesenchymal Transition

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    Zhou Yongchun [Department of Radiation Oncology, Xijing Hospital Fourth Military Medical University, Xi' an (China); Department of Radiation Medicine, College of Preventive Medicine, Xijing Hospital Fourth Military Medical University, Xi' an (China); Liu Junye; Li Jing; Zhang Jie [Department of Radiation Medicine, College of Preventive Medicine, Xijing Hospital Fourth Military Medical University, Xi' an (China); Xu Yuqiao [Department of Pathology, Xijing Hospital Fourth Military Medical University, Xi' an (China); Zhang Huawei; Qiu Lianbo; Ding Guirong [Department of Radiation Medicine, College of Preventive Medicine, Xijing Hospital Fourth Military Medical University, Xi' an (China); Su Xiaoming [Department of Radiation Oncology, 306th Hospital of PLA, Beijing (China); Mei Shi [Department of Radiation Oncology, Xijing Hospital Fourth Military Medical University, Xi' an (China); Guo Guozhen, E-mail: guozhenguo@hotmail.com [Department of Radiation Medicine, College of Preventive Medicine, Xijing Hospital Fourth Military Medical University, Xi' an (China)

    2011-12-01

    Purpose: To examine whether ionizing radiation enhances the migratory and invasive abilities of cancer cells through transforming growth factor (TGF-{beta})-mediated epithelial-mesenchymal transition (EMT). Methods and Materials: Six cancer cell lines originating from different human organs were irradiated by {sup 60}Co {gamma}-ray at a total dose of 2 Gy, and the changes associated with EMT, including morphology, EMT markers, migration and invasion, were observed by microscope, Western blot, immunofluorescence, scratch assay, and transwell chamber assay, respectively. Then the protein levels of TGF-{beta} in these cancer cells were detected by enzyme-linked immunosorbent assay, and the role of TGF-{beta} signaling pathway in the effect of ionizing radiation on EMT was investigate by using the specific inhibitor SB431542. Results: After irradiation with {gamma}-ray at a total dose of 2 Gy, cancer cells presented the mesenchymal phenotype, and compared with the sham-irradiation group the expression of epithelial markers was decreased and of mesenchymal markers was increased, the migratory and invasive capabilities were strengthened, and the protein levels of TGF-{beta} were enhanced. Furthermore, events associated with EMT induced by IR in A549 could be reversed through inhibition of TGF-{beta} signaling. Conclusions: These results suggest that EMT mediated by TGF-{beta} plays a critical role in IR-induced enhancing of migratory and invasive capabilities in cancer cells.

  7. Oct4 mediates tumor initiating properties in oral squamous cell carcinomas through the regulation of epithelial-mesenchymal transition.

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    Lo-Lin Tsai

    Full Text Available BACKGROUND: Overexpression of Oct4, an important transcription factor of embryonic stem cells (ESC, has been reported in several cancers. The aim of this study was to determine the emerging role of Oct4 in oral squamous cell carcinoma (OSCC both in vitro and in vivo. METHODOLOGY/PRINCIPAL FINDING: Tumourigenic activity and molecular mechanisms of Oct4 overexpression or knockdown by lentiviral infection in OSCC was investigated in vitro and in vivo. Initially, we demonstrated that Oct4 expression was increased in OSCC cell lines as compared to a normal oral epithelial cell line SG. Overexpression of Oct4 was demonstrated to enhance cell proliferation, invasiveness, anchorage-independent growth and xenotransplantation tumourigenicity. These findings were coupled with epithelial-mesenchymal transition (EMT transformation in OSCCs. In contrast, the silence of Oct4 significantly blocked the xenograft tumorigenesis of OSCC-derived cancer stem cells (OSCC-CSCs and significantly improved the recipient survival. Clinically, the level of Oct4 expression was higher in recurrent and metastatic OSCC specimens but lower in primary OSCC specimens. CONCLUSION/SIGNIFICANCE: Our results suggest that Oct4-mediated tumorigenecity is associated with the regulation of EMT. Oct4 might be a therapeutic target for OSCC.

  8. Tumor Budding Cells, Cancer Stem Cells and Epithelial-Mesenchymal Transition-type Cells in Pancreatic Cancer

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    Eva eKaramitopoulou

    2013-01-01

    Full Text Available Pancreatic ductal adenocarcinoma (PDAC is one of the most lethal cancers with a 5-year survival rate of less than 5%. Moreover, PDAC escapes early detection and resists treatment. Multiple combinations of genetic alterations are known to occur in PDAC including mutational activation of KRAS, inactivation of p16/CDKN2A and SMAD4 (DPC4 and dysregulation of PTEN/PI3K/AKT signaling. Through their interaction with WNT pathway, the downstream molecules of these pathways have been implicated in the promotion of epithelial-mesenchymal transition (EMT. Emerging evidence has demonstrated that cancer stem cells (CSCs, small populations of which have been identified in PDAC, and EMT-type cells play critical roles in drug resistance, invasion and metastasis in pancreatic cancer. EMT may be histologically represented by the presence of tumor budding which is described as the occurrence of single tumor cells or small clusters (<5 of dedifferentiated cells at the invasive front of gastrointestinal (including colorectal, oesophageal, gastric and ampullary carcinomas and is linked to poor prognosis. Tumor budding has recently been shown to occur frequently in PDAC and to be associated with adverse clinicopathological features and decreased disease-free and overall survival. The aim of this review is to present a short overview on the morphological and molecular aspects that underline the relationship between tumor budding cells, CSCs and EMT-type cells in PDAC.

  9. Snail involves in the transforming growth factor β1-mediated epithelial-mesenchymal transition of retinal pigment epithelial cells.

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    Hui Li

    Full Text Available BACKGROUND: The proliferation of retinal pigment epithelium (RPE cells resulting from an epithelial-mesenchymal transition (EMT plays a key role in proliferative vitreoretinopathy (PVR, which leads to complex retinal detachment and the loss of vision. Genes of Snail family encode the zinc finger transcription factors that have been reported to be essential in EMT during embryonic development and cancer metastasis. However, the function of Snail in RPE cells undergoing EMT is largely unknown. PRINCIPAL FINDINGS: Transforming growth factor beta(TGF-β-1 resulted in EMT in human RPE cells (ARPE-19, which was characterized by the expected decrease in E-cadherin and Zona occludin-1(ZO-1 expression, and the increase in fibronectin and α-smooth muscle actin (α-SMA expression, as well as the associated increase of Snail expression at both mRNA and protein levels. Furthermore, TGF-β1 treatment caused a significant change in ARPE-19 cells morphology, with transition from a typical epithelial morphology to mesenchymal spindle-shaped. More interestingly, Snail silencing significantly attenuated TGF-β1-induced EMT in ARPE-19 cells by decreasing the mesenchymal markers fibronectin and a-SMA and increasing the epithelial marker E-cadherin and ZO-1. Snail knockdown could effectively suppress ARPE-19 cell migration. Finally, Snail was activated in epiretinal membranes from PVR patients. Taken together, Snail plays very important roles in TGF-β-1-induced EMT in human RPE cells and may contribute to the development of PVR. SIGNIFICANCE: Snail transcription factor plays a critical role in TGF-β1-induced EMT in human RPE cells, which provides deep insight into the pathogenesis of human PVR disease. The specific inhibition of Snail may provide a new approach to treat and prevent PVR.

  10. Downregulation of β-catenin decreases the tumorigenicity, but promotes epithelial-mesenchymal transition in breast cancer cells

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    Kai Cai

    2014-01-01

    Full Text Available Background: Wnt/β-catenin signaling pathway plays a key role in human breast cancer progression. In this study, we down regulated β-catenin expression in human breast cancer MDA-MB-231 cells and investigated the effect of β-catenin knockdown on the cell biological characteristics. Materials and Methods: The recombinant plasmids of pSUPER-enhancement green fluorescent protein 1 (EGFP1-scrabble-β-catenin-short hairpin ribonucleic acid (shRNA and pSUPER-EGFP1-β-catenin-shRNA-1 were transfected into MDA-MB-231 cells, respectively, and the stably transfected cells were isolated from G418 selected clones. The β-catenin gene silenced efficiency was measured by quantitative reverse transcriptase polymerase chain reaction (QRT-PCR and Western blot. The biological characteristics of MDA-MB-231 cells with down regulated β-catenin were evaluated by analyzing cell proliferation, clonogenicity, cell mobility and tumorigenicity. The expression of E-cadherin and Vimentin was concurrently detected by QRT-PCR. Results: The β-catenin-shRNA-1 stably transfected MDA-MB-231 cells significantly decreased β-catenin expression, cell proliferation, clonogenicity, and tumorigenicity in Balb/c nude mice compared with the MDA-MB-231 cells transfected with pSUPER-EGFP1-scrabble-β-catenin-shRNA. Interestingly, knockdown of β-catenin led to the reduction of epithelial E-cadherin expression, the increase of cell mobility and mesenchymal vimentin expression in MDA-MB-231 cells, indicating an epithelial to mesenchymal transition. Conclusion: Knockdown of β-catenin expression in human breast cancer MDA-MB-231 cells inhibits cell tumorigenicity in mice, but promotes cell epithelial-mesenchymal transition.

  11. Epithelial mesenchymal transition is required for acquisition of anoikis resistance and metastatic potential in adenoid cystic carcinoma.

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    Jun Jia

    Full Text Available Human adenoid cystic carcinoma (ACC is characterized by diffused invasion of the tumor into adjacent organs and early distant metastasis. Anoikis resistance and epithelial mesenchymal transition (EMT are considered prerequisites for cancer cells to metastasize. Exploring the relationship between these processes and their underlying mechanism of action is a promising way to better understand ACC tumors. We initially established anoikis-resistant sublines of ACC cells; the variant cells revealed a mesenchymal phenotype through Slug-mediated EMT-like transformation and displayed enhanced metastatic potential both in vitro and in vivo. Suppression of EMT by knockdown of Slug significantly impaired anoikis resistance, migration, and invasion of the variant cells. With overexpression of Slug and Twist, we determined that induction of EMT in normal ACC cells could prevent anoikis, albeit partially. These findings strongly suggest that EMT is indispensable in anoikis resistance, at least in ACC cells. Furthermore, we found that the EGFR/PI3K/Akt pathway acts as the common regulator for EMT-like transformation and anoikis resistance, as confirmed by their specific inhibitors. Gefitinib and LY294003 restored the sensibilities of anoikis-resistant cells to anoikis and simultaneously impaired their metastatic potential. In addition, the results from our in vivo model of metastasis suggest that pretreatment with gefitinib promotes mouse survival by alleviating pulmonary metastasis. Most importantly, immunohistochemistry of human ACC specimens showed a correlation between the overexpression of Slug and EGFR staining. This study has demonstrated that Slug-mediated EMT-like transformation is required by human ACC cells to achieve anoikis resistance and their metastatic potential. Targeting the EGFR/PI3K/Akt pathway holds potential as a preventive strategy against distant metastasis of ACC.

  12. Muscadine grape skin extract reverts snail-mediated epithelial mesenchymal transition via superoxide species in human prostate cancer cells.

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    Burton, Liza J; Barnett, Petrina; Smith, Basil; Arnold, Rebecca S; Hudson, Tamaro; Kundu, Kousik; Murthy, Niren; Odero-Marah, Valerie A

    2014-03-12

    Snail transcription factor can induce epithelial-mesenchymal transition (EMT), associated with decreased cell adhesion-associated molecules like E-cadherin, increased mesenchymal markers like vimentin, leading to increased motility, invasion and metastasis. Muscadine grape skin extract (MSKE) has been shown to inhibit prostate cancer cell growth and induce apoptosis without affecting normal prostate epithelial cells. We investigated novel molecular mechanisms by which Snail promotes EMT in prostate cancer cells via Reactive Oxygen Species (ROS) and whether it can be antagonized by MSKE. ARCaP and LNCaP cells overexpressing Snail were utilized to examine levels of reactive oxygen species (ROS), specifically, superoxide, in vitro using Dihydroethidium (DHE) or HydroCy3 dyes. Mitosox staining was performed to determine whether the source of ROS was mitochondrial in origin. We also investigated the effect of Muscadine grape skin extract (MSKE) on EMT marker expression by western blot analysis. Migration and cell viability using MTS proliferation assay was performed following MSKE treatments. Snail overexpression in ARCaP and LNCaP cells was associated with increased concentration of mitochondrial superoxide, in vitro. Interestingly, MSKE decreased superoxide levels in ARCaP and LNCaP cells. Additionally, MSKE and Superoxide Dismutase (SOD) reverted EMT as evidenced by decreased vimentin levels and re-induction of E-cadherin expression in ARCaP-Snail cells after 3 days, concomitant with reduced cell migration. MSKE also decreased Stat-3 activity in ARCaP-Snail cells. This study shows that superoxide species may play a role in Snail transcription factor-mediated EMT. Therefore, therapeutic targeting of Snail with various antioxidants such as MSKE may prove beneficial in abrogating EMT and ROS-mediated tumor progression in human prostate cancer.

  13. CCL21/CCR7 axis activating chemotaxis accompanied with epithelial-mesenchymal transition in human breast carcinoma.

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    Li, Fei; Zou, Zhigeng; Suo, Ning; Zhang, Zongpu; Wan, Fangzhu; Zhong, Guangxin; Qu, Yan; Ntaka, Kwanele Siphelele; Tian, Hua

    2014-09-01

    Secondary lymphoid tissue chemokine (SLC/CCL21) and its receptor CCR7 have been implicated in lymph node metastasis, whereas the mechanism of which remains unclear. Epithelial-mesenchymal transition (EMT) plays an important role in invasion and migration of cancer cells. We presumed that CCL21/CCR7 axis activates EMT process to induce cancer cell invasion and metastasis. Firstly, the expressions of CCR7 and EMT markers were examined by immunohistochemical staining in the primary breast carcinoma tissues from 60 patients who underwent radical mastectomy. Then, we investigated whether CCL21/CCR7 induces EMT process during mediating cancer cell invasion or migration in vitro. By immunohistolochemistry, high expressions of CCR7, Slug and N-cadherin were seen in 60, 65, and 76.67 % of tumors, respectively, and significantly associated with lymph node metastases as well as clinical pathological stage. Furthermore, the CCR7 expression was significantly correlated to Slug and N-cadherin. In vitro, stimulating breast cancer cell lines 1428, MCF-7 and MDA-MB-231 with CCL21, the invasion and migration of tumor cells were promoted, and simultaneously, EMT phenotype of tumor cells was enhanced, including down-regulation of E-cadherin, up-regulation of Slug, Vimentin and N-cadherin at both protein and mRNA levels. Inversely, knockdown of CCR7 by shRNA suppressed tumor cell invasion, migration and EMT phenotype induced by CCL21. These results indicated that CCL21/CCR7 axis could activate EMT process during chemotaxis of breast carcinoma cells.

  14. Specific N-glycan alterations are coupled in epithelial-mesenchymal transition induced by EGF in GE11 epithelial cells.

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    Xu, Qingsong; Qu, Chen; Wang, Wenjing; Gu, Jianguo; Du, Yuguang; Song, Linsheng

    2017-02-01

    Epithelial-mesenchymal transition (EMT) is a phenomenon in cancer progression during which cancer cells undergo remarkable alteration acquiring highly invasive property. The aim of this study was to evaluate specific N-glycan alterations during EMT induced by epidermal growth factor (EGF) in GE11 epithelial cells. Herein, we demonstrated that EGF activated epidermal growth factor receptor (EGFR)/Akt/extracellular signal-regulated kinase (ERK) phosphorylation and promoted GE11 cell proliferation. Meanwhile, EGF stimulated the epithelial cells to undergo morphological alteration, destroying cell-cell inter-contact and exhibiting mesenchymal cells higher metastatic potential. A wound-healing assay showed the migratory ability increased 1.5-fold after EGF treatment. Moreover, the relative intensity of N-cadherin versus E-cadherin increased 2.6-fold, and the E-cadherin distribution in cell-cell junctions became jagged and faint after EGF incubation for 72 h. Interestingly, the amounts of bisecting GlcNAc structure were dramatically declined, by contrast, the formation of β1,6 GlcNAc branches on cell surface was upregulated during EMT induced by EGF. To understand the roles of N-glycans in EGF-induced EMT, the cells were stably transfected with N-acetylglucosaminyltransferase III (GnT-III), which catalyzes the bisecting GlcNAc structure formation. As the markers for EMT, EGF-induced E-cadherin decrease and fibronectin increase were delayed in GnT-III-overexpressing cells. Taken together, these results demonstrated that specific N-glycan alterations were coupled in EMT induced by EGF, which might be contributed to diagnosis and therapy of tumor metastasis.

  15. Epithelial mesenchymal transition of non-small-cell lung cancer cells A549 induced by SPHK1

    Institute of Scientific and Technical Information of China (English)

    Min Ni; Xiao-Lei Shi; Zhi-Gang Qu; Hong Jiang; Zi-Qian Chen; Jun Hu

    2015-01-01

    Objective:To explore the effect and molecular mechanism ofSPHK1 in the invasion and metastasis process of non-small-cell lung cancer cells(A549).Methods:Recombinant retrovirus was used to mediate the production ofA549/vector,A549/SPHK1,A549/scramble, andA549/SPHK1/RNAi that stably expressed or silencedSPHK1.The invasion and migration capacities of A549 cells overexpressing or silencingSPHK1 were determined usingTranswell invasion assay and scratch wound repair experiment.The protein and mRNA expression levels ofE-cadherin, fibronectin, vimentin inA549/vector,A549/SPHK1,A549/scramble,A549/SPHK1/RNAi were detected withWestern blot(WB) and quantitativePCR(QPCR) methods, respectively.Results:Transwell invasion assay and scratch wound repair experiments showed that over-expression of SPHK1 obviously enhanced the invasion and migration capacities ofA549 cells.WB andQPCR detection results showed that, the expression ofE-cadherin(a molecular marker of epithelial cells) and fibronectin, vimentin(molecular markers of mesenchymal cells) inA549 cells was upregulated after overexpression ofSPHK1; whileSPHK1 silencing significantly reduced the invasion and metastasis capacities ofA549cells, upregulated the expression of molecular marker of epithelial cells, and downregulated the expression of molecular marker of mesenchymal cells. Conclusions:SPHK1 promotes epithelial mesenchymal transition of non-small-cell lung cancer cells and affects the invasion and metastasis capacities of these cells.

  16. Trps1 regulates biliary epithelial-mesenchymal transition and has roles during biliary fibrosis in liver grafts: a preliminary study.

    Science.gov (United States)

    Zhe, Cheng; Yu, Fan; Tian, Ju; Zheng, Shuguo

    2015-01-01

    To investigate the role(s) of Trps1 in non-anastomotic biliary stricture (NABS) following liver transplantation. Immunohistochemical and histological techniques were used to detect Trps1, E-cadherin, CK19, vimentin, α-SMA, and collagen deposition. Human intrahepatic biliary epithelial cells (HIBECs) were infected with a Trps1 adenovirus, or transfected with Trps1 short-interfering RNAs (siRNAs). Reverse transcription polymerase chain reaction (RT-PCR) assays and western blotting were used to determine expression levels of epithelial and mesenchymal markers, and Trps1 in HIBECs. Expression of Trps1 and epithelial markers was down-regulated or absent in NABS liver samples. Mesenchymal markers were seen in biliary epithelial cells (BECs), with collagen deposited around the bile duct. Trps1 expression positively correlated with epithelial markers. Expression of epithelial marker mRNAs and proteins in HIBECs decreased with prolonged cold preservation (CP), while mesenchymal marker expression increased. A 12-h CP period led to increased Trps1 mRNA and protein levels. Expression of E-cadherin was increased in HIBECs following Trps1 adenovirus infection and CP/reperfusion injury (CPRI), with vimentin expression levels reduced and CPRI-mediated epithelial-mesenchymal transition (EMT) inhibited. Transfection of HIBECs with Trps1 siRNAs in conjunction with CPRI revealed that E-cadherin expression was decreased, vimentin expression was increased, and CPRI-mediated EMT was promoted. Trps1 is involved in NABS pathogenesis following liver transplantation and negatively correlates with BEC EMT and biliary fibrosis in liver grafts. Trps1 demonstrates antagonistic effects that could reverse EMT.

  17. CD133/Src axis mediates tumor initiating property and epithelial-mesenchymal transition of head and neck cancer.

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    Yu-Syuan Chen

    Full Text Available BACKGROUND: Head and Neck squamous cell carcinoma (HNSCC is a human lethal cancer with clinical, pathological, phenotypical and biological heterogeneity. Caner initiating cells (CICs, which are responsible for tumor growth and coupled with gain of epithelial-mesenchymal transition (EMT, have been identified. Previously, we enriched a subpopulation of head and neck cancer initiating cells (HN-CICs with up-regulation of CD133 and enhancement of EMT. Others demonstrate that Src kinase interacts with and phosphorylates the cytoplasmic domain of CD133. However, the physiological function of CD133/Src signaling in HNSCCs has not been uncovered. METHODOLOGY/PRINCIPAL FINDING: Herein, we determined the critical role of CD133/Src axis modulating stemness, EMT and tumorigenicity of HNSCC and HN-CICs. Initially, down-regulation of CD133 significantly reduced the self-renewal ability and expression of stemness genes, and promoted the differentiation and apoptotic capability of HN-CICs. Additionally, knockdown of CD133 in HN-CICs also lessened both in vitro malignant properties including cell migration/cell invasiveness/anchorage independent growth, and in vivo tumor growth by nude mice xenotransplantation assay. In opposite, overexpression of CD133 enhanced the stemness properties and tumorigenic ability of HNSCCs. Lastly, up-regulation of CD133 increased phosphorylation of Src coupled with EMT transformation in HNSCCs, on the contrary, silence of CD133 or treatment of Src inhibitor inversely abrogated above phenotypic effects, which were induced by CD133 up-regulation in HNSCCs or HN-CICs. CONCLUSION/SIGNIFICANCE: Our results suggested that CD133/Src signaling is a regulatory switch to gain of EMT and of stemness properties in HNSCC. Finally, CD133/Src axis might be a potential therapeutic target for HNSCC by eliminating HN-CICs.

  18. h-Prune is associated with poor prognosis and epithelial-mesenchymal transition in patients with colorectal liver metastases.

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    Hashimoto, Masakazu; Kobayashi, Tsuyoshi; Tashiro, Hirotaka; Arihiro, Koji; Kikuchi, Akira; Ohdan, Hideki

    2016-08-15

    The prognosis of patients with colorectal liver metastases (CRLM) remains low despite advances in chemotherapy and surgery. The expression of h-prune (human homolog of Drosophila prune protein; HGNC13420), an exopolyphosphatase, is correlated with progression and aggressiveness in several cancers and promotes migration and invasion. We investigated the role of h-prune in CRLM. To investigate the role of h-prune, immunohistochemical analysis for h-prune was performed in 87 surgically resected specimens of CRLM obtained between 2001 and 2009 at the Hiroshima University Hospital. Immunohistochemical analysis revealed positive staining for h-prune in 24 (28%) cases. The overall survival rate was significantly lower in h-prune-positive cases than in h-prune-negative cases (p = 0.003). Multivariate analysis showed that h-prune positivity was the only independent factor related to poor overall survival of patients after curative hepatectomy of CRLM. In vitro and in vivo, h-prune-knocked-down and h-prune-overexpressing cells were analyzed. In vitro, h-prune was associated with increased cell motility and upregulation of epithelial-mesenchymal transition (EMT) markers. In a mouse model, h-prune was associated with invasion of the tumor and distant metastases. In summary, h-prune expression is a useful marker to identify high-risk patients for resectable colorectal liver metastasis. h-Prune expression is necessary for cancer cell motility and EMT and is associated with liver and lung metastasis in colorectal cancer cells. h-Prune could be a new prognostic marker and molecular target for CRLM.

  19. The multifaceted role of the embryonic gene Cripto-1 in cancer, stem cells and epithelial-mesenchymal transition

    Science.gov (United States)

    Klauzinska, Malgorzata; Castro, Nadia P.; Rangel, Maria Cristina; Spike, Benjamin T.; Gray, Peter C.; Bertolette, Daniel; Cuttitta, Frank; Salomon, David

    2014-01-01

    Cripto-1 (CR-1)/Teratocarcinoma-derived growth factor1 (TDGF-1) is a cell surface glycosylphosphatidylinositol (GPI)-linked glycoprotein that can function either in cis (autocrine) or in trans (paracrine). The cell membrane cis form is found in lipid rafts and endosomes while the trans acting form lacking the GPI anchor is soluble. As a member of the epidermal growth factor (EGF)/Cripto-1-FRL-1-Cryptic (CFC) family, CR-1 functions as an obligatory co-receptor for the transforming growth factor-β (TGF-β) family members, Nodal and growth and differentiation factors 1 and 3 (GDF1/3) by activating Alk4/Alk7 signaling pathways that involve Smads 2, 3 and 4. In addition, CR-1 can activate non-Smad-dependent signaling elements such as PI3K, Akt and MAPK. Both of these pathways depend upon the 78 kDa glucose regulated protein (GRP78). Finally, CR-1 can facilitate signaling through the canonical Wnt/β-catenin and Notch/Cbf-1 pathways by functioning as a chaperone protein for LRP5/6 and Notch, respectively. CR-1 is essential for early embryonic development and maintains embryonic stem cell pluripotentiality. CR-1 performs an essential role in the etiology and progression of several types of human tumors where it is expressed in a population of cancer stem cells (CSCs) and facilitates epithelial-mesenchymal transition (EMT). In this context, CR-1 can significantly enhance tumor cell migration, invasion and angiogenesis. Collectively, these facts suggest that CR-1 may be an attractive target in the diagnosis, prognosis and therapy of several types of human cancer. PMID:25153355

  20. Interleukin-6 induces an epithelial-mesenchymal transition phenotype in human adamantinomatous craniopharyngioma cells and promotes tumor cell migration

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    Zhou, Jie; Zhang, Chao; Pan, Jun; Chen, Ligang; Qi, Song-Tao

    2017-01-01

    Total resection of adamantinomatous craniopharyngioma (ACP) is complex and often leads to postoperative recurrence. This is due to the tendency of the tumor to invade the surrounding brain tissue and the generation of a local inflammatory state between the tumor cells and parenchyma. While there is evidence to suggest that interleukin-6 (IL-6) induces craniopharyngioma (CP)-associated inflammation, particularly in ACP, the role of IL-6 in the progression of ACP remains unclear. The results of the present study demonstrated that CP inflammation was associated with pathological classification, extent of surgery, degree of calcification and postoperative hypothalamic status scale. Cytokine antibody arrays were conducted to measure the expression of IL-6 and other inflammatory factors in tumor tissues in response to various levels of inflammatory exposure. IL-6, IL-6 receptor (IL-6R) and glycoprotein 130 expression was detected by immunohistochemistry. In addition, an ELISA was performed to quantify the levels of soluble IL-6R (sIL-6R) in the cystic fluid and supernatants of ACP cells and tumor-associated fibroblasts. These measurements demonstrated that ACP cells produce IL-6 and its associated proteins. In addition, the results revealed that while the viability of ACP cells was not affected, the migration of ACP cells was promoted by IL-6 treatment in a concentration-dependent manner. Conversely, treatment with an IL-6-blocking monoclonal antibody significantly decreased the migration of ACP cells. In addition, IL-6 treatment increased the expression of vimentin and decreased the expression of E-cadherin in a dose-dependent manner. The findings of the present study demonstrate that IL-6 may promote migration in vitro via the classic- and trans-signaling pathways by inducing epithelial-mesenchymal transition in ACP cell cultures. PMID:28487953

  1. Investigating the link between epithelial-mesenchymal transition and the cancer stem cell phenotype: A mathematical approach.

    Science.gov (United States)

    Turner, C; Kohandel, M

    2010-08-07

    Under the cancer stem cell (CSC) hypothesis, sustained metastatic growth requires the dissemination of a CSC from the primary tumour followed by its re-establishment in a secondary site. The epithelial-mesenchymal transition (EMT), a differentiation process crucial to normal development, has been implicated in conferring metastatic ability on carcinomas. Balancing these two concepts has led researchers to investigate a possible link between EMT and the CSC phenotype-indeed, recent evidence indicates that, following induction of EMT in human breast cancer and related cell lines, stem cell activity increased, as judged by the presence of cells displaying the CD44(high)/CD24(low) phenotype and an increase in the ability of cells to form mammospheres. We mathematically investigate the nature of this increase in stem cell activity. A stochastic model is used when small number of cells are under consideration, namely in simulating the mammosphere assay, while a related continuous model is used to probe the dynamics of larger cell populations. Two scenarios of EMT-mediated CSC enrichment are considered. In the first, differentiated cells re-acquire a CSC phenotype-this model implicates fully mature cells as key subjects of de-differentiation and entails a delay period of several days before de-differentiation occurs. In the second, pre-existing CSCs experience accelerated division and increased proportion of self-renewing divisions; a lack of perfect CSC biomarkers and cell sorting techniques requires that this model be considered, further emphasizing the need for better characterization of the mammary (cancer) stem cell hierarchy. Additionally, we suggest the utility of comparing mammosphere data to computational mammosphere simulations in elucidating the growth characteristics of mammary (cancer) stem cells.

  2. Human papillomavirus oncoproteins differentially modulate epithelial-mesenchymal transition in 5-FU-resistant cervical cancer cells.

    Science.gov (United States)

    Vishnoi, Kanchan; Mahata, Sutapa; Tyagi, Abhishek; Pandey, Arvind; Verma, Gaurav; Jadli, Mohit; Singh, Tejveer; Singh, Sukh Mahendra; Bharti, Alok C

    2016-10-01

    Etiological role of viral proteins E6 and E7 of high-risk HPV in cervical carcinogenesis is well established. However, their contribution in chemoresistance and epithelial-mesenchymal transition (EMT) that leads to advanced metastatic lesions and chemoresistance is poorly defined. In the present study, contribution of viral oncoproteins in acquisition of EMT character during onset of chemoresistance was assessed. A chemoresistant cell line (SiHaCR) was developed from an established HPV16-positive cervical cancer cell line, SiHa, by escalating selection pressure of 5-fluorouracil (5-FU). Expression of Survivin, ABCG2, Snail, Slug, Twist, and Vimentin was examined in SiHa and SiHaCR cells by reverse transcriptase-PCR (RT-PCR) and immunoblotting assays. Mesenchymal phenotype in SiHaCR cells was confirmed by assessment of migration and invasion potentials. SiHaCR cells displayed elevated level of functional and molecular markers associated with chemoresistance (Survivin, ABCG2) and EMT (Snail, Slug, Twist, Vimentin) and reduced E-cadherin. SiHaCR also showed increased levels of HPV16 E6 and E7 transcripts. Specific silencing of HPV16 E6, but not E7 using corresponding siRNA, demonstrated a differential involvement of HPV oncogenes in manifestation of EMT. HPV16 E6 silencing resulted in reduction of Slug and Twist expression. However, the expression of Snail and Vimentin was only marginally affected. In contrast, there was an increase in the expression of E-cadherin. A reduced migration and invasion capabilities were observed only in E6-silenced SiHaCR cells, which further confirmed functional contribution of HPV16 E6 in manifestation of EMT. Taken together, our study demonstrated an active involvement of HPV16 E6 in regulation of EMT, which promotes chemoresistance in cervical cancer.

  3. Gefitinib inhibits invasive phenotype and epithelial-mesenchymal transition in drug-resistant NSCLC cells with MET amplification.

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    Silvia La Monica

    Full Text Available Despite the initial response, all patients with epidermal growth factor receptor (EGFR-mutant non-small cell lung cancer (NSCLC eventually develop acquired resistance to EGFR tyrosine kinase inhibitors (TKIs. The EGFR-T790M secondary mutation is responsible for half of acquired resistance cases, while MET amplification has been associated with acquired resistance in about 5-15% of NSCLCs. Clinical findings indicate the retained addiction of resistant tumors on EGFR signaling. Therefore, we evaluated the molecular mechanisms supporting the therapeutic potential of gefitinib maintenance in the HCC827 GR5 NSCLC cell line harbouring MET amplification as acquired resistance mechanism. We demonstrated that resistant cells can proliferate and survive regardless of the presence of gefitinib, whereas the absence of the drug significantly enhanced cell migration and invasion. Moreover, the continuous exposure to gefitinib prevented the epithelial-mesenchymal transition (EMT with increased E-cadherin expression and down-regulation of vimentin and N-cadherin. Importantly, the inhibition of cellular migration was correlated with the suppression of EGFR-dependent Src, STAT5 and p38 signaling as assessed by a specific kinase array, western blot analysis and silencing functional studies. On the contrary, the lack of effect of gefitinib on EGFR phosphorylation in the H1975 cells (EGFR-T790M correlated with the absence of effects on cell migration and invasion. In conclusion, our findings suggest that certain EGFR-mutated patients may still benefit from a second-line therapy including gefitinib based on the specific mechanism underlying tumor cell resistance.

  4. Trps1 regulates biliary epithelial-mesenchymal transition and has roles during biliary fibrosis in liver grafts: a preliminary study.

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    Cheng Zhe

    Full Text Available To investigate the role(s of Trps1 in non-anastomotic biliary stricture (NABS following liver transplantation.Immunohistochemical and histological techniques were used to detect Trps1, E-cadherin, CK19, vimentin, α-SMA, and collagen deposition. Human intrahepatic biliary epithelial cells (HIBECs were infected with a Trps1 adenovirus, or transfected with Trps1 short-interfering RNAs (siRNAs. Reverse transcription polymerase chain reaction (RT-PCR assays and western blotting were used to determine expression levels of epithelial and mesenchymal markers, and Trps1 in HIBECs.Expression of Trps1 and epithelial markers was down-regulated or absent in NABS liver samples. Mesenchymal markers were seen in biliary epithelial cells (BECs, with collagen deposited around the bile duct. Trps1 expression positively correlated with epithelial markers. Expression of epithelial marker mRNAs and proteins in HIBECs decreased with prolonged cold preservation (CP, while mesenchymal marker expression increased. A 12-h CP period led to increased Trps1 mRNA and protein levels. Expression of E-cadherin was increased in HIBECs following Trps1 adenovirus infection and CP/reperfusion injury (CPRI, with vimentin expression levels reduced and CPRI-mediated epithelial-mesenchymal transition (EMT inhibited. Transfection of HIBECs with Trps1 siRNAs in conjunction with CPRI revealed that E-cadherin expression was decreased, vimentin expression was increased, and CPRI-mediated EMT was promoted.Trps1 is involved in NABS pathogenesis following liver transplantation and negatively correlates with BEC EMT and biliary fibrosis in liver grafts. Trps1 demonstrates antagonistic effects that could reverse EMT.

  5. α-Solanine inhibits invasion of human prostate cancer cell by suppressing epithelial-mesenchymal transition and MMPs expression.

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    Shen, Kun-Hung; Liao, Alex Chien-Hwa; Hung, Jui-Hsiang; Lee, Wei-Jiunn; Hu, Kai-Chieh; Lin, Pin-Tsen; Liao, Ruei-Fang; Chen, Pin-Shern

    2014-08-11

    α-Solanine, a naturally occurring steroidal glycoalkaloid found in nightshade (Solanum nigrum Linn.), was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in suppression of cancer cell metastasis by α-solanine remains unclear. This study investigates the suppression mechanism of α-solanine on motility of the human prostate cancer cell PC-3. Results show that α-solanine reduces the viability of PC-3 cells. When treated with non-toxic doses of α-solanine, cell invasion is markedly suppressed by α-solanine. α-Solanine also significantly elevates epithelial marker E-cadherin expression, while it concomitantly decreases mesenchymal marker vimentin expression, suggesting it suppresses epithelial-mesenchymal transition (EMT). α-Solanine reduces the mRNA level of matrix metalloproteinase-2 (MMP-2), MMP-9 and extracellular inducer of matrix metalloproteinase (EMMPRIN), but increases the expression of reversion-inducing cysteine-rich protein with kazal motifs (RECK), and tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2. Immunoblotting assays indicate α-solanine is effective in suppressing the phosphorylation of phosphatidylinositide-3 kinase (PI3K), Akt and ERK. Moreover, α-solanine downregulates oncogenic microRNA-21 (miR-21) and upregulates tumor suppressor miR-138 expression. Taken together, the results suggest that inhibition of PC-3 cell invasion by α-solanine may be, at least in part, through blocking EMT and MMPs expression. α-Solanine also reduces ERK and PI3K/Akt signaling pathways and regulates expression of miR-21 and miR-138. These findings suggest an attractive therapeutic potential of α-solanine for suppressing invasion of prostate cancer cell.

  6. Ursolic acid inhibits the proliferation of human ovarian cancer stem-like cells through epithelial-mesenchymal transition.

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    Zhang, Jie; Wang, Wenjing; Qian, Lin; Zhang, Qiuwan; Lai, Dongmei; Qi, Cong

    2015-11-01

    Ovarian cancer is the most frequent cause of cancer-related death among all gynecological cancers. Increasing evidence suggests that human ovarian cancer stem-like cells could be enriched under serum-free culture conditions. In the present study, SKOV3 ovarian epithelial cancer cells were cultured for sphere cells. Ursolic acid (UA) with triterpenoid compounds exist widely in food, medicinal herbs and other plants. Evidence shows that UA has anticancer activities in human ovarian cancer cells, but he role of UA in ovarian cancer stem cells (CSCs) remains unknown. The aim of the present study was to investigate the anticancer effects of UA in combination with cisplatin in ovarian CSCs (in vitro and in vivo), along with the molecular mechanism of action. Treatment with UA at various concentrations was examined in combination with cisplatin in human ovarian CSCs. MTT assay and flow cytometry were used for cell viability and apoptosis analysis, and qRT-PCR for stem cell markers and epithelial-mesenchymal transition (EMT) markers for mRNA expression. Transwell assay was employed to observe the migration and invasion of SKOV3 cells and SKOV3 sphere cells after treatment. Moreover, athymic BALB/c-nu nude mice were injected with SKOV3 sphere cells to obtain a xenograft model for in vivo studies. The results showed that CSCs possessed mesenchymal characteristics and EMT ability, and the growth of SKOV3 and sphere cells was significantly inhibited by UA. Transplanted tumors were significantly reduced after injection of UA and UA plus cisplatin. Furthermore, we found that UA could play a role in enhancing the sensitivity of CSCs to cisplatin resistance. Our findings suggested that UA is involved in EMT mechanism to affect the proliferation and apoptosis of human ovarian cancer stem-like cells and it is a potent anti-ovarian cancer agent.

  7. The Cain and Abl of epithelial-mesenchymal transition and transforming growth factor-β in mammary epithelial cells.

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    Allington, Tressa M; Schiemann, William P

    2011-01-01

    Transforming growth factor-β (TGF-β) normally inhibits breast cancer development by preventing mammary epithelial cell (MEC) proliferation, by inducing MEC apoptosis, and by creating cell microenvironments that maintain MEC homeostasis and prevent their uncontrolled growth and motility. Mammary tumorigenesis elicits dramatic alterations in MEC architecture and microenvironment integrity, which collectively counteract the tumor-suppressing activities of TGF-β and enable its stimulation of breast cancer invasion and metastasis. How malignant MECs overcome the cytostatic actions imposed by normal microenvironments and TGF-β, and how abnormal microenvironments conspire with TGF-β to stimulate the development and progression of mammary tumors remains largely undefined. These knowledge gaps have prevented science and medicine from implementing treatments effective in simultaneously targeting abnormal cellular microenvironments, and in antagonizing the oncogenic activities of TGF-β in developing and progressing breast cancers. c-Abl is a ubiquitously expressed nonreceptor protein tyrosine kinase that essentially oversees all aspects of cell physiology, including the regulation of cell proliferation, migration and adhesion, as well as that of cell survival. Thus, the biological functions of c-Abl are highly reminiscent of those attributed to TGF-β, including the ability to function as either a suppressor or promoter of tumorigenesis. Interestingly, while dysregulated Abl activity clearly promotes tumorigenesis in hematopoietic cells, an analogous role for c-Abl in regulating solid tumor development, including those of the breast, remains controversial. Here, we review the functions of c-Abl in regulating breast cancer development and progression, and in alleviating the oncogenic activities of TGF-β and its stimulation of epithelial-mesenchymal transition during mammary tumorigenesis.

  8. IL-6 promotes growth and epithelial-mesenchymal transition of CD133+ cells of non-small cell lung cancer.

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    Lee, Soo Ok; Yang, Xiaodong; Duan, Shanzhou; Tsai, Ying; Strojny, Laura R; Keng, Peter; Chen, Yuhchyau

    2016-02-09

    We examined IL-6 effects on growth, epithelial-mesenchymal transition (EMT) process, and metastatic ability of CD133+ and CD133- cell subpopulations isolated from three non-small cell lung cancer (NSCLC) cell lines: A549, H157, and H1299. We developed IL-6 knocked-down and scramble (sc) control cells of A549 and H157 cell lines by lentiviral infection system, isolated CD133+ and CD133- sub-populations, and investigated the IL-6 role in self-renewal/growth of these cells. IL-6 showed either an inhibitory or lack of effect in modulating growth of CD133- cells depending on intracellular IL-6 levels, but there was higher self-renewal ability of IL-6 expressing CD133+ cells than IL-6 knocked down cells, confirming the promoter role of IL-6 in CD133+ cells growth. We then examined tumor growth of xenografts developed from CD133+ cells of A549IL-6si vs. A549sc cell lines. Consistently, there was retarded growth of tumors developed from A549IL-6si, CD133+ cells compared to tumors originating from A549sc, CD133+ cells. The effect of IL-6 in promoting CD133+ self-renewal was due to hedgehog (Hhg) and Erk signaling pathway activation and higher Bcl-2/Bcl-xL expression. We also investigated whether IL-6 regulates the EMT process of CD133- and CD133+ cells differently. Expression of the EMT/metastasis-associated molecules in IL-6 expressing cells was higher than in IL-6 knocked down cells. Together, we demonstrated dual roles of IL-6 in regulating growth of CD133- and CD133+ subpopulations of lung cancer cells and significant regulation of IL-6 on EMT/metastasis increase in CD133+ cells, not in CD133- cells.

  9. Expression of Angiogenesis Regulatory Proteins and Epithelial-Mesenchymal Transition Factors in Platelets of the Breast Cancer Patients

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    Hui Han

    2014-01-01

    Full Text Available Platelets play a role in tumor angiogenesis and growth and are the main transporters of several angiogenesis regulators. Here, we aimed to determine the levels of angiogenesis regulators and epithelial-mesenchymal transition factors sequestered by circulating platelets in breast cancer patients and age-matched healthy controls. Platelet pellets (PP and platelet-poor plasma (PPP were collected by routine protocols. Vascular endothelial growth factor (VEGF, platelet-derived growth factor BB (PDGF-BB, thrombospondin-1 (TSP-1, platelet factor 4 (PF4, and transforming growth factor-β1 (TGF-β1 were measured by enzyme-linked immunosorbent assay. Angiogenesis-associated expression of VEGF (2.1 pg/106 platelets versus 0.9 pg/106 platelets, P < 0.001, PF4 (21.2 ng/106 platelets versus 10.2 ng/106 platelets, P < 0.001, PDGF-BB (42.9 pg/106 platelets versus 19.1 pg/106 platelets, P < 0.001, and TGF-β1 (15.3 ng/106 platelets versus 4.3 ng/106 platelets, P < 0.001 differed in the PP samples of cancer and control subjects. In addition, protein concentrations were associated with clinical characteristics (P<0.05. Circulating platelets in breast cancer sequester higher levels of PF4, VEGF, PDGF-BB, and TGF-β1, suggesting a possible target for early diagnosis. VEGF, PDGF, and TGF-β1 concentrations in platelets may be associated with prognosis.

  10. EGF induces epithelial-mesenchymal transition through phospho-Smad2/3-Snail signaling pathway in breast cancer cells.

    Science.gov (United States)

    Kim, Jinkyoung; Kong, Jienan; Chang, Hyeyoon; Kim, Hayeon; Kim, Aeree

    2016-12-20

    Epithelial-mesenchymal transition (EMT) can contribute to tumor invasion, metastasis, and resistance to chemotherapy or hormone therapy. EMT may be induced by a variety of growth factors, such as epidermal growth factor (EGF). Most studies regarding EMT have focused on TGF-β-Smads signaling. The mechanism of EGF-induced EMT via activation of the Smad2/3 in breast cancer cells, MCF-7 and MDA-MB-231, remains unclear. The expression levels of Snail, vimentin, and fibronectin were increased by EGF treatment in a time-dependent manner, while the expression level of E-cadherin was decreased. EGF-induced nuclear co-localization of phospho-Smad2/3 and Snail and cancer cell migration were inhibited by pretreatment with an ERK1/2 inhibitor, PD98059 and a phospho-Smad2 inhibitor, SB203580. Knockdown of Smad2/3 expression suppressed EGF-induced expressions of Snail, vimentin, fibronectin, and cancer cell invasion, suggesting an acquisition of the mesenchymal and migratory phenotype in less aggressive MCF-7 cells. Moreover, MDA-MB-231 cells were shown that EGF-induced EMT, and cell invasion through ERK1/2-phospho-Smad2/3-Snail signaling pathway. We have discovered that EGF-stimulated activation of Smad2/3 upregulated several key EMT markers, inhibited E-cadherin expression, promoted EMT, enhanced migration and invasion in MCF-7 and MDA-MB-231 breast cancer cells. Identification of this molecular mechanism may provide new molecular targets for the development of therapies for metastatic breast cancer.

  11. The critical role of EGF-β-catenin signaling in the epithelial-mesenchymal transition in human glioblastoma

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    Wang X

    2017-05-01

    Full Text Available Xingqiang Wang, Shanshi Wang, Xiaolong Li, Shigang Jin, Feng Xiong, Xin Wang Department of Neurosurgery, People’s Hospital of Rizhao, Jining Medical University, Rizhao, China Abstract: To date, β-catenin has been reported to be implicated in mediating the epithelial-mesenchymal transition (EMT in a variety of human cancers, which can be triggered by EGF. However, the mechanisms underlying EGF-β-catenin pathway-induced EMT of glioblastoma multiforme (GBM have not been reported previously. In the present study, immunohistochemistry, reverse transcription polymerase chain reaction, and Western blot were applied to investigate the effect of EGF-β-catenin pathway on EMT of GBM. Here, we identified that β-catenin mRNA and protein levels were up-regulated in GBM tissues and four kinds of glioblastoma cell lines, including T98G, A172, U87, and U251 cells, compared with normal brain tissue and astrocytes. In U87 cell line, inhibition of β-catenin by siRNA suppressed EGF-induced proliferation, migration, invasiveness, and the expression of EMT activators (Snail and Slug. In addition, the expression of epithelial markers (E-cadherin was up-regulated and the expression of mesenchymal markers (N-cadherin and MMP9 was down-regulated. Finally, inhibitor of PI3K/Akt signaling pathways inactivated the EGF-β-catenin-induced EMT. In conclusion, β-catenin-EMT pathway induced by EGF is important for GBM progression by the PI3K/Akt pathways. Inhibition of β-catenin leads to suppression of EGF pathway-induced EMT, which provides a new way to treat GBM patients. Keywords: EGF, β-catenin, EMT, GBM

  12. Muscadine grape skin extract reverts snail-mediated epithelial mesenchymal transition via superoxide species in human prostate cancer cells

    Science.gov (United States)

    2014-01-01

    Background Snail transcription factor can induce epithelial-mesenchymal transition (EMT), associated with decreased cell adhesion-associated molecules like E-cadherin, increased mesenchymal markers like vimentin, leading to increased motility, invasion and metastasis. Muscadine grape skin extract (MSKE) has been shown to inhibit prostate cancer cell growth and induce apoptosis without affecting normal prostate epithelial cells. We investigated novel molecular mechanisms by which Snail promotes EMT in prostate cancer cells via Reactive Oxygen Species (ROS) and whether it can be antagonized by MSKE. Methods ARCaP and LNCaP cells overexpressing Snail were utilized to examine levels of reactive oxygen species (ROS), specifically, superoxide, in vitro using Dihydroethidium (DHE) or HydroCy3 dyes. Mitosox staining was performed to determine whether the source of ROS was mitochondrial in origin. We also investigated the effect of Muscadine grape skin extract (MSKE) on EMT marker expression by western blot analysis. Migration and cell viability using MTS proliferation assay was performed following MSKE treatments. Results Snail overexpression in ARCaP and LNCaP cells was associated with increased concentration of mitochondrial superoxide, in vitro. Interestingly, MSKE decreased superoxide levels in ARCaP and LNCaP cells. Additionally, MSKE and Superoxide Dismutase (SOD) reverted EMT as evidenced by decreased vimentin levels and re-induction of E-cadherin expression in ARCaP-Snail cells after 3 days, concomitant with reduced cell migration. MSKE also decreased Stat-3 activity in ARCaP-Snail cells. Conclusions This study shows that superoxide species may play a role in Snail transcription factor-mediated EMT. Therefore, therapeutic targeting of Snail with various antioxidants such as MSKE may prove beneficial in abrogating EMT and ROS-mediated tumor progression in human prostate cancer. PMID:24617993

  13. Notch-1 induces epithelial-mesenchymal transition consistent with cancer stem cell phenotype in pancreatic cancer cells.

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    Bao, Bin; Wang, Zhiwei; Ali, Shadan; Kong, Dejuan; Li, Yiwei; Ahmad, Aamir; Banerjee, Sanjeev; Azmi, Asfar S; Miele, Lucio; Sarkar, Fazlul H

    2011-08-01

    Activation of Notch-1 is known to be associated with the development and progression of human malignancies including pancreatic cancer. Emerging evidence suggest that the acquisition of epithelial-mesenchymal transition (EMT) phenotype and induction of cancer stem cell (CSC) or cancer stem-like cell phenotype are interrelated and contributes to tumor recurrence and drug resistance. The molecular mechanism(s) by which Notch-1 contributes to the acquisition of EMT phenotype and CSC self-renewal capacity has not been fully elucidated. Here we show that forced over-expression of Notch-1 leads to increased cell growth, clonogenicity, migration and invasion of AsPC-1 cells. Moreover, over-expression of Notch-1 led to the induction of EMT phenotype by activation of mesenchymal cell markers such as ZEB1, CD44, EpCAM, and Hes-1. Here we also report, for the first time, that over-expression of Notch-1 leads to increased expression of miR-21, and decreased expression of miR-200b, miR-200c, let-7a, let-7b, and let-7c. Re-expression of miR-200b led to decreased expression of ZEB1, and vimentin, and increased expression of E-cadherin. Over-expression of Notch-1 also increased the formation of pancreatospheres consistent with expression of CSC surface markers CD44 and EpCAM. Finally, we found that genistein, a known natural anti-tumor agent inhibited cell growth, clonogenicity, migration, invasion, EMT phenotype, formation of pancreatospheres and expression of CD44 and EpCAM. These results suggest that the activation of Notch-1 signaling contributes to the acquisition of EMT phenotype, which is in part mediated through the regulation of miR-200b and CSC self-renewal capacity, and these processes could be attenuated by genistein treatment.

  14. The multifaceted role of the embryonic gene Cripto-1 in cancer, stem cells and epithelial-mesenchymal transition.

    Science.gov (United States)

    Klauzinska, Malgorzata; Castro, Nadia P; Rangel, Maria Cristina; Spike, Benjamin T; Gray, Peter C; Bertolette, Daniel; Cuttitta, Frank; Salomon, David

    2014-12-01

    Cripto-1 (CR-1)/Teratocarcinoma-derived growth factor1 (TDGF-1) is a cell surface glycosylphosphatidylinositol (GPI)-linked glycoprotein that can function either in cis (autocrine) or in trans (paracrine). The cell membrane cis form is found in lipid rafts and endosomes while the trans acting form lacking the GPI anchor is soluble. As a member of the epidermal growth factor (EGF)/Cripto-1-FRL-1-Cryptic (CFC) family, CR-1 functions as an obligatory co-receptor for the transforming growth factor-β (TGF-β) family members, Nodal and growth and differentiation factors 1 and 3 (GDF1/3) by activating Alk4/Alk7 signaling pathways that involve Smads 2, 3 and 4. In addition, CR-1 can activate non-Smad-dependent signaling elements such as PI3K, Akt and MAPK. Both of these pathways depend upon the 78kDa glucose regulated protein (GRP78). Finally, CR-1 can facilitate signaling through the canonical Wnt/β-catenin and Notch/Cbf-1 pathways by functioning as a chaperone protein for LRP5/6 and Notch, respectively. CR-1 is essential for early embryonic development and maintains embryonic stem cell pluripotentiality. CR-1 performs an essential role in the etiology and progression of several types of human tumors where it is expressed in a population of cancer stem cells (CSCs) and facilitates epithelial-mesenchymal transition (EMT). In this context, CR-1 can significantly enhance tumor cell migration, invasion and angiogenesis. Collectively, these facts suggest that CR-1 may be an attractive target in the diagnosis, prognosis and therapy of several types of human cancer.

  15. Gli-1 is crucial for hypoxia-induced epithelial-mesenchymal transition and invasion of breast cancer.

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    Lei, Jianjun; Fan, Lin; Wei, Guangbing; Chen, Xin; Duan, Wanxing; Xu, Qinhong; Sheng, Wei; Wang, Kang; Li, Xuqi

    2015-04-01

    Hypoxia can induce HIF-1α expression and promote the epithelial-mesenchymal transition (EMT) and invasion of cancer cells. However, their mechanisms remain unclear. The objective of this study was to evaluate the role of Gli-1, an effector of the Hedgehog pathway, in the hypoxia-induced EMT and invasion of breast cancer cells. Human breast cancer MDA-MB-231 cells were transfected with HIF-1α or Gli-1-specific small interfering RNA (siRNA) and cultured under a normoxic or hypoxic condition. The relative levels of HIF-1α, Gli-1, E-cadherin, and vimentin in the cells were characterized by quantitative RT-PCR and Western blot assays, and the invasion of MDA-MB-231 cells was determined. Data was analyzed by Student T test, one-way ANOVA, and post hoc LSD test or Mann-Whitney U when applicable. We observed that hypoxia significantly upregulated the relative levels of vimentin expression, but downregulated E-cadherin expression and promoted the invasion of MDA-MB-231 cells, associated with upregulated HIF-1α translation and Gil-1 expression. Knockdown of HIF-1α mitigated hypoxia-modulated Gil-1, vimentin and E-cadherin expression, and invasion of MDA-MB-231 cells. Knockdown of Gil-1 did not significantly change hypoxia-upregulated HIF-1α translation but completely eliminated hypoxia-modulated vimentin and E-cadherin expression and invasion of MDA-MB-231 cells. These data indicate that Gil-1 is crucial for hypoxia-induced EMT and invasion of breast cancer cells and may be a therapeutic target for intervention of breast cancer metastasis.

  16. RYBP Inhibits Progression and Metastasis of Lung Cancer by Suppressing EGFR Signaling and Epithelial-Mesenchymal Transition

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    Xiaoxiao Dinglin

    2017-04-01

    Full Text Available Lung cancer (LC is a common lethal malignancy with rapid progression and metastasis, and Ring1 and YY1 binding protein (RYBP has been shown to suppress cell growth in human cancers. This study aimed to investigate the role of RYBP in LC progression and metastasis. In this study, a total of 149 LC patients were recruited, and the clinical stage of their tumors, metastasis status, survival time, presence of epidermal growth factor receptor (EGFR mutation, and RYBP expression levels were measured. RYBP silencing and overexpression were experimentally performed in LC cell lines and in nude mice, and the expressions of genes in EGFR-related signaling pathways and epithelial-mesenchymal transition (EMT were detected. The results showed that RYBP was downregulated in LC compared with adjacent normal tissues, and low RYBP expression was associated with a more severe clinical stage, high mortality, high metastasis risk, and poor survival. Cell proliferation and xenograft growth were inhibited by RYBP overexpression, whereas proliferation and xenograft growth were accelerated by RYBP silencing. EGFR and phosphorylated-EGFR levels were upregulated when RYBP was silenced, whereas EGFR, p-EGFR, p-AKT, and p-ERK were downregulated when RYBP was overexpressed. Low RYBP expression was related to a high metastasis risk, and metastasized tumors showed low RYBP levels. Cell migration and invasion were promoted by silencing RYBP but were inhibited by overexpressed RYBP. In addition, the EMT marker vimentin showed diminished expression, and E-cadherin was promoted by the overexpression of RYBP. In conclusion, our data suggest that RYBP suppresses cell proliferation and LC progression by impeding the EGFR-ERK and EGFR-AKT signaling pathways and thereby inhibiting cell migration and invasion and LC metastasis through the suppression of EMT.

  17. Influence of connective tissue growth factor antisense oligonucleotide on angiotensin Ⅱ-induced epithelial mesenchymal transition in HK2 cells

    Institute of Scientific and Technical Information of China (English)

    Long CHEN; Bi-cheng LIU; Xiao-liang ZHANG; Jian-dong ZHANG; Hong LIU; Min-xia LI

    2006-01-01

    Aim: The present study was designed to further investigate the effect of connective tissue growth factor antisense oligonucleotide (CTGF-AS) on angiotensin Ⅱ (Ang Ⅱ)-induced tubular cell epithelial mesenchymal transition (EMT) in vitro. Methods: The human proximal tubular cell line (HK2) was grown in Dulbecco's modified Eagle's medium containing 10% heat inactivated fetal calf serum. After being rested in serum-free medium for 24 h, the influence of CTGF-AS (20 μg/mL) on Ang Ⅱ-induced (1×10-7 mol/L) CTGF mRNA and the protein expression were examined by using reverse transcription-polymerase chain reaction and indirectimmunofluorescence. The effect of CTGF-AS on Ang Ⅱ-induced cellular ultrastructure was observed using a transmissive electronic microscope. The expression of α-smooth action (α-SMA) was assayed by immunocytochemistry. In all experiments, the control group was treated with scrambled oligonucleotide. Results: It was shown that Ang Ⅱ significantly induced the increasing expression of CTGF mRNA and protein (P<0.01, respectively), which were significantly abolished by treatment with CTGF-AS. After stimulating cells with Ang Ⅱ, the cellular ultrastructure showed mesenchymal features. These effects were partially inhibited by CTGF-AS. Ang Ⅱ significantly resulted in the expression of α-SMA in time dependent manner, which was markedly attenuated by the treatment with CTGF-AS (P<0.01, respectively). In contrast, no similar effects were observed in the control group treated with scrambled oligonucleotide. Conclusion: Ang Ⅱ-induced EMT in human proximal tubular epithelial cells (PTC) can be attenuated by treatment with CTGF-AS. Our data provides further evidence that CTGF might be involved in Ang Ⅱ-induced EMT in PTC.

  18. Epithelial-mesenchymal transition in keloid tissues and TGF-β1-induced hair follicle outer root sheath keratinocytes.

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    Yan, Li; Cao, Rui; Wang, Lianzhao; Liu, Yuanbo; Pan, Bo; Yin, Yanhua; Lv, Xiaoyan; Zhuang, Qiang; Sun, Xuejian; Xiao, Ran

    2015-01-01

    Keloid is a skin fibrotic disease with the characteristics of recurrence and invasion, its pathogenesis still remains unrevealed. The epithelial-mesenchymal transition (EMT) is critical for wound healing, fibrosis, recurrence, and invasion of cancer. We sought to investigate the EMT in keloid and the mechanism through which the EMT regulates keloid formation. In keloid tissues, the expressions of EMT-associated markers and transforming growth factor (TGF)-β1/Smad3 signaling were examined by immunohistochemistry. In the keloid epidermis and dermal tissue, the expressions of genes related to the regulation of skin homeostasis, fibroblast growth factor receptor 2 (FGFR2) and p63, were analyzed using quantitative real-time polymerase chain reaction. The results showed that accompanying the loss of the epithelial marker E-cadherin and the gain of the mesenchymal markers fibroblast-specific protein 1 (FSP1) and vimentin in epithelial cells from epidermis and skin appendages, and in endothelial cells from dermal microvessels, enhanced TGF-β1 expression and Smad3 phosphorylation were noted in keloid tissues. Moreover, alternative splicing of the FGFR2 gene switched the predominantly expressed isoform from FGFR2-IIIb to -IIIc, concomitant with the decreased expression of ΔNp63 and TAp63, which changes might partially account for abnormal epidermis and appendages in keloids. In addition, we found that TGF-β1-induced hair follicle outer root sheath keratinocytes (ORSKs) and normal skin epithelial cells underwent EMT in vitro with ORSKs exhibiting more obvious EMT changes and more similar expression profiles for EMT-associated and skin homeostasis-related genes as in keloid tissues, suggesting that ORSKs might play crucial roles in the EMT in keloids. Our study provided insights into the molecular mechanisms mediating the EMT pathogenesis of keloids.

  19. Transketolase Serves a Poor Prognosticator in Esophageal Cancer by Promoting Cell Invasion via Epithelial-Mesenchymal Transition

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    Chao, Yin-Kai; Peng, Ta-Lun; Chuang, Wen-Yu; Yeh, Chi-Ju; Li, Yan-Liang; Lu, Ya-Ching; Cheng, Ann-Joy

    2016-01-01

    Background: To characterize the potential function and clinical significance of Transketolase (TKT) in esophageal cancer. Methods: High invasive esophageal squamous cell carcinoma (ESCC) cell line CE48T/VGH was used. Cellular functions in response to TKT modulation were examined, including cell growth, migration and invasion. The underlying molecules involved in the TKT regulatory mechanism were determined by western blot and confocal microscopic analysis. Clinically, TKT expressions in 76 ESCC patients were assessed by immunohistochemical (IHC) method, and the association with treatment outcome was determined. Results: TKT silencing inhibited cell migration and invasion but had a minimal effect on cell growth. This TKT silencing also induced the reversion of epithelial-mesenchymal transition (EMT), as evidenced by the spindle to cuboidal morphological change, increased the expression of epithelial markers (γ-catenin), and decreased the levels of mesenchymal markers (fibronectin and N-cadherin). Mechanically, TKT was shown to modulate the EMT through the pERK-Slug/Snail-associated signaling pathway. Clinically, a high level of TKT in the cancer tissues of patients with esophageal squamous cell carcinoma was associated with poor survival (P = 0.042). In the multivariate analysis, a high TKT level was also shown to be an independent unfavorable prognostic factor (Odds ratio: 1.827, 95% confidence interval: 1.045-3.196, P = 0.035). Conclusions: TKT contributes to esophageal cancer by promoting cell invasion via meditating EMT process. Clinically, the over-expression of TKT in ESCC patients predicts poorer survival. TKT inhibition may be a useful strategy to intervene in cancer cell invasion and metastasis, which may lead to better prognosis for ESCC patients. PMID:27698919

  20. Cytokeratin 18 is necessary for initiation of TGF-β1-induced epithelial-mesenchymal transition in breast epithelial cells.

    Science.gov (United States)

    Jung, Hyejung; Kim, Bomin; Moon, Byung In; Oh, Eok-Soo

    2016-12-01

    During epithelial-mesenchymal transition (EMT), epithelial cells lose key phenotypic markers (e.g., E-cadherin and cytokeratin 18) and acquire mesenchymal markers (e.g., N-cadherin and vimentin). Although the loss of cytokeratin 18 is a hallmark of EMT, the regulatory role of cytokeratin 18 in EMT is not yet fully understood. Here, we report that cytokeratin 18 is involved in the regulation of transforming growth factor-beta1 (TGF-β1)-induced EMT in breast epithelial cells. When MCF10A cells were treated with TGF-β1 for 24 h, considerable morphological changes, indicative of the early stages of EMT (e.g., loss of cell-cell contact), were observed and cytokeratin 18 was downregulated. However, E-cadherin levels were not altered until a later time point. This suggests that cytokeratin 18 may play an active role during the earlier stages of EMT. Consistent with this notion, siRNA-mediated knockdown of cytokeratin 18 delayed TGF-β1-mediated EMT, and the associated downregulation of E-cadherin reduced the phosphorylation/nuclear localization of smad 2/3 and decreased the expression levels of snail and slug (which inhibit E-cadherin expression in epithelial cells as an early response to TGF-β1). Taken together, these results suggest that cytokeratin 18 critically contributes to initiating TGF-β1-induced EMT via the smad 2/3-mediated regulation of snail and slug expression in breast epithelial cells.

  1. α-Solanine Inhibits Invasion of Human Prostate Cancer Cell by Suppressing Epithelial-Mesenchymal Transition and MMPs Expression

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    Kun-Hung Shen

    2014-08-01

    Full Text Available α-Solanine, a naturally occurring steroidal glycoalkaloid found in nightshade (Solanum nigrum Linn., was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in suppression of cancer cell metastasis by α-solanine remains unclear. This study investigates the suppression mechanism of α-solanine on motility of the human prostate cancer cell PC-3. Results show that α-solanine reduces the viability of PC-3 cells. When treated with non-toxic doses of α-solanine, cell invasion is markedly suppressed by α-solanine. α-Solanine also significantly elevates epithelial marker E-cadherin expression, while it concomitantly decreases mesenchymal marker vimentin expression, suggesting it suppresses epithelial-mesenchymal transition (EMT. α-Solanine reduces the mRNA level of matrix metalloproteinase-2 (MMP-2, MMP-9 and extracellular inducer of matrix metalloproteinase (EMMPRIN, but increases the expression of reversion-inducing cysteine-rich protein with kazal motifs (RECK, and tissue inhibitor of metalloproteinase-1 (TIMP-1 and TIMP-2. Immunoblotting assays indicate α-solanine is effective in suppressing the phosphorylation of phosphatidylinositide-3 kinase (PI3K, Akt and ERK. Moreover, α-solanine downregulates oncogenic microRNA-21 (miR-21 and upregulates tumor suppressor miR-138 expression. Taken together, the results suggest that inhibition of PC-3 cell invasion by α-solanine may be, at least in part, through blocking EMT and MMPs expression. α-Solanine also reduces ERK and PI3K/Akt signaling pathways and regulates expression of miR-21 and miR-138. These findings suggest an attractive therapeutic potential of α-solanine for suppressing invasion of prostate cancer cell.

  2. Epithelial mesenchymal transition (EMT) and non-small cell lung cancer (NSCLC): a mutual association with airway disease.

    Science.gov (United States)

    Mahmood, Malik Quasir; Ward, Chris; Muller, Hans Konrad; Sohal, Sukhwinder Singh; Walters, Eugene Haydn

    2017-03-01

    NSCLC is a leading cause of morbidity and mortality worldwide. It includes adeno- and squamous cell carcinoma. In the background, COPD and smoking play a vital role in development of NSCLC. Local progression and metastasis of NSCLC has been associated with various mechanisms, but in particular by a process called epithelial mesenchymal transition (EMT), which is implicated in COPD pathogenesis. In this study, we have investigated whether expression of EGFR (activation marker) and S100A4, vimentin and N-cadherin (as EMT) is different both in central and leading edge of NSCLC and to what extent related to EMT activity of both small and large airways, stage and differentiation of NSCLC. We have investigated EMT biomarkers (S100A4, vimentin, and N-cadherin), an epithelial activation marker (EGFR) and a vascularity marker (Type-IV collagen) in surgically resected tissue from patients with NSCLC (adeno- and squamous cell carcinoma), and compared them with expression in the corresponding non-tumorous airways. EGFR, S100A4, vimentin, N-cadherin expression was higher in tumor cells located at the peripheral leading edge of NSCLC when compared with centrally located tumor cells of same subjects (P EMT markers in the leading edge were significantly related to airway EMT activity, while peripheral edge vascularity of squamous cell carcinoma only was significantly related to large airway Rbm vascularity (P EMT-related protein expression was markedly high in the peripheral leading edge of NSCLCs and related to tumor characteristics associated with poor prognosis. The relationships between EMT-related tumor biomarker expression and those in the airway epithelium and Rbm provide a background for utility of airway changes in clinical settings.

  3. Effect of artemisinin combined with cisplatin intervention on epithelial-mesenchymal transition, angiogenesis and ATP generation in MGC-803 gastric cancer cell lines

    Institute of Scientific and Technical Information of China (English)

    Meng-Xuan Wu

    2016-01-01

    Objective:To study the effect of artemisinin combined with cisplatin intervention on epithelial-mesenchymal transition, angiogenesis and ATP generation in MGC-803 gastric cancer cell lines.Methods: MGC-803 gastric cancer cell lines were cultured and divided into control group, cisplatin group, artemisinin group, cisplatin + artemisinin group, cisplatin + artemisinin + NAC group and NAC group, and after different conditions of treatment, cell viability, apoptosis rate as well as levels of epithelial-mesenchymal transition molecules, angiogenesis molecules and ATP were determined.Results: Cell viability, ATP generation as well as VEGFA, VEGFB, VEGFC, N-cadherin and Vimentin levels of cisplatin group, artemisinin group and cisplatin + artemisinin group were lower than those of control group, and early apoptosis rate, late apoptosis rate and E-cadherin levels were higher than those of control group; cell viability, ATP generation as well as VEGFA, VEGFB, VEGFC, N-cadherin and Vimentin levels of cisplatin + artemisinin group were lower than those of cisplatin group and artemisinin group, and early apoptosis rate, late apoptosis rate and E-cadherin level were higher than those of cisplatin group and artemisinin group. E-cadherin level of cisplatin + artemisinin + NAC group was lower than that of cisplatin + artemisinin group, and ATP generation as well as VEGFA, VEGFB, VEGFC, N-cadherin and Vimentin levels were higher than those of cisplatin + artemisinin group.Conclusion:Artemisinin combined with cisplatin intervention can synergistically exert the inhibitory effect on epithelial-mesenchymal transition, angiogenesis and ATP generation in MGC-803 gastric cancer cell lines, and the inhibiting effect is partially realized by increasing the generation of reactive oxygen species.

  4. miR-144 functions as a tumor suppressor in breast cancer through inhibiting ZEB1/2-mediated epithelial mesenchymal transition process

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    Pan Y

    2016-10-01

    Full Text Available Yuliang Pan,1,2 Jun Zhang,1 Huiqun Fu,1 Liangfang Shen2 1Department of Oncology, The Third Xiangya Hospital of Central South University, Changsha, Hunan, People’s Republic of China; 2Department of Oncology Radiotherapy, Xiangya Hospital of Central South University, Changsha, Hunan, People’s Republic of China Abstract: Breast cancer is the most common cancer in women worldwide. Local invasion, metastasis, and chemotherapy resistance are the obstacles for treatment of breast cancer. In this study, we aim to investigate the role of miR-144 in breast cancer. We demonstrate that the expression of miR-144 is downregulated in breast cancer and cell lines, and lower miR-144 expression is associated with poor differentiation, higher clinical stage, and lymph node metastasis in patients with breast cancer. The rescue of miR-144 expression is able to inhibit the cell proliferation and the ability of cell migration and invasion. In addition, we show that miR-144 can directly target at 3'-untranslation region of zinc finger E-box-binding homeobox 1 and 2, that is, ZEB1 and ZEB2, and regulate their expression at transcriptional and translational levels. Moreover, we also demonstrate that ectopic expression of miR-144 can inhibit the process of epithelial mesenchymal transition in MCF-7 and MDA-MB-231 cells. Thus, we here demonstrate that miR-144 functions as a tumor suppressor in breast cancer at least partly through inhibiting ZEB1/2-mediated epithelial mesenchymal transition process. Our findings indicate that the miR-144-ZEB1/2 signaling could represent a promising therapeutic target for breast cancer treatment. Keywords: breast cancer, miR-144, ZEB1, ZEB2, epithelial mesenchymal transition

  5. Benzyl Isothiocyanate Inhibits Epithelial-Mesenchymal Transition in Cultured and Xenografted Human Breast Cancer Cells

    OpenAIRE

    Sehrawat, Anuradha; Singh, Shivendra V.

    2011-01-01

    We showed previously that cruciferous vegetable constituent benzyl isothiocyanate (BITC) inhibits growth of cultured and xenografted human breast cancer cells, and suppresses mammary cancer development in a transgenic mouse model. We now demonstrate, for the first time, that BITC inhibits epithelial-to-mesenchymal transition (EMT) in human breast cancer cells. Exposure of estrogen-independent MDA-MB-231 and estrogen-responsive MCF-7 human breast cancer cell lines and a pancreatic cancer cell ...

  6. Translating epithelial mesenchymal transition markers into the clinic: Novel insights from proteomics

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    Vergara Daniele

    2016-03-01

    Full Text Available The growing understanding of the molecular mechanisms underlying epithelial-to-mesenchymal transition (EMT may represent a potential source of clinical markers. Despite EMT drivers have not yet emerged as candidate markers in the clinical setting, their association with established clinical markers may improve their specificity and sensitivity. Mass spectrometry-based platforms allow analyzing multiple samples for the expression of EMT candidate markers, and may help to diagnose diseases or monitor treatment efficiently. This review highlights proteomic approaches applied to elucidate the differences between epithelial and mesenchymal tumors and describes how these can be used for target discovery and validation.

  7. Cytoskeletal re-arrangement in TGF-β1-induced alveolar epithelial-mesenchymal transition studied by atomic force microscopy and high-content analysis.

    Science.gov (United States)

    Buckley, Stephen T; Medina, Carlos; Davies, Anthony M; Ehrhardt, Carsten

    2012-04-01

    Epithelial-mesenchymal transition (EMT) is closely implicated in the pathogenesis of idiopathic pulmonary fibrosis. Associated with this phenotypic transition is the acquisition of an elongated cell morphology and establishment of stress fibers. The extent to which these EMT-associated changes influence cellular mechanics is unclear. We assessed the biomechanical properties of alveolar epithelial cells (A549) following exposure to TGF-β1. Using atomic force microscopy, changes in cell stiffness and surface membrane features were determined. Stimulation with TGF-β1 gave rise to a significant increase in stiffness, which was augmented by a collagen I matrix. Additionally, TGF-β1-treated cells exhibited a rougher surface profile with notable protrusions. Simultaneous quantitative examination of the morphological attributes of stimulated cells using an image-based high-content analysis system revealed dramatic alterations in cell shape, F-actin content and distribution. Together, these investigations point to a strong correlation between the cytoskeletal-associated cellular architecture and the mechanical dynamics of alveolar epithelial cells undergoing EMT. From the Clinical Editor: Epithelial-mesenchymal transition is implicated in the pathogenesis of pulmonary fibrosis. Using atomic force microscopy, the authors demonstrate a strong correlation between the cytoskeletal-associated cellular architecture and the mechanical dynamics of alveolar epithelial cells undergoing mesenchymal transition.

  8. The ubiquitin–proteasome system and signal transduction pathways regulating Epithelial Mesenchymal transition of cancer

    Directory of Open Access Journals (Sweden)

    Voutsadakis Ioannis A

    2012-07-01

    Full Text Available Abstract Epithelial to Mesenchymal transition (EMT in cancer, a process permitting cancer cells to become mobile and metastatic, has a signaling hardwire forged from development. Multiple signaling pathways that regulate carcinogenesis enabling characteristics in neoplastic cells such as proliferation, resistance to apoptosis and angiogenesis are also the main players in EMT. These pathways, as almost all cellular processes, are in their turn regulated by ubiquitination and the Ubiquitin-Proteasome System (UPS. Ubiquitination is the covalent link of target proteins with the small protein ubiquitin and serves as a signal to target protein degradation by the proteasome or to other outcomes such as endocytosis, degradation by the lysosome or specification of cellular localization. This paper reviews signal transduction pathways regulating EMT and being regulated by ubiquitination.

  9. Upregulation of TrkB promotes epithelial-mesenchymal transition and anoikis resistance in endometrial carcinoma.

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    Wei Bao

    Full Text Available Mechanisms governing the metastasis of endometrial carcinoma (EC are poorly defined. Recent data support a role for the cell surface receptor tyrosine kinase TrkB in the progression of several human tumors. Here we present evidence for a direct role of TrkB in human EC. Immunohistochemical analysis revealed that TrkB and its secreted ligand, brain-derived neurotrophic factor (BDNF, are more highly expressed in EC than in normal endometrium. High TrkB levels correlated with lymph node metastasis (p<0.05 and lymphovascular space involvement (p<0.05 in EC. Depletion of TrkB by stable shRNA-mediated knockdown decreased the migratory and invasive capacity of cancer cell lines in vitro and resulted in anoikis in suspended cells. Conversely, exogenous expression of TrkB increased cell migration and invasion and promoted anoikis resistance in suspension culture. Furthermore, over-expression of TrkB or stimulation by BDNF resulted in altered the expression of molecular mediators of the epithelial-to-mesenchymal transition (EMT. RNA interference (RNAi-mediated depletion of the downstream regulator, Twist, blocked TrkB-induced EMT-like transformation. The use of in vivo models revealed decreased peritoneal dissemination in TrkB-depleted EC cells. Additionally, TrkB-depleted EC cells underwent mesenchymal-to-epithelial transition and anoikis in vivo. Our data support a novel function for TrkB in promoting EMT and resistance to anoikis. Thus, TrkB may constitute a potential therapeutic target in human EC.

  10. Curcumin Suppresses Intestinal Fibrosis by Inhibition of PPARγ-Mediated Epithelial-Mesenchymal Transition

    Science.gov (United States)

    Jiang, Bin; Wang, Hui; Shen, Cunsi; Chen, Hao

    2017-01-01

    Intestinal fibrotic stricture is a major complication of Crohn's disease (CD) and epithelial-to-mesenchymal transition (EMT) is considered as an important contributor to the formation of intestinal fibrosis by increasing extracellular matrix (ECM) proteins. Curcumin, a compound derived from rhizomes of Curcuma, has been demonstrated with a potent antifibrotic effect. However, its effect on intestinal fibrosis and the potential mechanism is not completely understood. Here we found that curcumin pretreatment significantly represses TGF-β1-induced Smad pathway and decreases its downstream α-smooth muscle actin (α-SMA) gene expression in intestinal epithelial cells (IEC-6); in contrast, curcumin increases expression of E-cadherin and peroxisome proliferator-activated receptor γ (PPARγ) in IEC-6. Moreover, curcumin promotes nuclear translocation of PPARγ and the inhibitory effect of curcumin on EMT could be reversed by PPARγ antagonist GW9662. Consistently, in the rat model of intestinal fibrosis induced by 2,4,5-trinitrobenzene sulphonic acid (TNBS), oral curcumin attenuates intestinal fibrosis by increasing the expression of PPARγ and E-cadherin and decreasing the expression of α-SMA, FN, and CTGF in colon tissue. Collectively, these results indicated that curcumin is able to prevent EMT progress in intestinal fibrosis by PPARγ-mediated repression of TGF-β1/Smad pathway. PMID:28203261

  11. Notch promotes epithelial-mesenchymal transition during cardiac development and oncogenic transformation

    Science.gov (United States)

    Timmerman, Luika A.; Grego-Bessa, Joaquín; Raya, Angel; Bertrán, Esther; Pérez-Pomares, José María; Díez, Juan; Aranda, Sergi; Palomo, Sergio; McCormick, Frank; Izpisúa-Belmonte, Juan Carlos; de la Pompa, José Luis

    2004-01-01

    Epithelial-to-mesenchymal transition (EMT) is fundamental to both embryogenesis and tumor metastasis. The Notch intercellular signaling pathway regulates cell fate determination throughout metazoan evolution, and overexpression of activating alleles is oncogenic in mammals. Here we demonstrate that Notch activity promotes EMT during both cardiac development and oncogenic transformation via transcriptional induction of the Snail repressor, a potent and evolutionarily conserved mediator of EMT in many tissues and tumor types. In the embryonic heart, Notch functions via lateral induction to promote a selective transforming growth factor-β (TGFβ)-mediated EMT that leads to cellularization of developing cardiac valvular primordia. Embryos that lack Notch signaling elements exhibit severely attenuated cardiac snail expression, abnormal maintenance of intercellular endocardial adhesion complexes, and abortive endocardial EMT in vivo and in vitro. Accordingly, transient ectopic expression of activated Notch1 (N1IC) in zebrafish embryos leads to hypercellular cardiac valves, whereas Notch inhibition prevents valve development. Overexpression of N1IC in immortalized endothelial cells in vitro induces EMT accompanied by oncogenic transformation, with corresponding induction of snail and repression of VE-cadherin expression. Notch is expressed in embryonic regions where EMT occurs, suggesting an intimate and fundamental role for Notch, which may be reactivated during tumor metastasis. PMID:14701881

  12. Cten promotes epithelial-mesenchymal transition through the post-transcriptional stabilization of Snail.

    Science.gov (United States)

    Thorpe, Hannah; Asiri, Abdulaziz; Akhlaq, Maham; Ilyas, Mohammad

    2017-07-10

    Cten promotes cell migration however the knowledge of underlying signalling pathways is sparse. We have shown that Cten downregulates E-cadherin, a feature of epithelial to mesenchymal transition (EMT). This prompted us to investigate whether Cten further contributed to EMT processes to regulate cell motility. The regulation of Snail by Cten was investigated following overexpression, knockdown (by RNA-interference) or knockout of Cten in HCT116, Caco-2 and SW620 colorectal cancer (CRC) cell lines. Subsequently, the cycloheximide (CHX) pulse chase assay was used to investigate changes in Snail protein stability and the functional relevance of Cten-Snail signalling was investigated. Snail was identified as a downstream target of Cten signalling using multiple approaches of Cten expression manipulation. Furthermore, this activity was mediated through the SH2 domain of Cten. The CHX assay confirmed that Cten was regulating Snail at a post transcriptional level and this was through the prevention of Snail degradation. Cell migration, invasion and colony formation efficiency were increased following forced expression of GFP-Cten but subsequently lost when Snail was knocked down, demonstrating a functional Cten-Snail signalling axis. In conclusion, we have described a novel Cten-Snail signaling pathway that contributes to cell motility in CRC, mediated by the stabilization of Snail protein. This finding potentially furthers the understanding of EMT regulatory networks in cancer metastasis. © 2017 Wiley Periodicals, Inc.

  13. Hepatitis C virus NS4B protein induces epithelial-mesenchymal transition by upregulation of Snail.

    Science.gov (United States)

    Hu, Bicheng; Xie, Shenggao; Hu, Yuqian; Chen, Wen; Chen, Xiaofan; Zheng, Yi; Wu, Xinxing

    2017-04-21

    Chronic hepatitis C virus (HCV) infection is an important cause of hepatocellular carcinoma (HCC). Epithelial to mesenchymal transition (EMT) is a key process associated with tumor metastasis and poor prognosis. HCV infection, HCV core and NS5A protein could induce EMT process, but the role of NS4B on EMT remains poorly understood. We overexpressed HCV NS4B protein in HepG2 cells or Huh7.5.1 cells infected by HCVcc, the E-cadherin expression, N-cadherin expression and the EMT-associated transcriptional factor Snail were determined. The migration and invasion capabilities of the transfected cells were evaluated using wound-healing assay. Additionally, we used Snail siRNA interference to confirm the relation of HCV NS4B and Snail on EMT promotion. HCV NS4B increased the expression of EMT related markers and promoted cell migration and invasion. Snail knock-down almost completely eliminated the function of NS4B protein in EMT changes and reversed cell migration capacity to lower level. HCV NS4B protein could reduce the expression of Scribble and Hippo signal pathway were subsequently inactivated, resulting in the activation of PI3K/AKT pathway, which may be the reason for the up-regulation of Snail. This study demonstrates that HCV NS4B protein induces EMT progression via the upregulation of Snail in HCC, which may be a novel underlying mechanism for HCV-associated HCC development, invasion and metastasis.

  14. Epithelial-mesenchymal transition in colorectal cancer metastasis: A system review.

    Science.gov (United States)

    Cao, Hui; Xu, Enping; Liu, Hong; Wan, Ledong; Lai, Maode

    2015-08-01

    Tumor metastasis is a multi-step process by which tumor cells disseminate from their primary site and form secondary tumors at a distant site. And metastasis is the major cause of death in the vast majority of cancer patients. However, the mechanisms underlying each step remain obscure. In the past decade, a developmental program epithelial-to-mesenchymal transition (EMT) has been increasingly recognized to play pivotal and intricate roles in promoting carcinoma invasion and metastasis. The EMT process is very complex and controlled by various families of transcriptional regulators through different signaling pathways. In this system review, we focus on the molecular network of the EMT program and its malignant phenotypes associated with metastasis in colorectal cancer (CRC), including cancer stem cells, tumor budding, circulating tumor cells and drug resistance. A better understanding of the molecular regulation of the dynamic EMT program during tumor metastasis will help to provide much-needed therapeutic interventions to target this program when treating metastatic CRC.

  15. LOXL2 drives epithelial-mesenchymal transition via activation of IRE1-XBP1 signalling pathway

    Science.gov (United States)

    Cuevas, Eva P.; Eraso, Pilar; Mazón, María J.; Santos, Vanesa; Moreno-Bueno, Gema; Cano, Amparo; Portillo, Francisco

    2017-01-01

    Epithelial-to-Mesenchymal Transition (EMT) is a key process contributing to the aggressiveness of cancer cells. EMT is triggered by activation of different transcription factors collectively known as EMT-TFs. Different cellular cues and cell signalling networks activate EMT at transcriptional and posttranscriptional level in different biological and pathological situations. Among them, overexpression of LOXL2 (lysyl oxidase-like 2) induces EMT independent of its catalytic activity. Remarkably, perinuclear/cytoplasmic accumulation of LOXL2 is a poor prognosis marker of squamous cell carcinomas and is associated to basal breast cancer metastasis by mechanisms no yet fully understood. Here, we report that overexpression of LOXL2 promotes its accumulation in the Endoplasmic Reticulum where it interacts with HSPA5 leading to activation of the IRE1-XBP1 signalling pathway of the ER-stress response. LOXL2-dependent IRE1-XBP1 activation induces the expression of several EMT-TFs: SNAI1, SNAI2, ZEB2 and TCF3 that are direct transcriptional targets of XBP1. Remarkably, inhibition of IRE1 blocks LOXL2-dependent upregulation of EMT-TFs thus hindering EMT induction. PMID:28332555

  16. The contribution of TGF-β in Epithelial-Mesenchymal Transition (EMT): Down-regulation of E-cadherin via snail.

    Science.gov (United States)

    Yu, H; Shen, Y; Hong, J; Xia, Q; Zhou, F; Liu, X

    2015-01-01

    TGF-β plays a central role in epithelial-mesenchymal transition (EMT), which is a highly conserved and reversible process that governs tumor development, invasion and metastasis. Through this transition, the epithelial cell acquires a migratory behavior which allows it to move away from the cell community and to integrate into the surrounding tissue. The cells lost epithelial phenotypes including the change of cell polarity and the loss of specialized cell-cell contacts, which related with the shortage of E-cadherin directly. The increasing reports indicated that TGF-β down-regulated the expression of E-cadherin through snail signaling pathway, which played an important role in the development of EMT. In this review, we summarized the contribution of TGF-β in EMT and discussed the molecular mechanism of snail signaling participating in the regulation of E-cadherin triggered by TGF-β. TGF-β, Epithelial-mesenchymal-transition (EMT), snail family, E-cadherin.

  17. Paeoniflorin suppresses TGF-β mediated epithelial-mesenchymal transition in pulmonary fibrosis through a Smad-dependent pathway

    Science.gov (United States)

    Ji, Yu; Dou, Yan-nong; Zhao, Qian-wen; Zhang, Ji-zhou; Yang, Yan; Wang, Ting; Xia, Yu-feng; Dai, Yue; Wei, Zhi-feng

    2016-01-01

    Aim: Paeoniflorin has shown to attenuate bleomycin-induced pulmonary fibrosis (PF) in mice. Because the epithelial-mesenchymal transition (EMT) in type 2 lung endothelial cells contributes to excessive fibroblasts and myofibroblasts during multiple fibrosis of tissues, we investigated the effects of paeoniflorin on TGF-β mediated pulmonary EMT in bleomycin-induced PF mice. Methods: PF was induced in mice by intratracheal instillation of bleomycin (5 mg/kg). The mice were orally treated with paeoniflorin or prednisone for 21 d. After the mice were sacrificed, lung tissues were collected for analysis. An in vitro EMT model was established in alveolar epithelial cells (A549 cells) incubated with TGF-β1 (2 ng/mL). EMT identification and the expression of related proteins were performed using immunohistochemistry, transwell assay, ELISA, Western blot and RT-qPCR. Results: In PF mice, paeoniflorin (50, 100 mg·kg−1·d−1) or prednisone (6 mg·kg−1·d−1) significantly decreased the expression of FSP-1 and α-SMA, and increased the expression of E-cadherin in lung tissues. In A549 cells, TGF-β1 stimulation induced EMT, as shown by the changes in cell morphology, the increased cell migration, and the increased vimentin and α-SMA expression as well as type I and type III collagen levels, and by the decreased E-cadherin expression. In contrast, effects of paeoniflorin on EMT disappeared when the A549 cells were pretreated with TGF-β1 for 24 h. TGF-β1 stimulation markedly increased the expression of Snail and activated Smad2/3, Akt, ERK, JNK and p38 MAPK in A549 cells. Co-incubation with paeoniflorin (1–30 μmol/L) dose-dependently attenuated TGF-β1-induced expression of Snail and activation of Smad2/3, but slightly affected TGF-β1-induced activation of Akt, ERK, JNK and p38 MAPK. Moreover, paeoniflorin markedly increased Smad7 level, and decreased ALK5 level in A549 cells. Conclusion: Paeoniflorin suppresses the early stages of TGF-β mediated EMT in alveolar

  18. Evidence for epithelial-mesenchymal transition in cancer stem cells of head and neck squamous cell carcinoma.

    Directory of Open Access Journals (Sweden)

    Chao Chen

    Full Text Available Initiation, growth, recurrence, and metastasis of head and neck squamous cell carcinomas (HNSCC have been related to the behavior of cancer stem cells (CSC that can be identified by their aldehyde-dehydrogenase-isoform-1 (ALDH1 activity. We quantified and enriched ALDH1(+ cells within HNSCC cell lines and subsequently characterized their phenotypical and functional properties like invasion capacity and epithelial-mesenchymal transition (EMT. Spheroid culture enriched CSC from five HNSCC cell lines by up to 5-fold. In spheroid-derived cells (SDC and the parental monolayer-derived cell line ALDH1, CD44, CD24, E-Cadherin, α-SMA, and Vimentin expression was compared by flow-cytometry and immunofluorescence together with proliferation and cell cycle analysis. Invasion activity was evaluated by Matrigel assay and expression of stemness-related transcription factors (TF Nanog, Oct3/4, Sox2 and EMT-related genes Snail1 and 2, and Twist by real-time PCR. All cell lines formed spheroids that could self-renew and be serially re-passaged. ALDH1 expression was significantly higher in SDC. ALDH1(+ cells showed increased colony-formation. The proportion of cells with a putative CSC marker constellation of CD44(+/CD24(- was highly variable (0.5% to 96% in monolayer and spheroid cultures and overlapped in 0%-33% with the CD44(+/CD24(-/ALDH1(+ cell subset. SDC had significantly higher invading activity. mRNA of the stemness-related genes Sox2, Nanog, and Oct3/4 was significantly increased in SDC of all cell lines. Twist was significantly increased in two while Snail2 showed a significant increase in one and a significant decrease in SDC of two cell lines. SDC had a higher G0 phase proportion, showed high-level expression of α-SMA and Vimentin, but significantly decreased E-Cadherin expression. HNSCC-lines harbor potential CSC, characterized by ALDH1 and stemness marker TF expression as well as properties like invasiveness, quiescence, and EMT. CSC can be

  19. aPKC-ι/P-Sp1/Snail signaling induces epithelial-mesenchymal transition and immunosuppression in cholangiocarcinoma.

    Science.gov (United States)

    Qian, Yawei; Yao, Wei; Yang, Tao; Yang, Yan; Liu, Yan; Shen, Qi; Zhang, Jian; Qi, Weipeng; Wang, Jianming

    2017-10-01

    Cholangiocarcinoma (CCA) is a highly malignant bile duct cancer that tends to invade and metastasize early. The epithelial-mesenchymal transition (EMT) has been implicated in cancer cell invasion and metastasis, as well as in cancer cell evasion of host immunity. In this study, we investigated the interaction between atypical protein kinase C-iota (aPKC-ι) and Snail in the regulation of EMT and its relationship to CCA immunosuppression. Our results demonstrated that aPKC-ι, Snail, and infiltrated immunosuppressive cells were significantly up-regulated in CCA tumor tissues and linked to poor prognosis. aPKC-ι induced EMT and immunosuppression by regulating Snail in vitro and in vivo, although aPKC-ι did not directly interact with Snail in coimmunoprecipitation experiments. To further clarify the molecular interaction between aPKC-ι and Snail in relation to EMT, quantitative iTRAQ-based phosphoproteomic analysis and liquid chromatography-tandem mass spectrometry were conducted to identify the substrates of aPKC-ι-dependent phosphorylation. Combined with coimmunoprecipitation, we showed that specificity protein 1 (Sp1) was directly phosphorylated by aPKC-ι on Ser59 (P-Sp1). Both Sp1 and P-Sp1 were up-regulated in CCA tumor tissues and associated with clinicopathological features and poor prognosis in CCA patients. Moreover, using chromatin immunoprecipitation assays, we found that P-Sp1 regulated Snail expression by increasing Sp1 binding to the Snail promoter. P-Sp1 also regulated aPKC-ι/Snail-induced EMT-like changes and immunosuppression in CCA cells. Our findings further indicated that CCA cells with EMT-like features appear to generate immunosuppressive natural T regulatory-like cluster of differentiation 4-positive (CD4(+) )CD25(-) cells rather than to increase CD4(+) CD25(+) natural T regulatory cells, in part by mediating T regulatory-inducible cytokines such as transforming growth factor β1 and interleukin 2. These results demonstrate that a

  20. Tanshinone IIA Inhibits Epithelial-Mesenchymal Transition in Bladder Cancer Cells via Modulation of STAT3-CCL2 Signaling

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    Sung-Ying Huang

    2017-07-01

    Full Text Available Tanshinone IIA (Tan-IIA is an extract from the widely used traditional Chinese medicine (TCM Danshen (Salvia miltiorrhiza, and has been found to attenuate the proliferation of bladder cancer (BCa cells (The IC50 were: 5637, 2.6 μg/mL; BFTC, 2 μg/mL; T24, 2.7 μg/mL, respectively.. However, the mechanism of the effect of Tan-IIA on migration inhibition of BCa cells remains unclear. This study investigates the anti-metastatic effect of Tan-IIA in human BCa cells and clarifies its molecular mechanism. Three human BCa cell lines, 5637, BFTC and T24, were used for subsequent experiments. Cell migration and invasion were evaluated by transwell assays. Real-time RT-PCR and western blotting were performed to detect epithelial-mesenchymal transition (EMT-related gene expression. The enzymatic activity of matrix metalloproteinases (MMP was evaluated by zymography assay. Tan-IIA inhibited the migration and invasion of human BCa cells. Tan-IIA suppressed both the protein expression and enzymatic activity of MMP-9/-2 in human BCa cells. Tan-IIA up-regulated the epithelial marker E-cadherin and down-regulated mesenchymal markers such as N-cadherin and Vimentin, along with transcription regulators such as Snail and Slug in BCa cells in a time- and dose-dependent manner. Mechanism dissection revealed that Tan-IIA-inhibited BCa cell invasion could function via suppressed chemokine (C-C motif ligand 2 (CCL2 expression, which could be reversed by the addition of CCL2 recombinant protein. Furthermore, Tan-IIA could inhibit the phosphorylation of the signal transducer and activator of transcription 3 (STAT3 (Tyr705, which cannot be restored by the CCL2 recombinant protein addition. These data implicated that Tan-IIA might suppress EMT on BCa cells through STAT3-CCL2 signaling inhibition. Tan-IIA inhibits EMT of BCa cells via modulation of STAT3-CCL2 signaling. Our findings suggest that Tan-IIA can serve as a potential anti-metastatic agent in BCa therapy.

  1. Epithelial-mesenchymal transition (EMT) phenotype at invasion front of squamous cell carcinoma of the penis influences oncological outcomes.

    Science.gov (United States)

    da Cunha, Isabela Werneck; Souza, Maria José L; da Costa, Walter Henriques; Amâncio, Alice M; Fonseca, Francisco Paulo; Zequi, Stenio de Cassio; Lopes, Ademar; Guimarães, Gustavo Cardoso; Soares, Fernando

    2016-10-01

    Our aims were to evaluate epithelial-mesenchymal transition (EMT) as a useful prognostic marker in penile carcinoma (PC), and establish an objective criterion to define EMT in PC specimens. A total of 149 consecutive cases surgically treated for PC were retrospectively selected. E-cadherin (E-CAD) and vimentin immunohistochemical expressions were evaluated. A combined analysis was performed using both markers to determine EMT status. To establish a normal control to E-CAD expression, we included 14 cases from circumcisions from patients without any neoplastic disease and 77 cases of tumor-free margins. The analyses of tumor samples were evaluated in 2 different areas of the tumor. The first one was in the tumor core. The second analyses were performed on the deepest infiltrative edge of the tumor, nominated invasion front. Cases were classified into EMT absent group, partial EMT group and complete EMT group. Overall survival (OS) and cancer-specific survival (CSS) were analyzed. Kaplan-Meier curves and the log-rank test were used. Cox proportional hazards model was used to determine which variables influenced survival. Tumor specimens presented a significant loss of expression of E-CAD when compared with normal epithelium. Vimentin expression in more than 10% of tumor cells was observed in 50 cases. EMT status was associated with histologic grade, pattern of invasion, lymph node metastasis, and perineural and vascular invasion. Further, 10-year OS and CSS rates in patients with presence and absence of complete EMT status were 38.0% and 55.6%; and 48.0% and 91.9%, respectively. EMT status significantly affected CSS and OS rates even after patients were grouped based on lymph node involvement status. The presence of complete EMT status was associated with both CSS and OS rates. Patients in the complete EMT group had a higher risk of death from cancer (hazard ratio = 7.6, PEMT in PC. We encourage the study of EMT markers in other centers to validate our findings and

  2. SRPX2 Enhances the Epithelial-Mesenchymal Transition and Temozolomide Resistance in Glioblastoma Cells.

    Science.gov (United States)

    Tang, Haitao; Zhao, Jiaxin; Zhang, Liangyu; Zhao, Jiang; Zhuang, Yongzhi; Liang, Peng

    2016-10-01

    Glioblastoma (GBM) is the most common and most aggressive central nervous system tumor in adults. Due to GBM cell invasiveness and resistance to chemotherapy, current medical interventions are not satisfactory, and the prognosis for GBM is poor. It is necessary to investigate the underlying mechanism of GBM metastasis and drug resistance so that more effective treatments can be developed for GBM patients. sushi repeat-containing protein, X-linked 2 (SRPX2) is a prognostic biomarker in many different cancer cell lines and is associated with poor prognosis in cancer patients. SRPX2 overexpression promotes interactions between tumor and endothelial cells, leading to tumor progression and metastasis. We hypothesize that SRPX2 also contributes to GBM chemotherapy resistance and metastasis. Our results revealed that GBM tumor samples from 42 patients expressed higher levels of SRPX2 than the control normal brain tissue samples. High-SRPX2 expression levels are correlated with poor prognosis in those patients, as well as resistance to temozolomide in cultured GBM cells. Up-regulating SRPX2 expression in cultured GBM cell lines facilitated invasiveness and migration of GBM cells, while down-regulating SRPX2 through RNA interference was inhibitory. These results suggest that SRPX2 plays an important role in GBM metastasis. Epithelial to mesenchymal transition (EMT) is one of the processes that facilitate GBM metastasis and resistance to chemotherapy. EMT marker expression was decreased in SRPX2 down-regulated GBM cells, and MAPK signaling pathway marker expression was also decreased when SRPX2 is knocked down in GBM-cultured cells. Blocking the MAPK signaling pathway inhibited GBM metastasis but did not inhibit cell invasion and migration in SRPX2 down-regulated cells. Our results indicate that SRPX2 facilitates GBM metastasis by enhancing the EMT process via the MAPK signaling pathway.

  3. Downregulation of microRNA-145 promotes epithelial-mesenchymal transition via regulating Snail in osteosarcoma.

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    Zhang, Z; Zhang, M; Chen, Qinghan; Zhang, Q

    2017-02-01

    Metastasis is the principal cause of cancer death and occurs through multiple, complex processes. Epithelial to mesenchymal transition (EMT) is an important process during embryonic development and has also been hypothesized to exhibit a significant role in cancer cell invasion and metastasis. MicroRNAs (miRNAs) are a class of widespread noncoding RNAs. In recent years, many studies have shown that miRNAs could influence the signaling pathways and downstream events that define EMT on a molecular level. However, the exact role and mechanisms of miR-145 in EMT of osteosarcoma (OS) was unknown. In the present study, miR-145 was downregulated in OS tissues and cell lines and it was shown that miR-145 expression was closely correlated with advanced tumor progression in patients of OS. In addition, miR-145 upregulation by miR-145 agomir significantly inhibited MG63 cells invasion and migration ability. MiR-145 was reported to be able to inhibit EMT in cancers. Following the examination of changes in cell epithelial and mesenchymal markers, it was found that upregulation of miR-145 strongly reversed EMT in MG63 cells. Meanwhile, the expression of Snail, a strong E-cadherin transcription repressor was also attenuated by miR-145 agomir. Furthermore, the decreased EMT and invasion and metastasis caused by miR-145 agomir could be restored by Snail siRNA. In conclusion, the results demonstrated that miR-145 could mediate EMT by targeting Snail and miR-145 might be a novel EMT regulating transcription factor that involved in the progression of OS. The specific drugs targeting miR-145-mediated EMT process might be new promising cancer therapies.

  4. Sulforaphane inhibits TGF-β-induced epithelial-mesenchymal transition of hepatocellular carcinoma cells via the reactive oxygen species-dependent pathway.

    Science.gov (United States)

    Wu, Jinsheng; Han, Jingli; Hou, Benxin; Deng, Chengwei; Wu, Huanliang; Shen, Liangfang

    2016-05-01

    Sulforaphane is recognized as a safe antitumor agent derived from various cruciferous vegetables, including broccoli. It has been demonstrated that sulforaphase is a potent antitumor agent in diverse cancers. However, its effect on hepatocellular carcinoma remains largely unknown. Here, we show that sulforaphane inhibits TGF-β-induced epithelial-mesenchymal transition of hepatocellular carcinoma cell via the reactive oxygen species-dependent pathway. We found sulforaphane inhibited hepatocellular carcinoma cell proliferation in a dose- and time-dependent manner. Sulforaphane induced G0/G1 phase cell cycle arrest and promoted cell apoptosis. A set of experiments showed that sulforaphase inhibited hepatocellular carcinoma cell migration and invasion, inhibited the formation of fibroblast like mesenchymal cells and the expression of Vimentin, but increased the expression of E-cadherin, suggesting sulforaphane suppresses epithelial-mesenchymal transition (EMT) process. Cotreatment with N-acetyl-L-cysteine inhibited sulforaphane-inhibited invasion and upregulation of E-cadherin and almost completely abolished the sulforaphane-induced expression of Vimentin. The effect of sulforaphane on the growth of hepatocellular carcinoma cells was confirmed by a xenograft tumor growth model. All our finding indicated that sulforaphane is a promising and safe strategy for treating hepatocellular carcinoma.

  5. Twist and YB-1 gene expression in cervical cancer and cervical intraepithelial neoplasia tissue as well as its correlation with epithelial-mesenchymal transition

    Institute of Scientific and Technical Information of China (English)

    Qin Liu; Hong Li; Yu Zhang

    2016-01-01

    Objective:To study the Twist and YB-1 gene expression in cervical cancer and cervical intraepithelial neoplasia tissue as well as its correlation with epithelial-mesenchymal transition. Methods:Normal cervical tissue, cervical intraepithelial neoplasia tissue and cervical cancer tissue were collected for study. ELISA kits were used to detect Twist, YB-1, E-cadherin,β-catenin, N-cadherin and Vimentin contents in cervical tissue, and immunohistochemistry was used to detect Twist and YB-1 expression levels in cervical tissue.Results:Twist and YB-1 contents, cell positive rate and immunohistochemical scores as well as N-cadherin and Vimentin contents in cervical cancer tissue and cervical intraepithelial neoplasia tissue were significantly higher than those in normal cervical tissue while E-cadherin andβ-catenin contents were lower than those in normal cervical tissue; Twist and YB-1 contents, cell positive rate and immunohistochemical scores as well as N-cadherin and Vimentin contents in cervical cancer tissue were significantly higher than those in cervical intraepithelial neoplasia tissue while E-cadherin andβ-catenin contents were lower than those in cervical intraepithelial neoplasia tissue; the higher the Twist and YB-1 expression levels in cervical cancer tissue, the lower the E-cadherin andβ-catenin contents, and the higher the N-cadherin and Vimentin contents.Conclusions: Twist and YB-1 gene overexpression can promote epithelial-mesenchymal transition to be involved in the occurrence of cervical cancer and cervical intraepithelial neoplasia.

  6. Oncogenic potential of histone-variant H2A.Z.1 and its regulatory role in cell cycle and epithelial-mesenchymal transition in liver cancer

    Science.gov (United States)

    Eun, Jung Woo; Shen, Qingyu; Kim, Hyung Seok; Shin, Woo Chan; Ahn, Young Min; Park, Won Sang; Lee, Jung Young; Nam, Suk Woo

    2016-01-01

    H2A.Z is a highly conserved H2A variant, and two distinct H2A.Z isoforms, H2A.Z.1 and H2A.Z.2, have been identified as products of two non-allelic genes, H2AFZ and H2AFV. H2A.Z has been reported to be overexpressed in breast, prostate and bladder cancers, but most studies did not clearly distinguish between isoforms. One recent study reported a unique role for the H2A.Z isoform H2A.Z.2 as a driver of malignant melanoma. Here we first report that H2A.Z.1 plays a pivotal role in the liver tumorigenesis by selectively regulating key molecules in cell cycle and epithelial-mesenchymal transition (EMT). H2AFZ expression was significantly overexpressed in a large cohort of hepatocellular carcinoma (HCC) patients, and high expression of H2AFZ was significantly associated with their poor prognosis. H2A.Z.1 overexpression was demonstrated in a subset of human HCC and cell lines. H2A.Z.1 knockdown suppressed HCC cell growth by transcriptional deregulation of cell cycle proteins and caused apoptotic cell death of HCC cells. We also observed that H2A.Z.1 knockdown reduced the metastatic potential of HCC cells by selectively modulating epithelial-mesenchymal transition regulatory proteins such as E-cadherin and fibronectin. In addition, H2A.Z.1 knockdown reduced the in vivo tumor growth rate in a mouse xenograft model. In conclusion, our findings suggest the oncogenic potential of H2A.Z.1 in liver tumorigenesis and that it plays established role in accelerating cell cycle transition and EMT during hepatocarcinogenesis. This makes H2A.Z.1 a promising target in liver cancer therapy. PMID:26863632

  7. Oncogenic potential of histone-variant H2A.Z.1 and its regulatory role in cell cycle and epithelial-mesenchymal transition in liver cancer.

    Science.gov (United States)

    Yang, Hee Doo; Kim, Pum-Joon; Eun, Jung Woo; Shen, Qingyu; Kim, Hyung Seok; Shin, Woo Chan; Ahn, Young Min; Park, Won Sang; Lee, Jung Young; Nam, Suk Woo

    2016-03-08

    H2A.Z is a highly conserved H2A variant, and two distinct H2A.Z isoforms, H2A.Z.1 and H2A.Z.2, have been identified as products of two non-allelic genes, H2AFZ and H2AFV. H2A.Z has been reported to be overexpressed in breast, prostate and bladder cancers, but most studies did not clearly distinguish between isoforms. One recent study reported a unique role for the H2A.Z isoform H2A.Z.2 as a driver of malignant melanoma. Here we first report that H2A.Z.1 plays a pivotal role in the liver tumorigenesis by selectively regulating key molecules in cell cycle and epithelial-mesenchymal transition (EMT). H2AFZ expression was significantly overexpressed in a large cohort of hepatocellular carcinoma (HCC) patients, and high expression of H2AFZ was significantly associated with their poor prognosis. H2A.Z.1 overexpression was demonstrated in a subset of human HCC and cell lines. H2A.Z.1 knockdown suppressed HCC cell growth by transcriptional deregulation of cell cycle proteins and caused apoptotic cell death of HCC cells. We also observed that H2A.Z.1 knockdown reduced the metastatic potential of HCC cells by selectively modulating epithelial-mesenchymal transition regulatory proteins such as E-cadherin and fibronectin. In addition, H2A.Z.1 knockdown reduced the in vivo tumor growth rate in a mouse xenograft model. In conclusion, our findings suggest the oncogenic potential of H2A.Z.1 in liver tumorigenesis and that it plays established role in accelerating cell cycle transition and EMT during hepatocarcinogenesis. This makes H2A.Z.1 a promising target in liver cancer therapy.

  8. Retraction: "Over-expression of FoxM1 leads to epithelial-mesenchymal transition and cancer stem cell phenotype in pancreatic cancer cells" by Bao et al.

    Science.gov (United States)

    2016-08-01

    The above article, published online on April 18, 2011 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the journal Editor in Chief, Gary S. Stein, and Wiley Periodicals, Inc. The retraction has been agreed following an investigation from Wayne State University involving the second author that found Figures 1C and 4C to be inappropriately re-used and re-labeled. REFERENCE Bao B, Wang Z, Ali S, Kong D, Banerjee S, Ahmad A, Li Y, Azmi AS, Miele L, Sarkar FH. 2011. Over-expression of FoxM1 leads to epithelial-mesenchymal transition and cancer stem cell phenotype in pancreatic cancer cells. J Cell Biochem 112:2296-2306; doi: 10.1002/jcb.23150.

  9. Molecules involved in epithelial-mesenchymal transition and epithelial-stromal interaction in phyllodes tumors: implications for histologic grade and prognosis.

    Science.gov (United States)

    Kwon, Ji Eun; Jung, Woo-Hee; Koo, Ja Seung

    2012-06-01

    The aim of this study was to investigate the expression of molecules associated with epithelial-mesenchymal transition (EMT) and epithelial-stromal interactions (ESI) and to evaluate their roles in phyllodes tumors (PTs). Tissue microarrays (TMAs) were constructed from 207 PT specimens (157 benign, 34 borderline and 16 malignant). The presence of EMT-related markers including N-cadherin, Twist, TGF-beta, HMGA2, S100A4 and Ezrin as well as ESI-related molecules such as SDF1 and CXCR4 among the TMAs was assessed immunohistochemically. Immunohistochemical results were analyzed in terms of clinicopathologic parameters. For higher grade PTs, expressions of Twist (p EMT-associated molecules, and CXCR4, an ESI-associated molecule, were increased in the stromal component of advanced grade PTs. Further, high expression of Twist in the stromal component was correlated with poorer prognoses.

  10. Elf5 inhibits the epithelial-mesenchymal transition in mammary gland development and breast cancer metastasis by transcriptionally repressing Snail2.

    Science.gov (United States)

    Chakrabarti, Rumela; Hwang, Julie; Andres Blanco, Mario; Wei, Yong; Lukačišin, Martin; Romano, Rose-Anne; Smalley, Kirsten; Liu, Song; Yang, Qifeng; Ibrahim, Toni; Mercatali, Laura; Amadori, Dino; Haffty, Bruce G; Sinha, Satrajit; Kang, Yibin

    2012-11-01

    The epithelial-mesenchymal transition (EMT) is a complex process that occurs during organogenesis and in cancer metastasis. Despite recent progress, the molecular pathways connecting the physiological and pathological functions of EMT need to be better defined. Here we show that the transcription factor Elf5, a key regulator of mammary gland alveologenesis, controls EMT in both mammary gland development and metastasis. We uncovered this role for Elf5 through analyses of Elf5 conditional knockout animals, various in vitro and in vivo models of EMT and metastasis, an MMTV-neu transgenic model of mammary tumour progression and clinical breast cancer samples. Furthermore, we demonstrate that Elf5 suppresses EMT by directly repressing the transcription of Snail2, a master regulator of mammary stem cells and a known inducer of EMT. These findings establish Elf5 not only as a key cell lineage regulator during normal mammary gland development, but also as a suppressor of EMT and metastasis in breast cancer.

  11. Extracellular vesicles from women with breast cancer promote an epithelial-mesenchymal transition-like process in mammary epithelial cells MCF10A.

    Science.gov (United States)

    Galindo-Hernandez, Octavio; Gonzales-Vazquez, Cristina; Cortes-Reynosa, Pedro; Reyes-Uribe, Emmanuel; Chavez-Ocaña, Sonia; Reyes-Hernandez, Octavio; Sierra-Martinez, Mónica; Salazar, Eduardo Perez

    2015-12-01

    Extracellular vesicles (EVs) mediate many stages of tumor progression including angiogenesis, escape from immune surveillance, and extracellular matrix degradation. We studied whether EVs from plasma of women with breast cancer are able to induce an epithelial-mesenchymal transition (EMT) process in mammary epithelial cells MCF10A. Our findings demonstrate that EVs from plasma of breast cancer patients induce a downregulation of E-cadherin expression and an increase of vimentin and N-cadherin expression. Moreover, EVs induce migration and invasion, as well as an increase of NFκB-DNA binding activity and MMP-2 and MMP-9 secretions. In summary, our findings demonstrate, for the first time, that EVs from breast cancer patients induce an EMT-like process in human mammary non-tumorigenic epithelial cells MCF10A.

  12. Overexpression of EphA2 correlates with epithelial-mesenchymal transition-related proteins in gastric cancer and their prognostic importance for postoperative patients.

    Science.gov (United States)

    Hou, Futao; Yuan, Weijie; Huang, Jin; Qian, Liyuan; Chen, Zhikang; Ge, Jie; Wu, Shaobin; Chen, Jinxiang; Wang, Jixu; Chen, Zihua

    2012-12-01

    The expression of EphA2 and three epithelial-mesenchymal transition-related proteins (E-cadherin, β-catenin and vimentin) was detected by immunohistochemistry in human gastric cancer and normal gastric mucosa. The expression of EphA2 and vimentin was significantly higher in gastric cancer tissues than in normal gastric mucosa tissues, and similar results were found for negative E-cadherin expression and ectopic β-catenin expression. Further analysis showed that the expression of EphA2 was closely correlated with the depth of tumor invasion, tumor-node-metastasis (TNM) stages and lymph node metastasis. Down-regulated expression of the epithelial protein E-cadherin, overexpression of the mesenchymal protein vimentin and ectopic expression of β-catenin were associated with the depth of tumor invasion, tumor differentiation, TNM stages and lymph node metastasis. The Spearman rank test indicated that the positive expression of EphA2 was negatively associated with E-cadherin expression and was positively correlated with β-catenin ectopic expression and vimentin expression. In addition, the Kaplan-Meier survival analysis showed that the overexpression of EphA2 and vimentin, ectopic expression of β-catenin and down-regulation of E-cadherin indicate a poor outcome. Moreover, multivariate Cox analysis showed that TNM stages, lymph node metastasis, EphA2 expression, E-cadherin expression and β-catenin ectopic expression were independent prognostic factors for postoperative gastric cancer. These findings indicate that the overexpression of EphA2 correlates with the loss of epithelial proteins and the appearance of mesenchymal proteins. Therefore, EphA2 may play a role in epithelial-mesenchymal transition in gastric cancer.

  13. Loss of prostasin (PRSS8 in human bladder transitional cell carcinoma cell lines is associated with epithelial-mesenchymal transition (EMT

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    Chai Karl X

    2009-10-01

    Full Text Available Abstract Background The glycosylphosphatidylinositol (GPI-anchored epithelial extracellular membrane serine protease prostasin (PRSS8 is expressed abundantly in normal epithelia and essential for terminal epithelial differentiation, but down-regulated in human prostate, breast, and gastric cancers and invasive cancer cell lines. Prostasin is involved in the extracellular proteolytic modulation of the epidermal growth factor receptor (EGFR and is an invasion suppressor. The aim of this study was to evaluate prostasin expression states in the transitional cell carcinomas (TCC of the human bladder and in human TCC cell lines. Methods Normal human bladder tissues and TCC on a bladder cancer tissue microarray (TMA were evaluated for prostasin expression by means of immunohistochemistry. A panel of 16 urothelial and TCC cell lines were evaluated for prostasin and E-cadherin expression by western blot and quantitative PCR, and for prostasin gene promoter region CpG methylation by methylation-specific PCR (MSP. Results Prostasin is expressed in the normal human urothelium and in a normal human urothelial cell line, but is significantly down-regulated in high-grade TCC and lost in 9 (of 15 TCC cell lines. Loss of prostasin expression in the TCC cell lines correlated with loss of or reduced E-cadherin expression, loss of epithelial morphology, and promoter DNA hypermethylation. Prostasin expression could be reactivated by demethylation or inhibition of histone deacetylase. Re-expression of prostasin or a serine protease-inactive variant resulted in transcriptional up-regulation of E-cadherin. Conclusion Loss of prostasin expression in bladder transitional cell carcinomas is associated with epithelial-mesenchymal transition (EMT, and may have functional implications in tumor invasion and resistance to chemotherapy.

  14. MicroRNA-616 promotes the migration, invasion and epithelial-mesenchymal transition of HCC by targeting PTEN.

    Science.gov (United States)

    Zhang, Di; Zhou, Peihua; Wang, Wei; Wang, Xiaolong; Li, Junhui; Sun, Xuejun; Zhang, Li

    2016-01-01

    MicroRNAs, which can post-transcriptionally regulate gene expression by binding to the 3'-untranslated regions of the mRNAs, have been found to be the critical regulators of the development and progression of hepatocellular carcinoma (HCC). The present study demonstrated for the first time that microRNA-616 (miR-616) was markedly upregulated in HCC tissues, and was associated with the recurrence and metastasis of HCC. Elevated level of miR-616 was correlated with adverse clinicopathological features and poor prognosis of HCC patients. Gain- and loss-of-function studies revealed that miR-616 could potentiate the migration, invasion and the epithelial-mesenchymal transtion (EMT) phenotype of HCC cells. Phosphatase and tensin homolog (PTEN), the predicted target of miR-616 by bioinformatics analysis, was confirmed as a direct downstream target of miR-616 through western blotting, luciferase reporter and immunohistochemical assays. Furthermore, we demonstrated that miR-616 exerted the promoting effects on EMT and metastatic ability of HCC cells through suppressing PTEN expression. Based on these results, we conclude that miR-616 is a promising prognostic biomarker of HCC and targeting miR-616 may be a potential option to prevent the progression of HCC.

  15. Multiwall carbon nanotubes directly promote fibroblast-myofibroblast and epithelial-mesenchymal transitions through the activation of the TGF-β/Smad signaling pathway.

    Science.gov (United States)

    Wang, Peng; Wang, Yue; Nie, Xin; Braïni, Céline; Bai, Ru; Chen, Chunying

    2015-01-27

    A number of studies have demonstrated that MWCNTs induce granuloma formation and fibrotic responses in vivo, and it has been recently reported that MWCNT-induced macrophage activation and subsequent TGF-β secretion contribute to pulmonary fibrotic responses. However, their direct effects against alveolar type-II epithelial cells and fibroblasts and the corresponding underlying mechanisms remain largely unaddressed. Here, MWCNTs are reported to be able to directly promote fibroblast-to-myofibroblast conversion and the epithelial-mesenchymal transition (EMT) through the activation of the TGF-β/Smad signaling pathway. Both of the cell transitions may play important roles in MWCNT-induced pulmonary fibrosis. Firstly, in-vivo and in-vitro data show that long MWCNTs can directly interact with fibroblasts and epithelial cells, and some of them may be uptaken into fibroblasts and epithelial cells by endocytosis. Secondly, long MWCNTs can directly activate fibroblasts and increase both the basal and TGF-β1-induced expression of the fibroblast-specific protein-1, α-smooth muscle actin, and collagen III. Finally, MWCNTs can induce the EMT through the activation of TGF-β/Smad2 signaling in alveolar type-II epithelial cells, from which some fibroblasts involved in pulmonary fibrosis are thought to originate. These observations suggest that the activation of the TGF-β/Smad2 signaling plays a critical role in the process of the fibroblast-to-myofibroblast transition and the EMT induced by MWCNTs.

  16. The Mu opioid receptor promotes opioid and growth factor-induced proliferation, migration and Epithelial Mesenchymal Transition (EMT in human lung cancer.

    Directory of Open Access Journals (Sweden)

    Frances E Lennon

    Full Text Available Recent epidemiologic studies implying differences in cancer recurrence based on anesthetic regimens raise the possibility that the mu opioid receptor (MOR can influence cancer progression. Based on our previous observations that overexpression of MOR in human non-small cell lung cancer (NSCLC cells increased tumor growth and metastasis, this study examined whether MOR regulates growth factor receptor signaling and epithelial mesenchymal transition (EMT in human NSCLC cells. We utilized specific siRNA, shRNA, chemical inhibitors and overexpression vectors in human H358 NSCLC cells that were either untreated or treated with various concentrations of DAMGO, morphine, fentanyl, EGF or IGF. Cell function assays, immunoblot and immunoprecipitation assays were then performed. Our results indicate MOR regulates opioid and growth factor-induced EGF receptor signaling (Src, Gab-1, PI3K, Akt and STAT3 activation which is crucial for consequent human NSCLC cell proliferation and migration. In addition, human NSCLC cells treated with opioids, growth factors or MOR overexpression exhibited an increase in snail, slug and vimentin and decrease ZO-1 and claudin-1 protein levels, results consistent with an EMT phenotype. Further, these effects were reversed with silencing (shRNA or chemical inhibition of MOR, Src, Gab-1, PI3K, Akt and STAT3 (p<0.05. Our data suggest a possible direct effect of MOR on opioid and growth factor-signaling and consequent proliferation, migration and EMT transition during lung cancer progression. Such an effect provides a plausible explanation for the epidemiologic findings.

  17. Real-time imaging of the epithelial-mesenchymal transition using microRNA-200a sequence-based molecular beacon-conjugated magnetic nanoparticles.

    Directory of Open Access Journals (Sweden)

    YoonSeok Choi

    Full Text Available The epithelial-mesenchymal transition (EMT plays important roles in tumor progression to metastasis. Thus, the development of an imaging probe that can monitor transient periods of the EMT process in live cells is required for a better understanding of metastatic process. Inspired by the fact that the mRNA expression levels of zinc finger E-box-binding homeobox 1 (ZEB1 increase when cells adopt mesenchyme characteristics and that microRNA-200a (miR-200a can bind to ZEB1 mRNA, we conjugated molecular beacon (MB mimicking mature miR-200a to magnetic nanoparticles (miR-200a-MB-MNPs and devised an imaging method to observe transitional changes in the cells during EMT. Transforming growth factor-β1 treated epithelial cells and breast cancer cell lines representing both epithelial and mesenchymal phenotypes were used for the validation of miR-200a-MB-MNPs as an EMT imaging probe. The real-time imaging of live cells acquired with the induction of EMT revealed an increase in fluorescence signals by miR-200a-MB-MNPs, cell morphology alterations, and the loss of cell-cell adhesion. Our results suggest that miR-200a-MB-MNPs can be used as an imaging probe for the real-time monitoring of the EMT process in live cells.

  18. Epithelial-Mesenchymal Transition (EMT and Regulation of EMT Factors by Steroid Nuclear Receptors in Breast Cancer: A Review and in Silico Investigation

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    Ioannis A. Voutsadakis

    2016-01-01

    Full Text Available Steroid Nuclear Receptors (SNRs are transcription factors of the nuclear receptor super-family. Estrogen Receptor (ERα is the best-studied and has a seminal role in the clinic both as a prognostic marker but also as a predictor of response to anti-estrogenic therapies. Progesterone Receptor (PR is also used in the clinic but with a more debatable prognostic role and the role of the four other SNRs, ERβ, Androgen Receptor (AR, Glucocorticoid Receptor (GR and Mineralocorticoid Receptor (MR, is starting only to be appreciated. ERα, but also to a certain degree the other SNRs, have been reported to be involved in virtually every cancer-enabling process, both promoting and impeding carcinogenesis. Epithelial-Mesenchymal Transition (EMT and the reverse Mesenchymal Epithelial Transition (MET are such carcinogenesis-enabling processes with important roles in invasion and metastasis initiation but also establishment of tumor in the metastatic site. EMT is governed by several signal transduction pathways culminating in core transcription factors of the process, such as Snail, Slug, ZEB1 and ZEB2, and Twist, among others. This paper will discuss direct regulation of these core transcription factors by SNRs in breast cancer. Interrogation of publicly available databases for binding sites of SNRs on promoters of core EMT factors will also be included in an attempt to fill gaps where other experimental data are not available.

  19. Epithelial-Mesenchymal Transitions and the Expression of Twist in MCF-7/ADR,Human Multidrug-Resistant Breast Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    Fei Zhang; Yurong Shi; Lin Zhang; Bin Zhang; Xiyin Wei; Yi Yang; RUi Wang; Ruifang Niu

    2007-01-01

    OBJECTIVE To study the expression levels of Twist and epithelialmesenchymal transitions in multidrug-resistant MCF-7/ADR breast cancer cells,and to study the relationship between multidrug resistance (MDR) and metastatic potential of the cells.METHODS RT-PCR,immunohislochemical and Western blotting methods were used to examine the changes of expression levels of the transcription factor Twist.E-cadherin and N-cadherin in the MCF-7 breast cancer cell line and its multidrug-resistant variant.MCF-7/ADR.RESULTS In MCF-7 cells,the expression of E-cadherin can be detected,but there is no expression of Twisl or N-cadherin.In MCF-7/ADR cells,E-cadherin expression is lost.bul the expression of two other genes was significantly positive.CONCLUSION Epithelial-mesenchymal transitions induced by Twist,may have a relationship with enhanced invasion and metastatic potential during the development of multidrug-resistant MCF-7/ADR breast cancer cells.

  20. Epithelial-Mesenchymal Transition (EMT) and Regulation of EMT Factors by Steroid Nuclear Receptors in Breast Cancer: A Review and in Silico Investigation

    Science.gov (United States)

    Voutsadakis, Ioannis A.

    2016-01-01

    Steroid Nuclear Receptors (SNRs) are transcription factors of the nuclear receptor super-family. Estrogen Receptor (ERα) is the best-studied and has a seminal role in the clinic both as a prognostic marker but also as a predictor of response to anti-estrogenic therapies. Progesterone Receptor (PR) is also used in the clinic but with a more debatable prognostic role and the role of the four other SNRs, ERβ, Androgen Receptor (AR), Glucocorticoid Receptor (GR) and Mineralocorticoid Receptor (MR), is starting only to be appreciated. ERα, but also to a certain degree the other SNRs, have been reported to be involved in virtually every cancer-enabling process, both promoting and impeding carcinogenesis. Epithelial-Mesenchymal Transition (EMT) and the reverse Mesenchymal Epithelial Transition (MET) are such carcinogenesis-enabling processes with important roles in invasion and metastasis initiation but also establishment of tumor in the metastatic site. EMT is governed by several signal transduction pathways culminating in core transcription factors of the process, such as Snail, Slug, ZEB1 and ZEB2, and Twist, among others. This paper will discuss direct regulation of these core transcription factors by SNRs in breast cancer. Interrogation of publicly available databases for binding sites of SNRs on promoters of core EMT factors will also be included in an attempt to fill gaps where other experimental data are not available. PMID:26797644

  1. The involvement of hematopoietic pre-B cell leukemia transcription factor-interacting protein in regulating epithelial-mesenchymal transition of human spinal glioblastoma.

    Science.gov (United States)

    Wang, Deliang; Wang, Li; Zhou, Yi; Zhao, Xinjun; Xiong, Hui

    2016-05-01

    To date, hematopoietic pre-B cell leukemia transcription factor-interacting protein (HPIP), a co-repressor for the transcription factor PBX, has been involved into the initiation and onset in a wide variety of cancers. However, the molecular mechanisms underlying HPIP-induced epithelial-mesenchymal transition (EMT) in the spinal glioblastoma have been under investigation. In the present study, spinal glioblastoma tissues, U87, and U251 cell lines were used and subjected to in vitro assays, such as RT-PCR, and Western blot. Here, in vitro assays revealed that HPIP mRNA and protein were highly expressed in five cases of spinal glioblastoma tissues, compared with non-tumor tissues. Subsequently, in vitro experiments demonstrated HPIP promoted the U87 and U251 cell growth and regulated the G1/S phase transitions in U87 and U251 cell cycle, respectively, accompanied by the increased expression of cyclin A2, cyclin B1, and cyclin D1. Furthermore, HPIP increased the expression of N-cadherin, Slug, and MMP2, and decreased the expression of E-cadherin. By contrast, knockdown of HPIP reversed HPIP-induced EMT biomarkers, migration, and invasion in U87 and U251 cells. In conclusion, our findings identified HPIP plays an important role in the progression and EMT of spinal glioblastoma, by which cell growth is improved. Thus, HPIP gene or protein could act as a useful target in the clinical practice.

  2. Beta-elemene blocks epithelial-mesenchymal transition in human breast cancer cell line MCF-7 through Smad3-mediated down-regulation of nuclear transcription factors.

    Directory of Open Access Journals (Sweden)

    Xian Zhang

    Full Text Available Epithelial-mesenchymal transition (EMT is the first step required for breast cancer to initiate metastasis. However, the potential of drugs to block and reverse the EMT process are not well explored. In the present study, we investigated the inhibitory effect of beta-elemene (ELE, an active component of a natural plant-derived anti-neoplastic agent in an established EMT model mediated by transforming growth factor-beta1 (TGF-β1. We found that ELE (40 µg/ml blocked the TGF-β1-induced phenotypic transition in the human breast cancer cell line MCF-7. ELE was able to inhibit TGF-β1-mediated upregulation of mRNA and protein expression of nuclear transcription factors (SNAI1, SNAI2, TWIST and SIP1, potentially through decreasing the expression and phosphorylation of Smad3, a central protein mediating the TGF-β1 signalling pathway. These findings suggest a potential therapeutic benefit of ELE in treating basal-like breast cancer.

  3. α-Mangostin Suppresses the Viability and Epithelial-Mesenchymal Transition of Pancreatic Cancer Cells by Downregulating the PI3K/Akt Pathway

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    Qinhong Xu

    2014-01-01

    Full Text Available α-Mangostin, a natural product isolated from the pericarp of the mangosteen fruit, has been shown to inhibit the growth of tumor cells in various types of cancers. However, the underlying molecular mechanisms are largely unclear. Here, we report that α-mangostin suppressed the viability and epithelial-mesenchymal transition (EMT of pancreatic cancer cells through inhibition of the PI3K/Akt pathway. Treatment of pancreatic cancer BxPc-3 and Panc-1 cells with α-mangostin resulted in loss of cell viability, accompanied by enhanced cell apoptosis, cell cycle arrest at G1 phase, and decrease of cyclin-D1. Moreover, Transwell and Matrigel invasion assays showed that α-mangostin significantly reduced the migration and invasion of pancreatic cancer cells. Consistent with these results, α-mangostin decreased the expression of MMP-2, MMP-9, N-cadherin, and vimentin and increased the expression of E-cadherin. Furthermore, we found that α-mangostin suppressed the activity of the PI3K/Akt pathway in pancreatic cancer cells as demonstrated by the reduction of the Akt phosphorylation by α-mangostin. Finally, α-mangostin significantly inhibited the growth of BxPc-3 tumor mouse xenografts. Our results suggest that α-mangostin may be potentially used as a novel adjuvant therapy or complementary alternative medicine for the management of pancreatic cancers.

  4. Long Non Coding RNA MALAT1 Promotes Tumor Growth and Metastasis by inducing Epithelial-Mesenchymal Transition in Oral Squamous Cell Carcinoma.

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    Zhou, Xuan; Liu, Su; Cai, Guoshuai; Kong, Lingping; Zhang, Tingting; Ren, Yu; Wu, Yansheng; Mei, Mei; Zhang, Lun; Wang, Xudong

    2015-11-02

    The prognosis of advanced oral squamous cell carcinoma (OSCC) patients remains dismal, and a better understanding of the underlying mechanisms is critical for identifying effective targets with therapeutic potential to improve the survival of patients with OSCC. This study aims to clarify the clinical and biological significance of metastasis-associated long non-coding RNA, metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in OSCC. We found that MALAT1 is overexpressed in OSCC tissues compared to normal oral mucosa by real-time PCR. MALAT1 served as a new prognostic factor in OSCC patients. When knockdown by small interfering RNA (siRNA) in OSCC cell lines TSCCA and Tca8113, MALAT1 was shown to be required for maintaining epithelial-mesenchymal transition (EMT) mediated cell migration and invasion. Western blot and immunofluorescence staining showed that MALAT1 knockdown significantly suppressed N-cadherin and Vimentin expression but induced E-cadherin expression in vitro. Meanwhile, both nucleus and cytoplasm levels of β-catenin and NF-κB were attenuated, while elevated MALAT1 level triggered the expression of β-catenin and NF-κB. More importantly, targeting MALAT1 inhibited TSCCA cell-induced xenograft tumor growth in vivo. Therefore, these findings provide mechanistic insight into the role of MALAT1 in regulating OSCC metastasis, suggesting that MALAT1 is an important prognostic factor and therapeutic target for OSCC.

  5. TLR4-mediated galectin-1 production triggers epithelial-mesenchymal transition in colon cancer cells through ADAM10- and ADAM17-associated lactate production.

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    Park, Ga Bin; Kim, Daejin

    2017-01-01

    Toll-like receptor 4 (TLR4) activation is a key contributor to the carcinogenesis of colon cancer. Overexpression of galectin-1 (Gal-1) also correlates with increased invasive activity of colorectal cancer. Lactate production is a critical predictive factor of risk of metastasis, but the functional relationship between intracellular lactate and Gal-1 expression in TLR4-activated colon cancer remains unknown. In this study, we investigated the underlying mechanism and role of Gal-1 in metastasis and invasion of colorectal cancer (CRC) cells after TLR4 stimulation. Exposure to the TLR4 ligand lipopolysaccharide (LPS) increased expression of Gal-1, induced EMT-related cytokines, triggered the activation of glycolysis-related enzymes, and promoted lactate production. Gene silencing of TLR4 and Gal-1 in CRC cells inhibited lactate-mediated epithelial-mesenchymal transition (EMT) after TLR4 stimulation. Gal-1-mediated activation of a disintegrin and metalloproteinase 10 (ADAM10) and ADAM 17 increased the invasion activity and expression of mesenchymal characteristics in LPS-activated CRC cells. Conversely, inhibition of ADAM10 or ADAM17 effectively blocked the generation of lactate and the migration capacity of LPS-treated CRC cells. Thus, the TLR4/Gal-1 signaling pathway regulates lactate-mediated EMT processes through the activation of ADAM10 and ADAM17 in CRC cells.

  6. Disulfiram inhibits TGF-β-induced epithelial-mesenchymal transition and stem-like features in breast cancer via ERK/NF-κB/Snail pathway.

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    Han, Dan; Wu, Gang; Chang, Chan; Zhu, Fang; Xiao, Yin; Li, Qiuhui; Zhang, Tao; Zhang, Liling

    2015-12-01

    Disulfiram (DSF), an anti-alcoholism drug, has been reported as an inhibitor of NF-κB. NF-κB is involved in epithelial-mesenchymal transition (EMT) and self-renewal of breast cancer stem cells (CSCs). In this study, we treated MCF-7 and MDA-MB-231 breast cancer cells with TGF-β to induce EMT and cancer stem-like features and studied whether DSF can reverse this process. We found that DSF inhibited TGF-β induced EMT in breast cancer cells in a dose-dependent manner. Also, DSF inhibited EMT-associated stem-like features, migration and invasion of tumor cells as well as tumor growth in xenograft model. The activation of NF-κB was linked with EMT and stem-like cells. We conclude that DSF can suppress NF-κB activity and downregulate ERK/NF-κB/Snail pathway, leading to reverse EMT and stem-like features. Our data suggest that DSF inhibits EMT and stem-like properties in breast cancer cells associated with inhibition of the ERK/NF-κB/Snail pathway.

  7. CCR7 enhances TGF-β1-induced epithelial-mesenchymal transition and is associated with lymph node metastasis and poor overall survival in gastric cancer.

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    Ma, Huiying; Gao, Lingling; Li, Shichao; Qin, Jie; Chen, Long; Liu, Xinzhou; Xu, Pingping; Wang, Fei; Xiao, Honglei; Zhou, Shuang; Gao, Qiang; Liu, Binbin; Sun, Yihong; Liang, Chunmin

    2015-09-15

    CCR7 is a G protein-coupled chemokine receptor. In this study, we used immunohistochemistry with tissue microarrays to measure CCR7 expression in tumor specimens from 122 patients with gastric cancer. We show that CCR7 expression is associated with lymph node metastasis (P = 0.022) and overall survival (OS; P = 0.025), and is an independent factor associated with poorer overall survival (P = 0.032). The CCR7 mechanism was predicted based on bioinformatic analysis and verified in gastric cancer cell lines and primary tumor samples. The data show that CCR7 contributes to TGF-β1-induced epithelial-mesenchymal transition (EMT) and that the effects of TGF-β1 are inhibited by a CCR7 neutralizing antibody or a NF-κB inhibitor. Increased TGF-β1 expression was accompanied by nuclear localization of NF-κB-p65 and higher levels of the mesenchymal marker vimentin in human gastric cancer samples. We conclude that the CCR7 axis mediates TGF-β1-induced EMT via crosstalk with NF-κB signaling, facilitating lymph node metastasis and poorer overall survival in patients with gastric cancer. These findings suggest CCR7 is a novel prognostic indicator and a potential target for gastric cancer therapy.

  8. Carnosic Acid Inhibits the Epithelial-Mesenchymal Transition in B16F10 Melanoma Cells: A Possible Mechanism for the Inhibition of Cell Migration

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    So Young Park

    2014-07-01

    Full Text Available Carnosic acid is a natural benzenediol abietane diterpene found in rosemary and exhibits anti-inflammatory, antioxidant, and anti-carcinogenic activities. In this study, we evaluated the effects of carnosic acid on the metastatic characteristics of B16F10 melanoma cells. When B16F10 cells were cultured in an in vitro Transwell system, carnosic acid inhibited cell migration in a dose-dependent manner. Carnosic acid suppressed the adhesion of B16F10 cells, as well as the secretion of matrix metalloproteinase (MMP-9, tissue inhibitor of metalloproteinase (TIMP-1, urokinase plasminogen activator (uPA, and vascular cell adhesion molecule (VCAM-1. Interestingly, secretion of TIMP-2 increased significantly in B16F10 cells treated with 10 μmol/L carnosic acid. Additionally, carnosic acid suppressed the mesenchymal markers snail, slug, vimentin, and N-cadherin and induced epithelial marker E-cadherin. Furthermore, carnosic acid suppressed phosphorylation of Src, FAK, and AKT. These results indicate that inhibition of the epithelial-mesenchymal transition may be important for the carnosic acid-induced inhibition of B16F10 cell migration.

  9. SDF-1/CXCR4 Axis Regulates Cell Cycle Progression and Epithelial-Mesenchymal Transition via Up-regulation of Survivin in Glioblastoma.

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    Liao, Anyan; Shi, Ranran; Jiang, Yuliang; Tian, Suqing; Li, Panpan; Song, Fuxi; Qu, Yalan; Li, Jinna; Yun, Haiqin; Yang, Xiangshan

    2016-01-01

    Stromal cell-derived factor 1 (SDF-1)/CXCR4 ligand-receptor axis is widely recommended as an attractive target for cancer therapy. Meanwhile, epithelial-mesenchymal transition (EMT) process is linked to disease pathophysiology. As one of inhibitors of apoptosis proteins, survivin is implicated in the onset and development of cancer. In the present study, we tried to determine the cause-effect associations between SDF-1/CXCR4 axis and survivin expression in glioblastoma U-251 cell line. Survivin activation and inhibition were induced with exogenous SDF-1 and survivin small interfering RNA (survivin siRNA), respectively. Western blot was used to detect relevant proteins in SDF-1/CXCR4 axis. Western blot analysis revealed that survivin expression in U-251 increased in a dose- and time-dependent manner in response to SDF-1 treatment. However, the interference with MEK/ERK and PI3K/AKT pathway prohibited SDF-1-induced survivin up-regulation. Importantly, survivin knockdown abrogated cell cycle progression and the expression of snail and N-cadherin, compared with non-transfectants. In conclusion, the present study shows that SDF-1 up-regulates survivin via MEK/ERK and PI3K/AKT pathway, leading to cell cycle progression and EMT occurrence dependent on survivin. The blockade of survivin will allow for the treatment of glioblastoma.

  10. Osteopontin Promotes Invasion, Migration and Epithelial-Mesenchymal Transition of Human Endometrial Carcinoma Cell HEC-1A Through AKT and ERK1/2 Signaling

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    Yinghua Li

    2015-10-01

    Full Text Available Background/Aims: Osteopontin (OPN is an Extracellular Matrix (ECM molecule and is involved in many physiologic and pathologic processes, including cell adhesion, angiogenesis and tumor metastasis. OPN is a well-known multifunctional factor involved in various aspects of cancer progression, including endometrial cancer. In this study, we examined the significance of OPN in endometrial cancer. Methods: The proliferation, migration and invasion ability of HEC-1A cells were detected by Cell Counting Kit-8 (CCK-8, Wound scratch assay and transwell. Western blots were employed to detect the expression of Matrix metalloproteinase-2 (MMP-2 and epithelial-mesenchymal transition (EMT-related factors in HEC-1A cells treated with rhOPN. Results: rhOPN promotes cell proliferation, migration and invasion in HEC-1A cells. rhOPN influenced EMT-related factors and MMP-2 expression in HEC-1A cells. rhOPN promoted HEC-1A cells migration, invasion and EMT through protein kinase B (PKB/AKT and Extracellular regulated protein kinases (ERK1/2 signaling pathway. Conclusions: These results may open up a novel therapeutic strategy for endometrial cancer: namely, rhOPN have important roles in controlling growth of endometrial of cancer cells and suggest a novel target pathway for treatment of this cancer.

  11. Expression of transcription factors Slug in the lens epithelial cells undergoing epithelial-mesenchymal transition induced by connective tissue growth factor

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    Ying-Na Wang

    2015-10-01

    Full Text Available AIM:To investigate the expression of transcription factors Slug in human lens epithelial cells (HLECs undergoing epithelial-mesenchymal transition (EMT induced by connective tissue growth factor (CTGF.METHODS: HLECs were treated with CTGF of different concentrations (20, 50 and 100 ng/mL or without CTGF (control for 24h. The morphological changes of HLECs were analysed by microscopy. The expression and cellular localization of Slug was evaluated by immumo-fluorescence. Expressions of Slug, E-cadherin and alpha smooth muscle actin (α-SMA were further determined by Western blot analysis. RESULTS: HLECs showed spidle fibrolasts-like characteristics and loosely connected each other after CTGF treatment. The immuno-fluorescence staining indicated that Slug was localized in the nuclei and its expression was induced by CTGF. The relative expressions of Slug protein were 1.64±0.11, 1.96 ±0.03, 3.12 ±0.10, and 4.08±0.14, respectively, in response to control group and treatment with CTGF of 20, 50 and 100 ng/mL (F=443.86, PCONCLUSION: Transcription factor Slug may be involved in EMT of HLECs induced by CTGF in vitro.

  12. Elucidation of epithelial-mesenchymal transition-related pathways in a triple-negative breast cancer cell line model by multi-omics interactome analysis.

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    Pauling, Josch K; Christensen, Anne G; Batra, Richa; Alcaraz, Nicolas; Barbosa, Eudes; Larsen, Martin R; Beck, Hans C; Leth-Larsen, Rikke; Azevedo, Vasco; Ditzel, Henrik J; Baumbach, Jan

    2014-11-01

    In life sciences, and particularly biomedical research, linking aberrant pathways exhibiting phenotype-specific alterations to the underlying physical condition or disease is an ongoing challenge. Computationally, a key approach for pathway identification is data enrichment, combined with generation of biological networks. This allows identification of intrinsic patterns in the data and their linkage to a specific context such as cellular compartments, diseases or functions. Identification of aberrant pathways by traditional approaches is often limited to biological networks based on either gene expression, protein expression or post-translational modifications. To overcome single omics analysis, we developed a set of computational methods that allow a combined analysis of data collections from multiple omics fields utilizing hybrid interactome networks. We apply these methods to data obtained from a triple-negative breast cancer cell line model, combining data sets of gene and protein expression as well as protein phosphorylation. We focus on alterations associated with the phenotypical differences arising from epithelial-mesenchymal transition in two breast cancer cell lines exhibiting epithelial-like and mesenchymal-like morphology, respectively. Here we identified altered protein signaling activity in a complex biologically relevant network, related to focal adhesion and migration of breast cancer cells. We found dysregulated functional network modules revealing altered phosphorylation-dependent activity in concordance with the phenotypic traits and migrating potential of the tested model. In addition, we identified Ser267 on zyxin, a protein coupled to actin filament polymerization, as a potential in vivo phosphorylation target of cyclin-dependent kinase 1.

  13. N-myc downstream regulated gene 1 (NDRG1 promotes metastasis of human scirrhous gastric cancer cells through epithelial mesenchymal transition.

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    Hiroki Ureshino

    Full Text Available Our recent study demonstrated that higher expression of N-myc downregulated gene 1 (NDRG1 is closely correlated with poor prognosis in gastric cancer patients. In this study, we asked whether NDRG1 has pivotal roles in malignant progression including metastasis of gastric cancer cells. By gene expression microarray analysis expression of NDRG1 showed the higher increase among a total of 3691 up-regulated genes in a highly metastatic gastric cancer cell line (58As1 than their parental low metastatic counterpart (HSC-58. The highly metastatic cell lines showed decreased expression of E-cadherin, together with enhanced expression of vimentin and Snail. This decreased expression of E-cadherin was restored by Snail knockdown in highly metastatic cell lines. We next established stable NDRG1 knockdown cell lines (As1/Sic50 and As1/Sic54 from the highly metastatic cell line, and both of these cell lines showed enhanced expression of E-cadherin and decreased expression of vimentin and Snail. And also, E-cadherin promoter-driven luciferase activity was found to be increased by NDRG1 knockdown in the highly metastatic cell line. NDRG1 knockdown in gastric cancer cell showed suppressed invasion of cancer cells into surround tissues, suppressed metastasis to the peritoneum and decreased ascites accumulation in mice with significantly improved survival rates. This is the first study to demonstrate that NDRG1 plays its pivotal role in the malignant progression of gastric cancer through epithelial mesenchymal transition.

  14. Microcystin-LR promotes epithelial-mesenchymal transition in colorectal cancer cells through PI3-K/AKT and SMAD2.

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    Ren, Yan; Yang, Mengli; Chen, Meng; Zhu, Qiangqiang; Zhou, Lihua; Qin, Wei; Wang, Ting

    2017-01-04

    Increasing evidences suggest that microcystins, a kind of toxic metabolites, produced by cyanobacteria in contaminated water may contribute to the aggravation of the human colorectal carcinoma. Our previous study showed that microcystin-LR (MC-LR) exposure caused significant invasion and migration of colorectal cancer cells. However, the roles of MC-LR in regulating epithelial-mesenchymal transition (EMT) in colorectal cancer cells remain unknown. In our study, we observed that MC-LR treatment decreased epithelial marker E-cadherin expression and up-regulated the levels of mesenchymal markers Vimentin and Snail in colorectal cancer cells. Moreover, MC-LR stimulated protein expression of SMAD2 and phospho-SMAD2 by PI3-K/AKT activation. The activated PI3-K/AKT and SMAD2 signaling largely accounted for MC-LR-induced EMT, which could be reversed by SMAD2 RNA interference or PI3-K/AKT chemical inhibitor in colorectal cancer cells. Our results show that MC-LR could induce SMAD2 expression to promote colorectal cancer cells EMT, which not only provides a mechanistic insight on MC-LR promotes EMT in colorectal cancer cells, but also support to the development of therapies aimed at SMAD2 in colorectal cancer induced by MC-LR. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  15. Loss of cellular polarity/cohesiveness in the invasive front of papillary thyroid carcinoma, a novel predictor for lymph node metastasis; possible morphological indicator of epithelial mesenchymal transition.

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    Liu, Zhiyan; Kakudo, Kennichi; Bai, Yanhua; Li, Yaqiong; Ozaki, Takashi; Miyauchi, Akira; Taniguchi, Emiko; Mori, Ichiro

    2011-04-01

    Loss of cellular polarity/cohesiveness (LOP/C) in the invasive front of papillary thyroid carcinoma (PTC) results in a high recurrence risk of PTC. To investigate the immunohistochemical features of LOP/C in PTC and to show that this feature is linked to a high association of lymph node metastasis (LNM) at surgery and tumour recurrence. The degree of LOP/C of the PTCs was evaluated histologically using a cut-off value of 20% and the immunohistochemical features of LOP/C were analysed using immunohistochemical staining for E-cadherin, β-catenin and vimentin. The relationship between the LOP/C and the other clinicopathological parameters was analysed. 43 cases of PTC with LOP/C (≥20%) were selected and 27 cases with LOP/C (LOP/C. LOP/C (≥20%) was significantly correlated with extrathyroid invasion (r=0.336, p=0.003), advanced tumour stage (r=0.275, p=0.017), LNM (r=0.389, pLOP/C were independent significant predictors of LNM. LOP/C may be a useful morphological feature of epithelial mesenchymal transition under H&E observation, and it is an important indicator of lymph node metastasis and aggressive clinical behaviour of PTC.

  16. Anti-Cancer Activity of Solanum nigrum (AESN through Suppression of Mitochondrial Function and Epithelial-Mesenchymal Transition (EMT in Breast Cancer Cells

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    Ying-Jang Lai

    2016-04-01

    Full Text Available Chemotherapy is the main approach for treating advanced and recurrent carcinoma, but the clinical performance of chemotherapy is limited by relatively low response rates, drug resistance, and adverse effects that severely affect the quality of life of patients. An association between epithelial-mesenchymal transition (EMT and chemotherapy resistance has been investigated in recent studies. Our recent studies have found that the aqueous extract of Solanum nigrum (AESN is a crucial ingredient in some traditional Chinese medicine formulas for treating various types of cancer patients and exhibits antitumor effects. We evaluated the suppression of EMT in MCF-7 breast cancer cells treated with AESN. The mitochondrial morphology was investigated using Mitotracker Deep-Red FM stain. Our results indicated that AESN markedly inhibited cell viability of MCF-7 breast cancer cells through apoptosis induction and cell cycle arrest mediated by activation of caspase-3 and production of reactive oxygen species. Furthermore, mitochondrial fission was observed in MCF-7 breast cancer cells treated with AESN. In addition to elevation of E-cadherin, downregulations of ZEB1, N-cadherin, and vimentin were found in AESN-treated MCF-7 breast cancer cells. These results suggested that AESN could inhibit EMT of MCF-7 breast cancer cells mediated by attenuation of mitochondrial function. AESN could be potentially beneficial in treating breast cancer cells, and may be of interest for future studies in developing integrative cancer therapy against proliferation, metastasis, and migration of breast cancer cells.

  17. Transforming growth factor-β1 regulates epithelial-mesenchymal transition in association with cancer stem-like cells in a breast cancer cell line.

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    Jia, Yongfeng; Wu, Di; Yun, Fen; Shi, Lin; Luo, Nianrong; Liu, Zhiyue; Shi, Yonghong; Sun, Qinnuan; Jiang, Lili; Wang, Shiqi; Du, Maolin

    2014-01-01

    Epithelial-mesenchymal transition (EMT) is associated with altered connection and junctions between cells and changes in abilities of invasion and migration. In this study, we investigated whether SK-BR-3 breast cancer cells induced to undergo EMT exhibit changes in morphological and invasion abilities after Transforming growth factor β1 (TGF-β1) treatment. Serum-deprived SK-BR-3 cells were treated with TGF-β1 (0, 10 ng/mL) for 24 h. The cells morphological changes were observed and imaged using inverted phase contrast microscope. Scratch experiment and invasion experiment were employed to detect changes of invasion ability, cell-flow experiment was used to assess cell cycle, immunohistochemistry technique was used to detect epithelial and mesenchymal markers after the crawling cells were fixed. Our research reveal that SK-BR-3 cells become larger and more messy, the elongated cells extend pseudopodia, the link of the cells became more loosely and cell gap widened after TGF-β1 treatment. SK-BR-3 cells showed faster growing and improved invasion abilities after TGF-β1 treatment, and reduced G1 phase cells proportion in the total number of cells after the conversion, in contrast the S phase cells accounted for the proportion of the total number of cells increased. These findings indicate that TGF-β1-induced EMT in breast cancer cells may be associated with major alterations in morphological and invasion abilities.

  18. Ginsenoside Rg1 Attenuates Cigarette Smoke-Induced Pulmonary Epithelial-Mesenchymal Transition via Inhibition of the TGF-β1/Smad Pathway

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    Sibin Guan

    2017-01-01

    Full Text Available Epithelial-mesenchymal transition (EMT is a process associated with airway remodeling in chronic obstructive pulmonary disease (COPD, which leads to progressive pulmonary destruction. Panax ginseng is a traditional herbal medicine that has been shown to improve pulmonary function and exercise capacity in patients with COPD. Ginsenoside Rg1 is one of the main active components and was shown to inhibit oxidative stress and inflammation. The present study investigated the hypothesis that ginsenoside Rg1 attenuates EMT in COPD rats induced by cigarette smoke (CS and human bronchial epithelial (HBE cells exposed to cigarette smoke extract (CSE. Our data showed that CS or CSE exposure increased expression of the mesenchymal marker α-smooth muscle actin (α-SMA and decreased expression of the epithelial marker epithelial cadherin (E-cad in both lung tissues and HBE cells, which was markedly suppressed by ginsenoside Rg1. Importantly, CS-induced upregulation of TGF-β1/Smad pathway components, including TGF-β1, TGF-βR1, phospho-Smad2, and phospho-Smad3, was also inhibited by ginsenoside Rg1. Additionally, ginsenoside Rg1 mimicked the effect of SB525334, a TGF-βR1-Smad2/3 inhibitor, on suppression of EMT in CSE-induced HBE cells. Collectively, we concluded that ginsenoside Rg1 alleviates CS-induced pulmonary EMT, in both COPD rats and HBE cells, via inhibition of the TGF-β1/Smad pathway.

  19. Glutamine inhibits CCl4 induced liver fibrosis in mice and TGF-β1 mediated epithelial-mesenchymal transition in mouse hepatocytes.

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    Shrestha, Nirajan; Chand, Lokendra; Han, Myung Kwan; Lee, Seung Ok; Kim, Chan Young; Jeong, Yeon Jun

    2016-07-01

    Glutamine, traditionally a non-essential amino acid, now has been considered as essential in serious illness and injury. It is a major precursor for glutathione synthesis. However, the anti-fibrotic effect of glutamine and its molecular mechanism in experimental liver fibrosis have not been explored. In the present study we aimed to examine the potential role of glutamine in carbon tetrachloride (CCl4) induced liver fibrosis and TGF-β1 mediated epithelial mesenchymal transition (EMT) and apoptosis in mouse hepatocytes. Liver fibrosis was induced by intraperitoneal injection of CCl4 three times a week for 10 weeks. Glutamine treatment effectively attenuated liver injury and oxidative stress. Collagen content was significantly decreased in liver sections of glutamine treated mice compared to CCl4 model mice. Furthermore, glutamine decreased expression level of α-SMA and TGF-β in liver tissue. Our in vitro study showed that TGF-β1 treatment in hepatocytes resulted in loss of E-cadherin and increased expression of mesenchymal markers and EMT related transcription factor. In addition, TGF-β1 increased the expression of apoptotic markers. However, glutamine interestingly suppressed TGF-β1 mediated EMT and apoptosis. In conclusion, our results suggest that glutamine ameliorates CCl4 induced liver fibrosis and suppresses TGF-β1 induced EMT progression and apoptosis.

  20. Calpain 1 regulates TGF-β1-induced epithelial-mesenchymal transition in human lung epithelial cells via PI3K/Akt signaling pathway

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    Tan, Wei-Jun; Tan, Qiu-Yue; Wang, Ting; Lian, Min; Zhang, Li; Cheng, Zhen-Shun

    2017-01-01

    Cell proliferation, transformation, and epithelial-mesenchymal transition (EMT) are key processes involved in the development of idiopathic pulmonary fibrosis (IPF). This study investigated the regulatory factors and signaling pathways that mediate EMT in the human type II alveolar epithelial A549 cell line. A549 cells were cultured in RPMI-1640 medium and allocated to the following four groups: blank control group or treated with transforming growth factor-β1 (TGF-β1), TGF-β1 + PD 150606 (a calpain 1 inhibitor), or PD 150606. We examined E-cadherin (E-cad), α-smooth muscle actin (α-SMA), and calpain 1 mRNA transcript and protein expression levels in these four groups by performing RT-PCR and western blot analyses. The results indicated that TGF-β1 treatment significantly downregulated E-cad and upregulated α-SMA expression compared with that of the blank control group (Pcells. However, TGF-β1-induced ETM was not correlated with the ERK and JNK signaling pathways. These combined results indicate that calpain 1 could regulate EMT in TGF-β1-treated A549 epithelial cells via the PI3K/Akt signaling pathway.

  1. FGF19 promotes epithelial-mesenchymal transition in hepatocellular carcinoma cells by modulating the GSK3β/β- catenin signaling cascade via FGFR4 activation.

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    Zhao, Huakan; Lv, Fenglin; Liang, Guizhao; Huang, Xiaobin; Wu, Gang; Zhang, Wenfa; Yu, Le; Shi, Lei; Teng, Yong

    2016-03-22

    Compelling evidence suggests that the epithelial-mesenchymal transition (EMT) correlates with aggressiveness of tumors and poor survival. FGF19 has been shown to be involved in EMT in cholangiocarcinoma and colorectal cancer, however, molecular mechanisms underlying FGF19-induced EMT process in hepatocellular carcinoma (HCC) remain largely unknown. Here, we show the expression of FGF19 is significantly elevated and negatively associated with the expression of E-cadherin in HCC tissues and cell lines. Ectopic FGF19 expression promotes EMT and invasion in epithelial-like HCC cells through repression of E-cadherin expression, whereas FGF19 knockdown enhances E-cadherin expression and hence diminishes EMT traits in mesenchymal-like HCC cells, suggesting FGF19 exerts its tumor progressing functions as an EMT inducer. Interestingly, depletion of FGF19 cannot abrogate EMT traits in the presence of GSK3β inhibitors. Furthermore, FGF19-induced EMT can be markedly attenuated when FGFR4 is knocked out. These observations clearly indicate that FGFR4/GSK3β/β-catenin axis may play a pivotal role in FGF19-induced EMT in HCC cells. As FGF19 and its specific receptor FGFR4 are frequently amplified in HCC cells, selective targeting this signaling node may lend insights into a potential effective therapeutic approach for blocking metastasis of HCC.

  2. Valproic acid (VPA) promotes the epithelial mesenchymal transition of hepatocarcinoma cells via transcriptional and post-transcriptional up regulation of Snail.

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    Wu, Lei; Feng, Hua; Hu, Jinhua; Tian, Xiangguo; Zhang, Chunqing

    2016-12-01

    Due to the low cost and favorable safety profile, valproic acid (VPA) has been considered as a potential candidate drug for therapy of various cancers. Our present study revealed that VPA, at the concentration (1mM) which has no effect on cell proliferation, can significantly increase the in vitro migration and invasion of hepatocarcinoma (HCC) HepG2 and Huh7 cells via induction of epithelial mesenchymal transition (EMT). VPA treatment can significantly increase the mRNA and protein expression of Snail, the key transcription factor of EMT. While knockdown of Snail can abolish VPA induced EMT of HCC cells. It suggested that Snail is essential for VPA induced EMT of HCC cells. VPA treatment also increased the phosphorylation of NF-κB p65. BAY 11-7082, the inhibitor of NF-κB, can significantly abolish VPA induced up regulation of Snail mRNA. Furthermore, VPA can increase the protein expression of Snail since 1h treatment via up regulation of half-lives of Snail protein. The increased protein stabilization of Snail can be attributed to VPA induced phosphorylation of Akt and GSK-3β. Collectively, our present study revealed that VPA can promote the EMT of HCC cells via up regulation of Snail through activation of NF-κB and Akt/GSK-3β signals. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  3. Overexpression of snail induces epithelial-mesenchymal transition and a cancer stem cell-like phenotype in human colorectal cancer cells.

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    Fan, Fan; Samuel, Shaija; Evans, Kurt W; Lu, Jia; Xia, Ling; Zhou, Yunfei; Sceusi, Eric; Tozzi, Federico; Ye, Xiang-Cang; Mani, Sendurai A; Ellis, Lee M

    2012-08-01

    Epithelial-mesenchymal transition (EMT) is a critical process providing tumor cells with the ability to migrate and escape from the primary tumor and metastasize to distant sites. Recently, EMT was shown to be associated with the cancer stem cell (CSC) phenotype in breast cancer. Snail is a transcription factor that mediates EMT in a number of tumor types, including colorectal cancer (CRC). Our study was done to determine the role of Snail in mediating EMT and CSC function in CRC. Human CRC specimens were stained for Snail expression, and human CRC cell lines were transduced with a retroviral Snail construct or vector control. Cell proliferation and chemosensitivity to oxaliplatin of the infected cells were determined by the MTT (colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Migration and invasion were determined in vitro using modified Boyden chamber assays. EMT and putative CSC markers were analyzed using Western blotting. Intravenous injection of tumor cells was done to evaluate their metastatic potential in mice. Snail was overexpressed in human CRC surgical specimens. This overexpression induced EMT and a CSC-like phenotype in human CRC cells and enhanced cell migration and invasion (P Snail overexpression also led to an increase in metastasis formation in vivo (P Snail-overexpressing CRC cells were more chemoresistant to oxaliplatin than control cells. Increased Snail expression induces EMT and the CSC-like phenotype in CRC cells, which enhance cancer cell invasion and chemoresistance. Thus, Snail is a potential therapeutic target in metastatic CRC.

  4. Stabilization of Snail through AKT/GSK-3β signaling pathway is required for TNF-α-induced epithelial-mesenchymal transition in prostate cancer PC3 cells.

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    Wang, Hao; Fang, Rui; Wang, Xian-Feng; Zhang, Fan; Chen, Dan-Yang; Zhou, Binhua; Wang, Hong-Sheng; Cai, Shao-Hui; Du, Jun

    2013-08-15

    Metastasis induced by chronic inflammation has been considered as a major challenge during cancer therapy. Epithelial-mesenchymal transition (EMT) is associated with cancer invasion and metastasis promoted by pro-inflammatory cytokine TNFα. However, the mechanisms underlying TNFα-induced EMT in prostate cancer cells is not entirely clear. Here we showed that EMT induced by longstanding stimulation with TNFα in prostate cancer PC3 cells is mediated by up-regulation of the transcriptional repressor Snail. TNFα-mediated EMT was characterized by acquiring mesenchymal fusiform morphology, increasing the expression of Vimentin and decreasing the expression of E-cadherin. Exposure to TNFα increased the expression of transcription factor Snail via post-transcriptional regulation process and induced Snail nuclear localization in PC3 cells. Moreover, overexpressed Snail in PC3 cells induced EMT. Conversely, suppressing Snail expression abrogated TNFα-induced EMT, suggesting that Snail plays a crucial role in TNFα-induced EMT in prostate cancer cells. Finally, we showed that TNFα time-dependently activated NF-κB, AKT, ERK, p38 MAPK signaling pathways, and elevated Snail stability by activating AKT pathway that subsequently inhibited GSK-3β activity. Taken together, these results reveal that stabilization of Snail via AKT/GSK-3β signaling pathway is required for TNFα-induced EMT in prostate cancer cells. This study offers a better understanding of TNFα-induced metastasis and provides an effective therapeutic strategy for prostate cancer treatment. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Ginsenoside 20(S-Rg3 targets HIF-1α to block hypoxia-induced epithelial-mesenchymal transition in ovarian cancer cells.

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    Ting Liu

    Full Text Available The prognosis of patients with ovarian cancer has remained poor mainly because of aggressive cancer progression. Since epithelial-mesenchymal transition (EMT is an important mechanism mediating invasion and metastasis of cancer cells, targeting the EMT process with more efficacious and less toxic compounds to inhibit metastasis is of great therapeutic value for the treatment of ovarian cancer. We have found for the first time that the ginsenoside 20(S-Rg3, a pharmacologically active component of the traditional Chinese herb Panax ginseng, potently blocks hypoxia-induced EMT of ovarian cancer cells in vitro and in vivo. Mechanistic studies confirm the mode of action of 20(S-Rg3, which reduces the expression of hypoxia-inducible factor 1α (HIF-1α by activating the ubiquitin-proteasome pathway to promote HIF-1α degradation. A decrease in HIF-1α in turn leads to up-regulation, via transcriptional suppression of Snail, of the epithelial cell-specific marker E-cadherin and down-regulation of the mesenchymal cell-specific marker vimentin under hypoxic conditions. Importantly, 20(S-Rg3 effectively inhibits EMT in nude mouse xenograft models of ovarian cancer, promising a novel therapeutic agent for anticancer therapy.

  6. miR-143 suppresses epithelial-mesenchymal transition and inhibits tumor growth of breast cancer through down-regulation of ERK5.

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    Zhai, Limin; Ma, Chuanxiang; Li, Wentong; Yang, Shuo; Liu, Zhijun

    2016-12-01

    Epithelial-mesenchymal transition (EMT) plays a pivotal role in the development of cancer invasion and metastasis. Many studies have significantly enhanced the knowledge on EMT through the characterization of microRNAs (miRNAs) influencing the signaling pathways and downstream events that define EMT on a molecular level. In this study, we found that miR-143 suppressed EMT. Up-regulating miR-143 enhanced E-cadherin-mediated cell-cell adhesion ability, reduced mesenchymal markers, and decreased cell proliferation, migration, and invasion in vitro. In vivo, the xenograft mouse model also unveiled the suppressive effects of miR-143 on tumor growth. Additionally, we demonstrated that up-regulating extracellular signal regulated kinase 5 (ERK5) was associated with poor prognosis of breast cancer patients. Moreover, we observed an inverse correlation between miR-143 and ERK5 in breast cancer tissues. miR-143 directly targeted seed sequences in the 3'-untranslated regions of ERK5. Furthermore, we revealed that the downstream molecules of glycogen synthase kinase 3 beta (GSK-3β)/Snail signaling were involved in EMT and modulated by ERK5. In summary, our findings demonstrated that miR-143 down-regulated its target ERK5, leading to the suppression of EMT induced by GSK-3β/Snail signaling of breast cancer. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

  7. Epithelial mesenchymal transition and pancreatic tumor initiating CD44+/EpCAM+ cells are inhibited by γ-secretase inhibitor IX.

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    Vindhya Palagani

    Full Text Available Pancreatic ductal adenocarcinoma (PDAC is an aggressive disease with a high rate of metastasis. Recent studies have indicated that the Notch signalling pathway is important in PDAC initiation and maintenance, although the specific cell biological roles of the pathway remain to be established. Here we sought to examine this question in established pancreatic cancer cell lines using the γ-secretase inhibitor IX (GSI IX to inactivate Notch. Based on the known roles of Notch in development and stem cell biology, we focused on effects on epithelial mesenchymal transition (EMT and on pancreatic tumor initiating CD44+/EpCAM+ cells. We analyzed the effect of the GSI IX on growth and epithelial plasticity of human pancreatic cancer cell lines, and on the tumorigenicity of pancreatic tumor initiating CD44+/EpCAM+ cells. Notably, apoptosis was induced after GSI IX treatment and EMT markers were selectively targeted. Furthermore, under GSI IX treatment, decline in the growth of pancreatic tumor initiating CD44+/EpCAM+ cells was observed in vitro and in a xenograft mouse model. This study demonstrates a central role of Notch signalling pathway in pancreatic cancer pathogenesis and identifies an effective approach to inhibit selectively EMT and suppress tumorigenesis by eliminating pancreatic tumor initiating CD44+/EpCAM+ cells.

  8. Inhibition of invasion and metastasis of MHCC97H cells by expression of snake venom cystatin through reduction of proteinases activity and epithelial-mesenchymal transition.

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    Tang, Nanhong; Xie, Qun; Wang, Xiaoqian; Li, Xiujin; Chen, Yanlin; Lin, Xu; Lin, Jianyin

    2011-05-01

    Snake venom cystatin (sv-cystatin) is a member of the cystatin family of cysteine protease inhibitors. To further evaluate the possibility of sv-cystatin in cancer therapy, this study examined the effects of sv-cystatin on the invasion and metastasis of liver cancer cells (MHCC97H) in vitro and in vivo as well as the underlying mechanism. sv-cystatin cDNA was transfected into MHCC97H cells and the anti-invasion and antimetastasis effects of sv-cystatin were determined using migration and matrigel invasion assays and a lung-metastasis mice model. The results suggest that sv-cyst clone (sv-cystatin expression in MHCC97H cells) delayed the invasion and metastasis in vitro and in vivo compared to the parental, mock and si-sv-cyst clone cells (inhibited sv-cystatin expression by siRNA). The decreased activities of cathepsin B, MMP-2 and MMP-9 and EMT change index including higher E-cadherin, lower N-cadherin and decreased Twist activity were observed in the sv-cyst clone, which contributes to the change in invasion and metastasis ability of MHCC97H cells. This study provides evidence that expression of the sv-cystatin gene in MHCC97H cells inhibits tumor cell invasion and metastasis through the reduction of the proteinases activity and Epithelial-Mesenchymal Transition (EMT), which might contribute to the anticancer research of the sv-cystatin protein.

  9. Knockdown of Collagen Triple Helix Repeat Containing 1 (CTHRC1) Inhibits Epithelial-Mesenchymal Transition and Cellular Migration in Glioblastoma Cells.

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    Liu, Jianpeng; Li, Wei; Liu, Shunshun; Zheng, Xu; Shi, Lin; Zhang, Weitao; Yang, Hongfa

    2017-01-26

    Collagen triple helix repeat containing 1 (CTHRC1), an extracellular matrix-related protein, has been found to be upregulated in many solid tumors and contributes to tumorigenesis. We found that CTHRC1 is overexpressed in glioblastoma tissues and cells. By using the technique of RNA interference, the expression of CTHRC1 in the human glioblastoma U-87MG cell line was downregulated, and the proliferation and migration of U-87MG cells were examined. The results showed that the knockdown of CTHRC1 exerts inhibitory effects on the proliferation and migration ability of U-87MG cells. Knockdown of CTHRC1 expression in U-87MG cells resulted in upregulation in the expression of E-cadherin and downregulation in the expression of N-cadherin, SNAIL, and Slug, suggesting that CTHRC1 inhibits glioblastoma cell migration by suppressing epithelial-mesenchymal transition (EMT). Knockdown of CTHRC1 led to remarkably decreased β-catenin protein levels in the nucleus. These results indicate that CTHRC1 might play an important role in the development of glioblastoma and offer a candidate molecular target for glioblastoma prevention and therapy.

  10. Epidermal growth factor promotes protein degradation of epithelial protein lost in neoplasm (EPLIN), a putative metastasis suppressor, during epithelial-mesenchymal transition.

    Science.gov (United States)

    Zhang, Shumin; Wang, Xu; Iqbal, Shareen; Wang, Yanru; Osunkoya, Adeboye O; Chen, Zhengjia; Chen, Zhuo; Shin, Dong M; Yuan, Hongwei; Wang, Yongqiang A; Zhau, Haiyen E; Chung, Leland W K; Ritenour, Chad; Kucuk, Omer; Wu, Daqing

    2013-01-18

    Aberrant expression of EGF receptors has been associated with hormone-refractory and metastatic prostate cancer (PCa). However, the molecular mechanism for EGF signaling in promoting PCa metastasis remains elusive. Using experimental models of PCa metastasis, we demonstrated that EGF could induce robust epithelial-mesenchymal transition (EMT) and increase invasiveness. Interestingly, EGF was found to be capable of promoting protein turnover of epithelial protein lost in neoplasm (EPLIN), a putative suppressor of EMT and tumor metastasis. Mechanistic study revealed that EGF could activate the phosphorylation, ubiquitination, and degradation of EPLIN through an extracellular signal-regulated kinase 1/2 (ERK1/2)-dependent signaling cascade. Pharmacological inhibition of the ERK1/2 pathway effectively antagonized EGF-induced EPLIN degradation. Two serine residues, i.e. serine 362 and serine 604, were identified as putative ERK1/2 phosphorylation sites in human EPLIN, whose point mutation rendered resistance to EGF-induced protein turnover. This study elucidated a novel molecular mechanism for EGF regulation of EMT and invasiveness in PCa cells, indicating that blockade of EGF signaling could be beneficial in preventing and retarding PCa metastasis at early stages.

  11. Knockdown of Snail inhibits epithelial-mesenchymal transition of human laryngeal squamous cell carcinoma Hep-2 cells through VDR signaling pathway.

    Science.gov (United States)

    Zhao, Xue; Yu, Dan; Yang, Jingpu; Xue, Kai; Liu, Yan; Jin, Chunshun

    2017-08-14

    It has been well-documented that Snail plays a decisive role in various tumors. However, the direct effect of Snail on laryngeal squamous cell carcinoma (LSCC) has not been elaborated. In this study, we firstly detected the expression of Snail in 14 samples of patients with LSCC and found that its content was high in cancer tissues compared with adjacent tissues. Then we established LSCC Hep-2 cells with Snail silencing and validated the knockdown efficiency by western blotting and real-time PCR. Results showed that silencing of Snail significantly inhibited the ability of adhesion, migration, and invasion of Hep-2 cells. Further study revealed that knockdown of Snail suppressed the epithelial-mesenchymal transition process of Hep-2 cells, as evidenced by downregulation of matrix metallopeptidase (MMP)-2, MMP-9, integrin subunit beta 1 (ITGβ1), β-catenin, vimentin, N-cadherin, and fibronectin (FN), while upregulation of vitamin D receptor (VDR) and E-cadherin. Additionally, transfection with the small interfering RNA of VDR reversed the effect induced by Snail silencing in Hep-2 cells. Taken together, these results demonstrate that knockdown of Snail can inhibit the EMT process of LSCC cells through VDR signaling pathway in vitro.

  12. Mechanisms of AEG-1 and CXCR4 gene expression regulating the epithelial-mesenchymal transition pathway involved in brain metastases of breast cancer.

    Science.gov (United States)

    Chen, Yanjun; Wang, Xinjun

    2017-01-01

    To study the astrocyte elevated gene (AEG)-1 in breast cancer and the mechanism of the chemokine receptor CXCR4 regulating the epithelial-mesenchymal transition (EMT) involved in brain metastases of breast cancer. A total of 20 breast cancer patients with and 40 without brain metastases were recruited. The expressions of AEG-1, CXCR4, E-cadherin, N-cadherin and α-SMA were detected by immunohistochemical staining, real-time (RT) quantitative (q) PCR and Western blot respectively in cancer and adjacent normal tissues. The expressions detected in the adjacent normal tissues from both groups showed no significant difference (p>0.05). In the group with brain metastases, AEG-1, CXCR4, N-cadherin, α-SMA, mRNA and the relative expression level of protein were higher than those of patients without brain metastases, while E-cadherin showed the opposite trend. AEG-1 and CXCR4 activate and regulate the EMT pathway to participate in brain metastases.

  13. Reversibility of epithelial-mesenchymal transition (EMT) induced in breast cancer cells by activation of urokinase receptor-dependent cell signaling.

    Science.gov (United States)

    Jo, Minji; Lester, Robin D; Montel, Valerie; Eastman, Boryana; Takimoto, Shinako; Gonias, Steven L

    2009-08-21

    Hypoxia induces expression of the urokinase receptor (uPAR) and activates uPAR-dependent cell signaling in cancer cells. This process promotes epithelial-mesenchymal transition (EMT). uPAR overexpression in cancer cells also promotes EMT. In this study, we tested whether uPAR may be targeted to reverse cancer cell EMT. When MDA-MB 468 breast cancer cells were cultured in 1% O(2), uPAR expression increased, as anticipated. Cell-cell junctions were disrupted, vimentin expression increased, and E-cadherin was lost from cell surfaces, indicating EMT. Transferring these cells back to 21% O(2) decreased uPAR expression and reversed the signs of EMT. In uPAR-overexpressing MDA-MB 468 cells, EMT was reversed by silencing expression of endogenously produced urokinase-type plasminogen activator (uPA), which is necessary for uPAR-dependent cell signaling, or by targeting uPAR-activated cell signaling factors, including phosphatidylinositol 3-kinase, Src family kinases, and extracellular signal-regulated kinase. MDA-MB 231 breast cancer cells express high levels of uPA and uPAR and demonstrate mesenchymal cell morphology under normoxic culture conditions (21% O(2)). Silencing uPA expression in MDA-MB-231 cells decreased expression of vimentin and Snail, and induced changes in morphology characteristic of epithelial cells. These results demonstrate that uPAR-initiated cell signaling may be targeted to reverse EMT in cancer.

  14. Inhibition of SK4 Potassium Channels Suppresses Cell Proliferation, Migration and the Epithelial-Mesenchymal Transition in Triple-Negative Breast Cancer Cells.

    Science.gov (United States)

    Zhang, Panshi; Yang, Xiaowei; Yin, Qian; Yi, Jilin; Shen, Wenzhuang; Zhao, Lu; Zhu, Zhi; Liu, Jinwen

    2016-01-01

    Treatments for triple-negative breast cancer (TNBC) are limited; intermediate-conductance calcium-activated potassium (SK4) channels are closely involved in tumor progression, but little is known about these channels in TNBC. We aimed to investigate whether SK4 channels affect TNBC. First, by immunohistochemistry (IHC) and western blotting (WB), increased SK4 protein expression in breast tumor tissues was detected relative to that in non-tumor breast tissues, but there was no apparent expression difference between various subtypes of breast cancer (p>0.05). Next, functional SK4 channels were detected in the TNBC cell line MDA-MB-231 using WB, real-time PCR, immunofluorescence and patch-clamp recording. By employing SK4 specific siRNAs and blockers, including TRAM-34 and clotrimazole, in combination with an MTT assay, a colony-formation assay, flow cytometry and a cell motility assay, we found that the suppression of SK4 channels significantly inhibited cell proliferation and migration and promoted apoptosis in MDA-MB-231 cells (pMDA-MB-231 cells to undergo the epithelial-mesenchymal transition (EMT) and to show increased SK4 mRNA expression. In addition, the down-regulation of SK4 expression inhibited the EMT markers Vimentin and Snail1. Collectively, our findings suggest that SK4 channels are expressed in TNBC and are involved in the proliferation, apoptosis, migration and EMT processes of TNBC cells.

  15. 3D models of epithelial-mesenchymal transition in breast cancer metastasis: high-throughput screening assay development, validation, and pilot screen.

    Science.gov (United States)

    Li, Qun; Chen, Chaoyu; Kapadia, Amit; Zhou, Qiong; Harper, Mary Kay; Schaack, Jerome; LaBarbera, Daniel V

    2011-02-01

    Despite advancements in therapies developed for the treatment of cancer, patient prognosis and mortality rates have improved minimally, and metastasis remains the primary cause of cancer mortality worldwide. An underlying mechanism promoting metastasis in many types of cancer is epithelial-mesenchymal transition (EMT). Here the authors report a novel 3D model of EMT and metastatic breast cancer suitable for high-throughput screening (HTS) drug discovery. The primary assay incorporates the expression of the prognostic biomarker vimentin, as a luciferase reporter of EMT, in basil-like/triple-negative MDA-MB-231 breast carcinoma spheroids. Using this model, the authors developed a number of known antitumor agents as control modulators of EMT. U0126, PKC412, PF2341066, dasatinib, and axitinib downregulated vimentin expression by 70% to 90% as compared to untreated spheroids. Counterassays were developed to measure spheroid viability and the invasive potential of MDA-MB-231 spheroids after small-molecule treatment and used to confirm hits from primary screening. Finally, the authors conducted a pilot screen to validate this model for HTS using a purified library of marine secondary metabolites. From 230 compounds screened, they obtained a Z' score of 0.64, indicative of an excellent assay, and confirmed 4 hits, including isonaamidine B, papuamine, mycalolide E, and jaspamide. This HTS model demonstrates the potential to identify small-molecule modulators of EMT that could be used to discover novel antimetastatic agents for the treatment of cancer.

  16. Cardiotoxin III Inhibits Hepatocyte Growth Factor-Induced Epithelial-Mesenchymal Transition and Suppresses Invasion of MDA-MB-231 Cells.

    Science.gov (United States)

    Tsai, Pei-Chien; Fu, Yaw-Syan; Chang, Long-Sen; Lin, Shinne-Ren

    2016-01-01

    The epithelial-mesenchymal transition (EMT) is the first step required for breast cancer to initiate metastasis. In this study, hepatocyte growth factor (HGF) was used as a metastatic inducer of MDA-MB-231 cells. Cardiotoxin III (CTX III) inhibited HGF-induced morphological changes and upregulation of E-cadherin with the concomitant decrease in N-cadherin and Vimentin protein levels, resulting in inhibition of cell migration and invasion. CTX III-induced downregulation of transcription factors, Snail, Twist, and Slug, in MDA-MB-231 cells. CTX III suppressed c-Met phosphorylation and downstream activation of phosphatidylinositol 3-kinase (PI3K)/Akt and extracellular signal-regulated kinase (ERK)1/2. The c-Met specific inhibitor PHA665752 attenuated ERK1/2 and Akt phosphorylation, cell migration and invasion, as well as the expressional changes of EMT markers induced by HGF. Taken together, our data suggest that CTX III suppresses HGF/c-Met-induced cell migration and invasion by reversing EMT, which involves the inactivation of the HGF/c-Met-mediated ERK1/2 and PI3K/Akt pathways in MDA-MB-231 cells.

  17. Suberoylanilide hydroxamic acid (SAHA) promotes the epithelial mesenchymal transition of triple negative breast cancer cells via HDAC8/FOXA1 signals.

    Science.gov (United States)

    Wu, Shao; Luo, Zhi; Yu, Peng-Jiu; Xie, Hui; He, Yu-Wen

    2016-01-01

    Inhibitor of histone deacetylases (HDACIs) have great therapeutic value for triple negative breast cancer (TNBC) patients. Interestingly, our present study reveals that suberoyl anilide hydroxamic acid (SAHA), one of the most advanced pan-HDAC inhibitor, can obviously promote in vitro motility of MDA-MB-231 and BT-549 cells via induction of epithelial-mesenchymal transition (EMT). SAHA treatment significantly down-regulates the expression of epithelial markers E-cadherin (E-Cad) while up-regulates the mesenchymal markers N-cadherin (N-Cad), vimentin (Vim) and fibronectin (FN). However, SAHA has no effect on the expression and nuclear translocation of EMT related transcription factors including Snail, Slug, Twist and ZEB. While SAHA treatment down-regulates the protein and mRNA expression of FOXA1 and then decreases its nuclear translocation. Over-expression of FOXA1 markedly attenuates SAHA induced EMT of TNBC cells. Further, silence of HDAC8, while not HDAC6, alleviates the down-regulation of FOXA1 and up-regulation of N-Cad and Vim in MDA-MB-231 cells treated with SAHA. Collectively, our present study reveals that SAHA can promote EMT of TNBC cells via HDAC8/FOXA1 signals, which suggests that more attention should be paid when SAHA is used as anti-cancer agent for cancer treatment.

  18. HAb18G/CD147 is involved in TGF-β-induced epithelial-mesenchymal transition and hepatocellular carcinoma invasion.

    Science.gov (United States)

    Ru, Ning-Yu; Wu, Jiao; Chen, Zhi-Nan; Bian, Huijie

    2015-01-01

    Epithelial-mesenchymal transition (EMT) induced by the transforming growth factor beta (TGF-β) is involved in hepatocarcinogenesis and hepatocellular carcinoma (HCC) metastasis. HAb18G/CD147, a member of the immunoglobulin family, plays an important role in tumor invasion and metastasis. HAb18G/CD147 promotes EMT of hepatocytes through TGF-β signaling and is transcriptionally regulated by Slug. We investigated the role of HAb18G/CD147 in TGF-β-induced EMT in HCC invasion. Two human HCC cell lines, SMMC-7721 and HepG2, were used to determine the role of HAb18G/CD147 in EMT. Upregulation of HAb18G/CD147 induced by the high doses of TGF-β1 in SMMC-7721 (5 ng/mL) and HepG2 cells (10 ng/mL) (P CD147 upregulation was coupled with upregulation of Snail1 and Slug. CD147 knockout significantly decreased the expression of N-cadherin and vimentin, and colony formation ability of SMMC-7721 cells. TGF-β1 enhanced the migration capacity of SMMC-7721 cells, which was markedly attenuated by CD147 knockdown. Thus, HAb18G/CD147 is involved in TGF-β-induced EMT and HCC invasion.

  19. Long noncoding RNA LINC01186, regulated by TGF-β/SMAD3, inhibits migration and invasion through Epithelial-Mesenchymal-Transition in lung cancer.

    Science.gov (United States)

    Hao, Yajing; Yang, Xinling; Zhang, Dongdong; Luo, Jianjun; Chen, Runsheng

    2017-04-15

    Accumulating evidence suggests that long noncoding RNAs (lncRNAs) are crucial regulators of the Epithelial-Mesenchymal-Transition (EMT). TGF-β signaling is a major inducer of EMT and can facilitate lung cancer metastasis. However, the role of lncRNAs in this process remains largely unknown. Here, we have identified 291 lncRNAs which were differentially expressed in lung cancer tissues compared with adjacent normal tissues. Of these, the gene body or vicinity of 19 transcripts were also bound by SMAD3. The expression of LINC01186 was significantly decreased in A549 cells treated with TGF-β1. Furthermore, LINC01186 was stably down-regulated in lung cancer tissues compared with normal tissues in TCGA data sets and another published lung cancer data sets. The bioinformatics analysis suggested that LINC01186 was associated with TGF-β and might participate in EMT process. Moreover, knocking-down LINC01186 promoted cell migration and invasion, whereas, LINC01186 overexpression prevented cell metastasis. Importantly, LINC01186 expression was regulated by SMAD3. And LINC01186 affected several EMT markers expression. These findings suggest that LINC01186, a mediator of TGF-β signaling, can play a significant role in the regulation of EMT and lung cancer cell migration and invasion.

  20. MicroRNA-145 Inhibits Cell Migration and Invasion and Regulates Epithelial-Mesenchymal Transition (EMT) by Targeting Connective Tissue Growth Factor (CTGF) in Esophageal Squamous Cell Carcinoma.

    Science.gov (United States)

    Han, Qiang; Zhang, Hua-Yong; Zhong, Bei-Long; Wang, Xiao-Jing; Zhang, Bing; Chen, Hua

    2016-10-23

    BACKGROUND This study investigated the mechanism of miR-145 in targeting connective tissue growth factor (CTGF), which affects the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of ESCC cells. MATERIAL AND METHODS A total of 50 ESCC tissues and their corresponding normal adjacent esophageal tissue samples were collected. Then, miR-145 expression in both ESCC clinical specimens and cell lines was detected using quantitative real-time PCR. CTGF protein was detected using immunohistochemistry. Dual luciferase reporter gene assay was employed to assess the effect of miR-145 on the 3'UTR luciferase activity of CTGF. Eca109 cells were transfected with miR-145 mimics and CTGF siRNA, respectively, and changes in cellular proliferation, migration, and invasion were detected via MTT assay, wound-healing assay, and Transwell assay, respectively. Western blotting assay was used to detect the expression of marker genes related to EMT. RESULTS MiR-145 was significantly down-regulated in ESCC tissues and cell lines compared with normal tissues and cell lines (Ptissues was than in normal adjacent esophageal tissues (Ptissues and cell lines, while the protein expression of CTGF exhibited the opposite trend. MiR-145 inhibited the proliferation, migration, invasiveness, and the EMT process of ESCC cells through targeted regulation of CTGF expression.

  1. Upregulation of long noncoding RNA SPRY4-IT1 promotes metastasis of esophageal squamous cell carcinoma via induction of epithelial-mesenchymal transition.

    Science.gov (United States)

    Zhang, Chun-Yang; Li, Ren-Ke; Qi, Yu; Li, Xiang-Nan; Yang, Yang; Liu, Dong-Lei; Zhao, Jia; Zhu, Deng-Yan; Wu, Kai; Zhou, Xu-Dong; Zhao, Song

    2016-10-01

    Esophageal squamous cell carcinoma (ESCC) is one of the prevalent and deadly cancers worldwide, especially in Eastern Asia. Recent studies show that long noncoding RNAs (lncRNAs) have critical roles in diverse biological processes, including tumorigenesis. In the present study, we find that the expression of lncRNA SPRY4-IT1 is significantly upregulated in ESCC cell lines as compared with human esophageal epithelial cell line HEEC. Overexpression of SPRY4-IT1 can increase in vitro motility of ESCC cells via induction of epithelial-mesenchymal transition (EMT), which is characterized by increasing the expression of vimentin (Vim) and fibronectin (FN) with a concomitant decrease of E-cadherin (E-Cad) and ZO-1, while silencing of SPRY4-IT1 significantly inhibits the in vitro motility of ESCC cells. Further, the knockdown of SPRY4-IT1 also significantly attenuates TFG-β-induced EMT of ESCC cells. Further, lncRNA SPRY4-IT1 can directly increase the transcription, expression, and nuclear localization of Snail, one key transcription factor during the EMT processes of cancer cells, while siRNA-mediated specific knockdown of Snail can significantly attenuate SPRY4-IT1-induced EMT of ESCC cells. Our results suggest that lncRNA SPRY4-IT1 might be considered as a novel oncogene involved in ESCC progression.

  2. Long Non-Coding RNA MALAT1 Mediates Transforming Growth Factor Beta1-Induced Epithelial-Mesenchymal Transition of Retinal Pigment Epithelial Cells.

    Directory of Open Access Journals (Sweden)

    Shuai Yang

    Full Text Available To study the role of long non-coding RNA (lncRNA MALAT1 in transforming growth factor beta 1 (TGF-β1-induced epithelial-mesenchymal transition (EMT of retinal pigment epithelial (RPE cells.ARPE-19 cells were cultured and exposed to TGF-β1. The EMT of APRE-19 cells is confirmed by morphological change, as well as the increased expression of alpha-smooth muscle actin (αSMA and fibronectin, and the down-regulation of E-cadherin and Zona occludin-1(ZO-1 at both mRNA and protein levels. The expression of lncRNA MALAT1 in RPE cells were detected by quantitative real-time PCR. Knockdown of MALAT1 was achieved by transfecting a small interfering RNA (SiRNA. The effect of inhibition of MALAT1 on EMT, migration, proliferation, and TGFβ signalings were observed. MALAT1 expression was also detected in primary RPE cells incubated with proliferative vitreoretinopathy (PVR vitreous samples.The expression of MALAT1 is significantly increased in RPE cells incubated with TGFβ1. MALAT1 silencing attenuates TGFβ1-induced EMT, migration, and proliferation of RPE cells, at least partially through activating Smad2/3 signaling. MALAT1 is also significantly increased in primary RPE cells incubated with PVR vitreous samples.LncRNA MALAT1 is involved in TGFβ1-induced EMT of human RPE cells and provides new understandings for the pathogenesis of PVR.

  3. Overexpression of forkhead Box C2 promotes tumor metastasis and indicates poor prognosis in colon cancer via regulating epithelial-mesenchymal transition.

    Science.gov (United States)

    Li, Qingguo; Wu, Jitao; Wei, Ping; Xu, Ye; Zhuo, Changhua; Wang, Yuwei; Li, Dawei; Cai, Sanjun

    2015-01-01

    Forkhead box protein C2 (FOXC2) plays a vital role in carcinogenesis; however, its significance and prognostic value in colon cancer remain unclear. In this study, FOXC2 expression was analyzed in a tissue microarray (TMA) containing 185 samples of primary colon cancer tumor samples and in human colon cancer cell lines. The effect of FOXC2 on cell proliferation, tumorigenesis, and metastasis was examined in vitro and in vivo. FOXC2 was overexpressed in human colon cancer cells and tissues, and correlated with colon cancer progression and patient survival. Functional study demonstrated that FOXC2 promoted cell growth, cell migration, and tumor formation in nude mice, whereas knockdown of FOXC2 by short hairpin RNA (shRNAs) significantly suppressed cell growth, cell migration and tumor formation. Further study found that FOXC2 enhanced AKT activity with subsequent GSK-3β phosphorylation and Snail stabilization, and then induced epithelial-mesenchymal transition (EMT) and promoted tumor invasion and metastasis. Collectively, FOXC2 promotes colon cancer metastasis by facilitating EMT and acts as a potential prognostic factor and therapeutic target in colon cancer.

  4. Kaempferol, a phytoestrogen, suppressed triclosan-induced epithelial-mesenchymal transition and metastatic-related behaviors of MCF-7 breast cancer cells.

    Science.gov (United States)

    Lee, Geum-A; Choi, Kyung-Chul; Hwang, Kyung-A

    2017-01-01

    As a phytoestrogen, kaempferol is known to play a chemopreventive role inhibiting carcinogenesis and cancer progression. In this study, the influences of triclosan, an anti-bacterial agent recently known for an endocrine disrupting chemical (EDC), and kaempferol on breast cancer progression were examined by measuring their effects on epithelial-mesenchymal transition (EMT) and metastatic-related behaviors of MCF-7 breast cancer cells. Morphological changes of MCF-7 cells were observed, and a wound-healing assay was performed after the treatment of triclosan and kaempferol. The effects of triclosan and kaempferol on protein expression of EMT-related markers such as E-cadherin, N-cadherin, Snail, and Slug and metastasis-related markers such as cathepsin B, D, MMP-2 and -9 were investigated by Western blot assay. In microscopic observations, triclosan (10(-6)M) or E2 (10(-9)M) induced transition to mesenchymal phenotype of MCF-7 cells compared with the control. Co-treatment of ICI 182,780 (10(-8)M), an ER antagonist, or kaempferol (25μM) with E2 or triclosan restored the cellular morphology to an epithelial phenotype. In a wound-healing scratch and a transwell migration assay, triclosan enhanced migration and invasion of MCF-7 cells, but co-treatment of kaempferol or ICI 182,780 reduced the migration and invasion ability of MCF-7 cells to the control level. In addition, kaempferol effectively suppressed E2 or triclosan-induced protein expressions of EMT and metastasis promoting markers. Taken together, triclosan may be a distinct xenoestrogenic EDC to promote EMT, migration, and invasion of MCF-7 breast cancer cells through ER. On the other hand, kaempferol can be an alternative chemopreventive agent to effectively suppress the metastatic behavior of breast cancer induced by an endogenous estrogen as well as exogenous xenoestrogenic compounds including triclosan.

  5. Down-regulation of Transducin-Like Enhancer of Split protein 4 in hepatocellular carcinoma promotes cell proliferation and epithelial-Mesenchymal-Transition

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    Wu, Xiao-cai; Xiao, Cui-cui; Li, Hua [Department of Hepatic Surgery, 3rd Affiliated Hospital of Sun Yat-sen University, Guangzhou (China); Guangdong Provincial Key Laboratory of Liver Disease Research, Guangzhou (China); Tai, Yan; Zhang, Qi [Guangdong Provincial Key Laboratory of Liver Disease Research, Guangzhou (China); Yang, Yang, E-mail: yysysu2@163.com [Department of Hepatic Surgery, 3rd Affiliated Hospital of Sun Yat-sen University, Guangzhou (China)

    2016-08-19

    Background: Transducin-Like Enhancer of Split protein 4 (TLE4) has been reported to be involved in some subsets of acute myeloid leukemia and colorectal cancer. In the present study, we aimed to explore the role of TLE4 in tumorigenesis and cancer progression in hepatocellular carcinoma (HCC). Methods: The expression pattern of TLE4 in HCC was determined by Western-blot and qRT-PCR, gain-of-function and loss-of-function was used to explore the biological role of TLE4 in HCC cells. A xenograft model was established to confirm its effects on proliferation. Results: The protein expression levels of TLE4 were significantly down-regulated in HCC tissues compared to matched adjacent normal liver tissues. In vitro, down-regulation of TLE4 in Huh7 or SMMC-7721 promoted cell proliferation and ectopical expression of TLE4 in Hep3B or Bel-7404 suppressed cell proliferation. In addition, the cell colony formation ability was enhanced after down-regulation of TLE4 expression in Huh-7 but suppressed after over-expression in Hep3B. Furthermore, down-regulation of TLE4 increased the cell invasion ability, as well as increased the expression level of Vimentin and decreased that of E-cadherin, indicating a phenotype of epithelial-mesenchymal transition (EMT) in HCC cells. On the contrary, ectopical expression of TLE4 in HCC cells decreased the cell invasion ability and inhibited EMT. In vivo, compared to control group, xenograft tumor volumes were significantly decreased in TLE4 overexpression group. Conclusions: These results demonstrated that TLE4 might play important regulatory roles in cellular proliferation and EMT process in HCC. - Highlights: • TLE4 is significantly down-regulated in HCC samples. • Down regulated of TLE4 in HCC cells promotes cell proliferation. • Down regulated of TLE4 in HCC cells promotes epithelial-to-mesenchymal transition.

  6. CCAAT/enhancer binding protein beta (C/EBPβ) isoform balance as a regulator of epithelial-mesenchymal transition in mouse mammary epithelial cells

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    Miura, Yuka; Hagiwara, Natsumi [Department of Bioscience, Graduate School of Science and Technology, Kwansei Gakuin University, Hyogo, 2-1 Gakuen, Sanda 669-1337 Japan (Japan); Radisky, Derek C. [Department of Cancer Biology, Mayo Clinic, Jacksonville, FL 32225 (United States); Hirai, Yohei, E-mail: y-hirai@kwansei.ac.jp [Department of Bioscience, Graduate School of Science and Technology, Kwansei Gakuin University, Hyogo, 2-1 Gakuen, Sanda 669-1337 Japan (Japan)

    2014-09-10

    Activation of the epithelial-mesenchymal transition (EMT) program promotes cell invasion and metastasis, and is reversed through mesenchymal-epithelial transition (MET) after formation of distant metastases. Here, we show that an imbalance of gene products encoded by the transcriptional factor C/EBPβ, LAP (liver-enriched activating protein) and LIP (liver-enriched inhibitory protein), can regulate both EMT- and MET-like phenotypic changes in mouse mammary epithelial cells. By using tetracycline repressive LIP expression constructs, we found that SCp2 cells, a clonal epithelial line of COMMA1-D cells, expressed EMT markers, lost the ability to undergo alveolar-like morphogenesis in 3D Matrigel, and acquired properties of benign adenoma cells. Conversely, we found that inducible expression of LAP in SCg6 cells, a clonal fibroblastic line of COMMA1-D cells, began to express epithelial keratins with suppression of proliferation. The overexpression of the C/EBPβ gene products in these COMMA1-D derivatives was suppressed by long-term cultivation on tissue culture plastic, but gene expression was maintained in cells grown on Matrigel or exposed to proteasome inhibitors. Thus, imbalances of C/EBPβ gene products in mouse mammary epithelial cells, which are affected by contact with basement membrane, are defined as a potential regulator of metastatic potential. - Highlights: • We created a temporal imbalance of C/EBPβ gene products in the mammary model cells. • The temporal up-regulation of LIP protein induced EMT-like cell behaviors. • The temporal up-regulation of LAP protein induced MET-like cell behaviors. • Excess amount of C/EBPβ gene products were eliminated by proteasomal-degradation. • Basement membrane components attenuated proteasome-triggered protein elimination.

  7. Over-expression of miR-106b promotes cell migration and metastasis in hepatocellular carcinoma by activating epithelial-mesenchymal transition process.

    Directory of Open Access Journals (Sweden)

    Wing Lung Yau

    Full Text Available Hepatocellular carcinoma (HCC is one the the most fatal cancers worldwide. The poor prognosis of HCC is mainly due to the developement of distance metastasis. To investigate the mechanism of metastasis in HCC, an orthotopic HCC metastasis animal model was established. Two sets of primary liver tumor cell lines and corresponding lung metastasis cell lines were generated. In vitro functional analysis demonstrated that the metastatic cell line had higher invasion and migration ability when compared with the primary liver tumor cell line. These cell lines were subjected to microRNA (miRNAs microarray analysis to identify differentially expressed miRNAs which were associated with the developement of metastasis in vivo. Fifteen human miRNAs, including miR-106b, were differentially expressed in 2 metastatic cell lines compared with the primary tumor cell lines. The clinical significance of miR-106b in 99 HCC clinical samples was studied. The results demonstrated that miR-106b was over-expressed in HCC tumor tissue compared with adjacent non-tumor tissue (p = 0.0005, and overexpression of miR-106b was signficantly correlated with higher tumor grade (p = 0.018. Further functional studies demonstrated that miR-106b could promote cell migration and stress fiber formation by over-expressing RhoGTPases, RhoA and RhoC. In vivo functional studies also showed that over-expression of miR-106b promoted HCC metastasis. These effects were related to the activation of the epithelial-mesenchymal transition (EMT process. Our results suggested that miR-106b expression contributed to HCC metastasis by activating the EMT process promoting cell migration in vitro and metastasis in vivo.

  8. Sanguiin H6 suppresses TGF-β induction of the epithelial-mesenchymal transition and inhibits migration and invasion in A549 lung cancer.

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    Ko, Hyeonseok; Jeon, Hyelin; Lee, Dahae; Choi, Hyo-Kyoung; Kang, Ki Sung; Choi, Kyung-Chul

    2015-12-01

    In the epithelial-mesenchymal transition (EMT), an important cellular process, epithelial cells become mesenchymal cells. This process is also critically involved in cancer metastasis. Sanguiin H6 is a compound derived from ellagitannin, which is found in berries. Sanguiin H6 shows various pharmacological properties, including anti-angiogenic activity. Because the possible role of sanguiin H6 in the EMT and the underlying molecular mechanisms are unclear, we investigated the effect of sanguiin H6 on the EMT. Transforming growth factor-beta 1 (TGF-β1) induces the EMT and promotes lung adenocarcinoma migration and invasion through the Smad2/3 signaling pathway. Thus, to understand the inhibitory effects of sanguiin H6 on lung cancer migration and invasion, we investigated the ability of sanguiin H6 to inhibit TGF-β1-induced EMT in the A549 cell line. We found that sanguiin H6 significantly prevented the activation of Smad2/3 signaling pathway by TGF-β1. Additionally, sanguiin H6 increased the expression of the epithelial marker E-cadherin and repressed the expression of Snail and the mesenchymal marker N-cadherin during TGF-β1-induced EMT. Moreover, sanguiin H6 regulated the expression of EMT-dependent genes induced by TGF-β1. Finally, sanguiin H6 inhibited the migration and invasion of TGF-β1-stimulated A549 cells. Taken together, our findings provide new evidence that sanguiin H6 suppresses lung cancer migration and invasion in vitro by inhibiting TGF-β1 induction of the EMT.

  9. Atrial natriuretic peptide: A novel mediator for TGF-β1-induced epithelial-mesenchymal transition in 16HBE-14o and A549 cells.

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    Chu, Shuyuan; Zhang, Xiufeng; Sun, Yabing; Yu, Yuanyuan; Liang, Yaxi; Jiang, Ming; Huang, Jianwei; Ma, Libing

    2017-02-13

    Atrial natriuretic peptide (ANP) is increasingly expressed on airway and inhibits pulmonary arterial remodeling. However, the role of ANP in remodeling of respiratory system is still unclear. The role of ANP on airway remodeling and the possible mechanism was explored in this study. Both human bronchial epithelial 16HBE-14o cells and alveolar epithelial A549 cells were stimulated by TGF-β1, ANP, cGMP inhibitor, PKG inhibitor, and cGMP analogue. The expressions of epithelial markers, mesenchymal markers, and Smad3 were assessed by quantitative real-time PCR and western blotting. Immunohistochemical staining was employed to assess Smad3 expression once it was silenced by siRNA in 16HBE-14o or A549 cells. Our results showed that the mRNA and protein expressions of E-Cadherin were decreased, whereas α-SMA expressions were increased after induction by TGF-β1 in 16HBE-14o and A549 cells. The E-Cadherin expressions were increased and α-SMA expressions were decreased after ANP stimulation. Inhibition of cGMP or PKG decreased E-Cadherin expression but increased α-SMA expression, which could be reversed by cGMP analogue. Moreover, the phosphorylated Smad3 expression was consistent with α-SMA expression. After smad3 was silenced, Smad3 was mostly expressed in cytoplasm instead of nucleus as non-silenced cells during epithelial-mesenchymal transition (EMT). In conclusion, ANP inhibits TGF-β1-induced EMT in 16HBE-14o and A549 cells through cGMP/PKG signaling, by which it targets TGF-β1/Smad3 via attenuating phosphorylation of Smad3. These findings suggest the potential of ANP in the treatment on pulmonary diseases with airway remodeling.

  10. Paeonol attenuates aging MRC-5 cells and inhibits epithelial-mesenchymal transition of premalignant HaCaT cells induced by aging MRC-5 cell-conditioned medium.

    Science.gov (United States)

    Yang, Lihua; Xing, Shangping; Wang, Kun; Yi, Hua; Du, Biaoyan

    2017-08-12

    Senescence-associated secretory phenotype (SASP) factors, such as IL-6 and IL-8, are extremely critical in tissue microenvironment. Senescent human fibroblasts facilitate epithelial-mesenchymal transition (EMT) in premalignant epithelial cells mainly through the secretion of SASP factors. Meanwhile, premalignant human HaCaT Keratinocyte (HaCaT) cells as immortal epithelial cells are susceptible to malignant transformation. Paeonol, an herbal phenolic component found in peonies, exerts anti-aging and anti-tumor efficacies, while the molecular mechanisms of paeonol on EMT in premalignant HaCaT cells induced by SASP factors are unclear. In this study, we first established a senescent human fetal lung fibroblast MRC-5 cell model using hydrogen peroxide evaluated by senescence-associated β-galactosidase assay. Upon paeonol treatment, intracellular reactive oxygen species levels in aging MRC-5 cells were significantly decreased via regulation of nuclear translocation of Nrf2. Then we curiously studied whether the aging MRC-5 cell-conditioned medium could induce EMT in premalignant HaCaT cells, and the results showed that paeonol significantly reduced the clonogenic, migratory, and invasive capacities of premalignant HaCaT cells potentially induced by IL-6 and IL-8. Moreover, we found that paeonol notably altered pluripotency of EMT-associated markers via the modulation of ERK and TGF-β1/Smad pathway in premalignant HaCaT cells. These findings suggest that paeonol may be used as an adjuvant therapy for SASP factor-mediated EMT in premalignant lesion.

  11. Role of nuclear factor kappa B and reactive oxygen species in the tumor necrosis factor-a-induced epithelial-mesenchymal transition of MCF-7 cells

    Directory of Open Access Journals (Sweden)

    R. Dong

    2007-08-01

    Full Text Available The microenvironment of the tumor plays an important role in facilitating cancer progression and activating dormant cancer cells. Most tumors are infiltrated with inflammatory cells which secrete cytokines such as tumor necrosis factor-a (TNF-a. To evaluate the role of TNF-a in the development of cancer we studied its effects on cell migration with a migration assay. The migrating cell number in TNF-a -treated group is about 2-fold of that of the control group. Accordingly, the expression of E-cadherin was decreased and the expression of vimentin was increased upon TNF-a treatment. These results showed that TNF-a can promote epithelial-mesenchymal transition (EMT of MCF-7 cells. Further, we found that the expression of Snail, an important transcription factor in EMT, was increased in this process, which is inhibited by the nuclear factor kappa B (NFkB inhibitor aspirin while not affected by the reactive oxygen species (ROS scavenger N-acetyl cysteine. Consistently, specific inhibition of NFkB by the mutant IkBa also blocked the TNF-a-induced upregulation of Snail promoter activity. Thus, the activation of NFkB, which causes an increase in the expression of the transcription factor Snail is essential in the TNF-a-induced EMT. ROS caused by TNF-a seemed to play a minor role in the TNF-a-induced EMT of MCF-7 cells, though ROS per se can promote EMT. These findings suggest that different mechanisms might be responsible for TNF-a - and ROS-induced EMT, indicating the need for different strategies for the prevention of tumor metastasis induced by different stimuli.

  12. Valproic acid inhibits irradiation-induced epithelial-mesenchymal transition and stem cell-like characteristics in esophageal squamous cell carcinoma

    Science.gov (United States)

    Kanamoto, Ayako; Ninomiya, Itasu; Harada, Shinichi; Tsukada, Tomoya; Okamoto, Koichi; Nakanuma, Shinichi; Sakai, Seisho; Makino, Isamu; Kinoshita, Jun; Hayashi, Hironori; Oyama, Katsunobu; Miyashita, Tomoharu; Tajima, Hidehiro; Takamura, Hiroyuki; Fushida, Sachio; Ohta, Tetsuo

    2016-01-01

    Esophageal carcinoma is one of the most aggressive malignancies, and is characterized by poor response to current therapy and a dismal survival rate. In this study we investigated whether irradiation induces epithelial-mesenchymal transition (EMT) in esophageal squamous cell carcinoma (ESCC) TE9 cells and whether the classic histone deacetylase (HDAC) inhibitor valproic acid (VPA) suppresses these changes. First, we showed that 2 Gy irradiation induced spindle cell-like morphologic changes, decreased expression of membranous E-cadherin, upregulated vimentin expression, and altered the localization of β-catenin from its usual membrane-bound location to cytoplasm in TE9 cells. Irradiation induced upregulation of transcription factors including Slug, Snail, and Twist, which regulate EMT. Stimulation by irradiation resulted in increased TGF-β1 and HIF-1α expression and induced Smad2 and Smad3 phosphorylation. Furthermore, irradiation enhanced CD44 expression, indicating acquisition of cancer stem-like cell properties. In addition, irradiation enhanced invasion and migration ability with upregulation of matrix metalloproteinases. These findings indicate that single-dose irradiation can induce EMT in ESCC cells. Second, we found that treatment with 1 mM VPA induced reversal of EMT caused by irradiation in TE9 cells, resulting in attenuated cell invasion and migration abilities. These results suggest that VPA might have clinical value to suppress irradiation-induced EMT. The reversal of EMT by HDAC inhibitors may be a new therapeutic strategy to improve the effectiveness of radiotherapy in ESCC by inhibiting the enhancement of invasion and metastasis.

  13. Stromal Fibroblasts from the Interface Zone of Triple Negative Breast Carcinomas Induced Epithelial-Mesenchymal Transition and its Inhibition by Emodin

    Science.gov (United States)

    Wang, Hao-Yu; Hung, Chao-Ming; Lin, Ying-Chao; Ho, Chi-Tang; Way, Tzong-Der

    2017-01-01

    “Triple negative breast cancer” (TNBC) is associated with a higher rate and earlier time of recurrence and worse prognosis after recurrence. In this study, we aimed to examine the crosstalk between fibroblasts and TNBC cells. The fibroblasts were isolated from TNBC patients’ tissue in tumor burden zones, distal normal zones and interface zones. The fibroblasts were indicated as cancer-associated fibroblasts (CAFs), normal zone fibroblasts (NFs) and interface zone fibroblasts (INFs). Our study found that INFs grew significantly faster than NFs and CAFs in vitro. The epithelial BT20 cells cultured with the conditioned medium of INFs (INFs-CM) and CAFs (CAFs-CM) showed more spindle-like shape and cell scattering than cultured with the conditioned medium of NFs (NFs-CM). These results indicated that factors secreted by INFs-CM or CAFs-CM could induce the epithelial-mesenchymal transition (EMT) phenotype in BT20 cells. Using an in vitro co-culture model, INFs or CAFs induced EMT and promoted cancer cell migration in BT20 cells. Interestingly, we found that emodin inhibited INFs-CM or CAFs-CM-induced EMT programming and phenotype in BT20 cells. Previous studies reported that CAFs and INFs-secreted TGF-β promoted human breast cancer cell proliferation, here; our results indicated that TGF-β initiated EMT in BT20 cells. Pretreatment with emodin significantly suppressed the TGF-β-induced EMT and cell migration in BT20 cells. These results suggest that emodin may be used as a novel agent for the treatment of TNBC. PMID:28060811

  14. Tumor cell heterogeneity in Small Cell Lung Cancer (SCLC: phenotypical and functional differences associated with Epithelial-Mesenchymal Transition (EMT and DNA methylation changes.

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    Alexander Krohn

    Full Text Available Small Cell Lung Cancer (SCLC is a specific subtype of lung cancer presenting as highly metastatic disease with extremely poor prognosis. Despite responding initially well to chemo- or radiotherapy, SCLC almost invariably relapses and develops resistance to chemotherapy. This is suspected to be related to tumor cell subpopulations with different characteristics resembling stem cells. Epithelial-Mesenchymal Transition (EMT is known to play a key role in metastatic processes and in developing drug resistance. This is also true for NSCLC, but there is very little information on EMT processes in SCLC so far. SCLC, in contrast to NSCLC cell lines, grow mainly in floating cell clusters and a minor part as adherent cells. We compared these morphologically different subpopulations of SCLC cell lines for EMT and epigenetic features, detecting significant differences in the adherent subpopulations with high levels of mesenchymal markers such as Vimentin and Fibronectin and very low levels of epithelial markers like E-cadherin and Zona Occludens 1. In addition, expression of EMT-related transcription factors such as Snail/Snai1, Slug/Snai2, and Zeb1, DNA methylation patterns of the EMT hallmark genes, functional responses like migration, invasion, matrix metalloproteases secretion, and resistance to chemotherapeutic drug treatment all differed significantly between the sublines. This phenotypic variability might reflect tumor cell heterogeneity and EMT during metastasis in vivo, accompanied by the development of refractory disease in relapse. We propose that epigenetic regulation plays a key role during phenotypical and functional changes in tumor cells and might therefore provide new treatment options for SCLC patients.

  15. Cigarette smoke extracts induced the colon cancer migration via regulating epithelial mesenchymal transition and metastatic genes in human colon cancer cells.

    Science.gov (United States)

    Kim, Cho-Won; Go, Ryeo-Eun; Lee, Hae-Miru; Hwang, Kyung-A; Lee, Kyuhong; Kim, Bumseok; Lee, Moo-Yeol; Choi, Kyung-Chul

    2017-02-01

    There was considerable evidence that exposure to cigarette smoke is associated with an increased risk for colon cancer. Nevertheless, the mechanism underlying the relationship between cigarette smoking and colon cancer remains unclear. Moreover, there were only a few studies on effects of complexing substance contained in cigarette smoke on colon cancer. Thus, we further investigated whether cigarette smoke extract (CSE) affects the cell cycle, apoptosis and migration of human metastatic colon cancer cells, SW-620. MTT assay revealed that SW-620 cell proliferation was significantly inhibited following treatments with all CSEs, 3R4F, and two-domestic cigarettes, for 9 days in a concentration-dependent manner. Moreover, CSE treatments decreased cyclin D1 and E1, and increased p21 and p27 proteins by Western blot analysis in SW-620 cells. Additionally, the treatment of the cells with CSE contributed to these effects expressing by apoptosis-related proteins. An increased migration or invasion ability of SW-620 cells following CSE treatment was also confirmed by a scratch or fibronectin invasion assay in vitro. In addition, the protein levels of E-cadherin as an epithelial maker were down-regulated, while the mesenchymal markers, N-cadherin, snail, and slug, were up-regulated in a time-dependent manner. A metastatic marker, cathepsin D, was also down-regulated by CSE treatment. Taken together, these results indicate that CSE exposure in colon cancer cells may deregulate the cell growth by altering the expression of cell cycle-related proteins and pro-apoptotic protein, and stimulate cell metastatic ability by altering epithelial-mesenchymal transition (EMT) markers and cathepsin D expression. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 690-704, 2017.

  16. miR156a Mimic Represses the Epithelial-Mesenchymal Transition of Human Nasopharyngeal Cancer Cells by Targeting Junctional Adhesion Molecule A.

    Directory of Open Access Journals (Sweden)

    Yunhong Tian

    Full Text Available MicroRNAs (miRNAs have been documented as having an important role in the development of cancer. Broccoli is very popular in large groups of the population and has anticancer properties. Junctional adhesion molecule A (JAMA is preferentially concentrated at tight junctions and influences cell morphology and migration. Epithelial-mesenchymal transition (EMT is a developmental program associated with cancer progression and metastasis. In this study we aimed to investigate the role of miRNAs from broccoli in human nasopharyngeal cancer (NPC. We demonstrated that a total of 84 conserved miRNAs and 184 putative novel miRNAs were found in broccoli by sequencing technology. Among these, miR156a was expressed the most. In addition, synthetic miR156a mimic inhibited the EMT of NPC cells in vitro. Furthermore, it was confirmed that JAMA was the target of miR156a mimic as validated by 3' UTR luciferase reporter assays and western blotting. Knockdown of JAMA was consistent with the effects of miR156a mimic on the EMT of NPC, and the up-regulation of JAMA could partially restore EMT repressed by miR156a mimic. In conclusion, these results indicate that the miR156a mimic inhibits the EMT of NPC cells by targeting the 3' UTR of JAMA. These miRNA profiles of broccoli provide a fundamental basis for further research. Moreover, the discovery of miR156a may have clinical implications for the treatment of patients with NPC.

  17. Ultrasound-targeted microbubble destruction-mediated downregulation of CD133 inhibits epithelial-mesenchymal transition, stemness and migratory ability of liver cancer stem cells.

    Science.gov (United States)

    Liu, Yan-Min; Li, Xuan-Fei; Liu, Hao; Wu, Xiao-Ling

    2015-12-01

    Hepatocellular carcinoma (HCC) is an aggressive disease with a poor outcome due to the high incidence of metastasis. Cancer stem cells (CSCs) have been identified to be responsible for tumor progression and may be generated by epithelial-mesenchymal transition (EMT) characteristics. CD133 is a specific surface marker for liver cancer stem cells (LCSCs), which is also considered as an important functional factor for tumorigenesis and overall survival in HCC. Ultrasound-targeted microbubble destruction (UTMD) has recently been used as a novel, safe and effective gene transfection technology. The aim of the present study was to elucidate the regulatory mechanism of CD133 and EMT in LCSCs and whether the UTMD-based shRNA delivery system facilitated gene delivery in LCSCs. In the present study, CD133+ cells were isolated from the SMMC-7721 HCC cell line and then transfected with shCD133 mediated by UTMD and liposomes, respectively. Compared to the liposomes group, the UTMD group resulted in significantly improved transfection efficiency. The downregulation of CD133 reversed the EMT program, attenuated self-renewal, proliferation and migration of CD133+ LCSCs and suppressed the growth of CSC tumor xenografts. Additionally, the downregulation of CD133 led to downregulation of the nuclear factor-κB (NF-κB) pathway. The present study demonstrated that CD133 plays a critical role in the regulation of the EMT process, tumor-initiating properties and migratory ability of LCSCs. The UTMD technique targeted for CD133 downregulation may be examined as a potential therapeutic strategy for HCC.

  18. Blockade of Jagged/Notch pathway abrogates transforming growth factor β2-induced epithelial-mesenchymal transition in human retinal pigment epithelium cells.

    Science.gov (United States)

    Chen, X; Xiao, W; Liu, X; Zeng, M; Luo, L; Wu, M; Ye, S; Liu, Y

    2014-05-01

    The epithelial-mesenchymal transition (EMT) of retinal pigment epithelium (RPE) cells plays a key role in proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR), which lead to the loss of vision. The Jagged/Notch pathway has been reported to be essential in EMT during embryonic development, fibrotic diseases and cancer metastasis. However, the function of Jagged/Notch signaling in EMT of RPE cells is unknown. Thus, we hypothesized that a crosstalk between Notch and transforming growth factor β2 (TGF-β2) signaling could induce EMT in RPE cells, which subsequently contributes to PVR and PDR. Here, we demonstrate that Jagged-1/Notch pathway is involved in the TGF-β2-mediated EMT of human RPE cells. Blockade of Notch pathway with DAPT (a specific inhibitor of Notch receptor cleavage) and knockdown of Jagged-1 expression inhibited TGF-β2-induced EMT through regulating the expression of Snail, Slug and ZEB1. Besides the canonical Smad signaling pathway, the noncanonical PI3K/Akt and MAPK pathway also contributed to TGF-β2-induced up-regulation of Jagged-1 in RPE cells. Overexpression of Jagged-1 could mimic TGF-β2 induce EMT. Our data suggest that the Jagged-1/Notch signaling pathway plays a critical role in TGF-β2-induced EMT in human RPE cells, and may contribute to the development of PVR and PDR. Inhibition of the Jagged/Notch signaling pathway, therefore, may have therapeutic value in the prevention and treatment of PVR and PDR.

  19. SIRT1 induces tumor invasion by targeting epithelial mesenchymal transition-related pathway and is a prognostic marker in triple negative breast cancer.

    Science.gov (United States)

    Jin, Min-Sun; Hyun, Chang Lim; Park, In Ae; Kim, Ji Young; Chung, Yul Ri; Im, Seock-Ah; Lee, Kyung-Hun; Moon, Hyeong-Gon; Ryu, Han Suk

    2016-04-01

    Absence of therapeutic targets poses a critical hurdle in improving prognosis for patients with triple negative breast cancer (TNBC). We evaluated interaction between SIRT1 and epithelial mesenchymal transition (EMT)-associated proteins as well as the role of combined protein expression as a predictor of lymph node metastasis and clinical outcome in TNBC through in vivo and vitro studies. Three hundred nineteen patients diagnosed with TNBC were chosen, immunohistochemical staining for SIRT1 and EMT-related markers' expression was performed on tissue microarrays, and in vitro experiments with each of the three human TNBC cell lines were carried out. The cohort was reclassified according to the use of adjuvant chemotherapy, tumor size, and AJCC stage to analyze the prognostic role of SIRT1 and EMT-related proteins' expression considering different therapeutic modalities and AJCC stages. Combination of four proteins including SIRT1 and three EMT-related proteins was revealed to be a statistically significant independent predictor of lymph node metastasis in the tumor size cohort as well as in the total patient population. Upon Cox regression analysis, increased expression level of the combined proteins correlated with decreased disease-free survival in the total patients as well as those who received adjuvant chemotherapy and those who had early stage breast cancer. In additional in vitro experiments, inhibition of SIRT1 expression with small interfering RNA (siRNA) suppressed tumor invasion in three different TNBC cell lines, and altered expression levels of EMT-related proteins following SIRT1 gene inhibition were identified on western blotting and fluorescence activated cell sorting (FACS) analysis; on the other hand, no change in expression levels of the cell cycle-related factors was observed. Our analysis showed the potential role of SIRT1 in association with EMT-related factors on tumor invasion, metastasis, and disease-free survival in TNBC, SIRT1, and

  20. Loss of TAK1 increases cell traction force in a ROS-dependent manner to drive epithelial-mesenchymal transition of cancer cells.

    Science.gov (United States)

    Lam, C R I; Tan, C; Teo, Z; Tay, C Y; Phua, T; Wu, Y L; Cai, P Q; Tan, L P; Chen, X; Zhu, P; Tan, N S

    2013-10-10

    Epithelial-mesenchymal transition (EMT) is a crucial step in tumor progression, and the TGFβ-SMAD signaling pathway as an inductor of EMT in many tumor types is well recognized. However, the role of non-canonical TGFβ-TAK1 signaling in EMT remains unclear. Herein, we show that TAK1 deficiency drives metastatic skin squamous cell carcinoma earlier into EMT that is conditional on the elevated cellular ROS level. The expression of TAK1 is consistently reduced in invasive squamous cell carcinoma biopsies. Tumors derived from TAK1-deficient cells also exhibited pronounced invasive morphology. TAK1-deficient cancer cells adopt a more mesenchymal morphology characterized by higher number of focal adhesions, increase surface expression of integrin α5β1 and active Rac1. Notably, these mutant cells exert an increased cell traction force, an early cellular response during TGFβ1-induced EMT. The mRNA level of ZEB1 and SNAIL, transcription factors associated with mesenchymal phenotype is also upregulated in TAK1-deficient cancer cells compared with control cancer cells. We further show that TAK1 modulates Rac1 and RhoA GTPases activities via a redox-dependent downregulation of RhoA by Rac1, which involves the oxidative modification of low-molecular weight protein tyrosine phosphatase. Importantly, the treatment of TAK1-deficient cancer cells with Y27632, a selective inhibitor of Rho-associated protein kinase and antioxidant N-acetylcysteine augment and hinders EMT, respectively. Our findings suggest that a dysregulated balance in the activation of TGFβ-TAK1 and TGFβ-SMAD pathways is pivotal for TGFβ1-induced EMT. Thus, TAK1 deficiency in metastatic cancer cells increases integrin:Rac-induced ROS, which negatively regulated Rho by LMW-PTP to accelerate EMT.

  1. Apobec-1 Complementation Factor (A1CF Inhibits Epithelial-Mesenchymal Transition and Migration of Normal Rat Kidney Proximal Tubular Epithelial Cells

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    Liyuan Huang

    2016-02-01

    Full Text Available Apobec-1 complementation factor (A1CF is a member of the heterogeneous nuclear ribonucleoproteins (hnRNP family, which participates in site-specific posttranscriptional RNA editing of apolipoprotein B (apoB transcript. The posttranscriptional editing of apoB mRNA by A1CF in the small intestine is required for lipid absorption. Apart from the intestine, A1CF mRNA is also reported to be highly expressed in the kidneys. However, it is remained unknown about the functions of A1CF in the kidneys. The aim of this paper is to explore the potential functions of A1CF in the kidneys. Our results demonstrated that in C57BL/6 mice A1CF was weakly expressed in embryonic kidneys from E15.5dpc while strongly expressed in mature kidneys after birth, and it mainly existed in the tubules of inner cortex. More importantly, we identified A1CF negatively regulated the process of epithelial-mesenchymal transition (EMT in kidney tubular epithelial cells. Our results found ectopic expression of A1CF up-regulated the epithelial markers E-cadherin, and down-regulated the mesenchymal markers vimentin and α-smooth muscle actin (α-SMA in NRK52e cells. In addition, knockdown of A1CF enhanced EMT contrary to the overexpression effect. Notably, the two A1CF variants led to the similar trend in the EMT process. Taken together, these data suggest that A1CF may be an antagonistic factor to the EMT process of kidney tubular epithelial cells.

  2. The expression of syndecan-1 and -2 is associated with Gleason score and epithelial-mesenchymal transition markers, E-cadherin and beta-catenin, in prostate cancer.

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    Contreras, Hector R; Ledezma, Rodrigo A; Vergara, Jorge; Cifuentes, Federico; Barra, Cristina; Cabello, Pablo; Gallegos, Ivan; Morales, Bernardo; Huidobro, Christian; Castellón, Enrique A

    2010-01-01

    The epithelial-mesenchymal transition (EMT) is considered a key step in tumor progression, where the invasive cancer cells change from epithelial to mesenchymal phenotype. During this process, a decrease or loss in adhesion molecules expression and an increase in migration molecules expression are observed. The aim of this work was to determine the expression and cellular distribution of syndecan-1 and -2 (migration molecules) and E-cadherin and beta-catenin (adhesion molecules) in different stages of prostate cancer progression. A quantitative immunohistochemical study of these molecules was carried out in tissue samples from benign prostatic hyperplasia and prostate carcinoma, with low and high Gleason score, obtained from biopsies archives of the Clinic Hospital of the University of Chile and Dipreca Hospital. Polyclonal specific antibodies and amplification system of estreptavidin-biotin peroxidase and diaminobenzidine were used. Syndecan-1 was uniformly expressed in basolateral membranes of normal epithelium, changing to a granular cytoplasmatic expression pattern in carcinomas. Syndecan-2 was observed mainly in a cytoplasmatic granular pattern, with high immunostaining intensity in areas of low Gleason score. E-cadherin was detected in basolateral membrane of normal epithelia showing decreased expression in high Gleason score samples. beta-Catenin was found in cell membranes of normal epithelia changing its distribution toward the nucleus and cytoplasm in carcinoma samples. We concluded that changes in expression and cell distribution of E-cadherin and beta-catenin correlated with the progression degree of prostate adenocarcinoma, suggesting a role of these molecules as markers of progression and prognosis. Furthermore, changes in the pattern expression of syndecan-1 and -2 indicate that both molecules may be involved in the EMT and tumor progression of prostate cancer.

  3. Anti-metastatic potential of resveratrol and its metabolites by the inhibition of epithelial-mesenchymal transition, migration, and invasion of malignant cancer cells.

    Science.gov (United States)

    Kim, Cho-Won; Hwang, Kyung-A; Choi, Kyung-Chul

    2016-12-15

    Increased epithelial-mesenchymal transition (EMT) and cell migration and invasion abilities of cancer cells play important roles in the metastatic process of cancer. Resveratrol is a stilbenoid, a type of natural polyphenol found in the skin of grapes, berries, and peanuts. A number of experiments have examined resveratrol's ability to target diverse pathways associated with carcinogenesis and cancer progression. This article aims to present updated overview of the knowledge that resveratrol and its metabolites or analogs have the potential to inhibit metastasis of cancer via affecting many signaling pathways related with EMT, cancer migration, and invasion in diverse organs of the body. This article starts with a short introduction describing diverse beneficial effects of resveratrol including cancer prevention and the aim of the present study. To address the effects of resveratrol on cancer metastasis, mechanisms of EMT, migration, invasion, and their relevance with cancer metastasis, anti-metastatic effects of resveratrol through EMT-related signaling pathways and inhibitory effects of resveratrol on migration and invasion are highlighted. In addition, anti-metastatic potential of resveratrol metabolites and analogs is addressed. Resveratrol was demonstrated to turn back the EMT process induced by diverse signaling pathways in several cellular and animal cancer models. In addition, resveratrol can exert chemopreventive efficacies on migration and invasion of cancer cells by inhibiting the related pathways and target molecules. Although these findings display the anti-metastatic potential of resveratrol, more patient-oriented clinical studies demonstrating the marked efficacies of resveratrol in humans are still needed. Copyright © 2016 Elsevier GmbH. All rights reserved.

  4. Evidence for embryonic stem-like signature and epithelial-mesenchymal transition features in the spheroid cells derived from lung adenocarcinoma.

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    Roudi, Raheleh; Madjd, Zahra; Ebrahimi, Marzieh; Najafi, Ali; Korourian, Alireza; Shariftabrizi, Ahmad; Samadikuchaksaraei, Ali

    2016-09-01

    Identification of the cellular and molecular aspects of lung cancer stem cells (LCSCs) that are suggested to be the main culprit of tumor initiation, maintenance, drug resistance, and relapse is a prerequisite for targeted therapy of lung cancer. In the current study, LCSCs subpopulation of A549 cells was enriched, and after characterization of the spheroid cells, complementary DNA (cDNA) microarray analysis was applied to identify differentially expressed genes (DEGs) between the spheroid and parental cells. Microarray results were validated using quantitative real-time reverse transcription-PCR (qRT-PCR), flow cytometry, and western blotting. Our results showed that spheroid cells had higher clonogenic potential, up-regulation of stemness gene Sox2, loss of CD44 expression, and gain of CD24 expression compared to parental cells. Among a total of 160 genes that were differentially expressed between the spheroid cells and the parental cells, 104 genes were up-regulated and 56 genes were down-regulated. Analysis of cDNA microarray revealed an embryonic stem cell-like signature and over-expression of epithelial-mesenchymal transition (EMT)-associated genes in the spheroid cells. cDNA microarray results were validated at the gene expression level using qRT-PCR, and further validation was performed at the protein level by flow cytometry and western blotting. The embryonic stem cell-like signature in the spheroid cells supports two important notions: maintenance of CSCs phenotype by dedifferentiating mechanisms activated through oncogenic pathways and the origination of CSCs from embryonic stem cells (ESCs). PI3/AKT3, as the most common up-regulated pathway, and other pathways related to aggressive tumor behavior and EMT process can confer to the spheroid cells' high potential for metastasis and distant seeding.

  5. Epithelial-mesenchymal transition (EMT) induced by TNF-α requires AKT/GSK-3β-mediated stabilization of snail in colorectal cancer.

    Science.gov (United States)

    Wang, Hao; Wang, Hong-Sheng; Zhou, Bin-Hua; Li, Cui-Lin; Zhang, Fan; Wang, Xian-Feng; Zhang, Ge; Bu, Xian-Zhang; Cai, Shao-Hui; Du, Jun

    2013-01-01

    Chronic inflammation-promoted metastasis has been considered as a major challenge in cancer therapy. Pro-inflammatory cytokine TNFα can induce cancer invasion and metastasis associated with epithelial-mesenchymal transition (EMT). However, the underlying mechanisms are not entirely clear. In this study, we showed that TNFα induces EMT in human HCT116 cells and thereby promotes colorectal cancer (CRC) invasion and metastasis. TNFα-induced EMT was characterized by acquiring mesenchymal spindle-like morphology and increasing the expression of N-cadherin and fibronectin with a concomitant decrease of E-cadherin and Zona occludin-1(ZO-1). TNFα treatment also increased the expression of transcription factor Snail, but not Slug, ZEB1 and Twist. Overexpression of Snail induced a switch from E-cadherin to N-cadherin expression in HCT116 cells, which is a characteristic of EMT. Conversely, knockdown of Snail significantly attenuated TNFα-induced EMT in HCT116 cells, suggesting that Snail plays a crucial role in TNFα-induced EMT. Interestingly, exposure to TNFα rapidly increased Snail protein expression and Snail nuclear localization but not mRNA level upregulation. Finally, we demonstrated that TNFα elevated Snail stability by activating AKT pathway and subsequently repressing GSK-3β activity and decreasing the association of Snail with GSK-3β. Knockdown of GSK-3β further verified our finding. Taken together, these results revealed that AKT/GSK-3β-mediated stabilization of Snail is required for TNFα-induced EMT in CRC cells. Our study provides a better understanding of inflammation-induced CRC metastasis.

  6. TGF-β and EGF induced HLA-I downregulation is associated with epithelial-mesenchymal transition (EMT) through upregulation of snail in prostate cancer cells.

    Science.gov (United States)

    Chen, Xiao-Hui; Liu, Zong-Cai; Zhang, Ge; Wei, Wei; Wang, Xiao-Xiong; Wang, Hao; Ke, Hong-Peng; Zhang, Fan; Wang, Hong-Sheng; Cai, Shao-Hui; Du, Jun

    2015-05-01

    Human leukocyte antigen class I antigens (HLA-I) is essential in immune response by presenting antigenic peptides to cytotoxic T lymphocytes. Downregulation of HLA-I is observed in primary and metastatic prostate cancers, which facilitates them escape from immune surveillance, thereby promotes prostate cancer progression. In addition, elevated level of growth factors like TGF-β or EGF in microenvironment is related to the prostate cancer deterioration. Thus, we wondered whether TGF-β or EGF was involved in the regulation of HLA-I during the development of prostate cancer cells. In this study, we demonstrated that TGF-β and EGF both downregulated the expression of HLA-I, thereby attenuated the cytotoxic T cell mediated lysis of prostate cancer cells. Next, we revealed that TGF-β and EGF induced downregulation of HLA-I is associated with classical epithelial-mesenchymal transition (EMT) morphological changes and expression profiles. We further illustrated that overexpression of Snail is crucial for HLA-I downregulation and its association with EMT. At last, we discussed that NF-κB/p65 is the plausible target for Snail to induce HLA-I downregulation. Taken together, this is the first evidence to reveal that both TGF-β and EGF can induce HLA-I downregulation which is then proven to be associated with EMT in prostate cancer cells. These discoveries provide a deeper understanding of growth factors induced immune escape and introduce potential therapeutic targets for prostate cancers. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Snail regulated by PKC/GSK-3β pathway is crucial for EGF-induced epithelial-mesenchymal transition (EMT) of cancer cells.

    Science.gov (United States)

    Liu, Zong-cai; Chen, Xiao-hui; Song, Hai-xing; Wang, Hong-sheng; Zhang, Ge; Wang, Hao; Chen, Dan-yang; Fang, Rui; Liu, Hao; Cai, Shao-hui; Du, Jun

    2014-11-01

    Cancer metastasis is considered a major challenge in cancer therapy. Recently, epidermal growth factor (EGF)/epidermal growth factor receptor (EGFR) signaling has been shown to induce epithelial-mesenchymal transition (EMT) and thereby to promote cancer metastasis. However, the underlying mechanism has not been fully elucidated. We demonstrate that EGF can induce EMT in human prostate and lung cancer cells and thus promote invasion and migration. EGF-induced EMT has been characterized by the cells acquiring mesenchymal spindle-like morphology and increasing their expression of N-cadherin and fibronectin, with a concomitant decrease of E-cadherin. Both protein and mRNA expression of transcription factor Snail rapidly increases after EGF treatment. The knockdown of Snail significantly attenuates EGF-induced EMT, suggesting that Snail is crucial for this process. To determine the way that Snail is accumulated, we demonstrate (1) that EGF promotes the stability of Snail via inhibiting the activity of glycogen synthase kinase 3 beta (GSK-3β), (2) that protein kinase C (PKC) rather than the phosphatidylinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway is responsible for GSK-3β inhibition and (3) that GSK-3β inhibition promotes the transcription of Snail. Taken together, these results reveal that the PKC/GSK-3β signaling pathway controls both the stability and transcription of Snail, which is crucial for EMT induced by EGF in PC-3 and A549 cells. Our study suggests a novel signaling pathway for Snail regulation and provides a better understanding of growth-factor-induced tumor EMT and metastasis.

  8. N-myc downstream-regulated gene 2 (NDRG2) suppresses the epithelial-mesenchymal transition (EMT) in breast cancer cells via STAT3/Snail signaling.

    Science.gov (United States)

    Kim, Myung-Jin; Lim, Jihyun; Yang, Young; Lee, Myeong-Sok; Lim, Jong-Seok

    2014-11-01

    Although NDRG2 has recently been found to be a candidate tumor suppressor, its precise role in the epithelial-mesenchymal transition (EMT) is not well understood. In the present study, we demonstrated that NDRG2 overexpression in MDA-MB-231 cells down-regulated the expression of Snail, a transcriptional repressor of E-cadherin and a key regulator of EMT, as well as the phosphorylation of signal transducer and activator of transcription 3 (STAT3), an oncogenic transcription factor that is activated in many human malignancies including breast cancer. In addition, we confirmed that the expression of Snail and phospho-STAT3 was recovered when NDRG2 was knocked down by siRNA in MCF7 cells in which NDRG2 is endogenously expressed. Interestingly, MDA-MB-231-NDRG2 cells showed remarkably decreased Snail expression after treatment with JSI-124 (also known as cucurbitacin I) or Stattic, STAT3 inhibitors, compared to MDA-MB-231-mock cells. Moreover, STAT3 activation by EGF treatment induced higher Snail expression, and NDRG2 overexpression resulted in the inhibition of Snail expression in MDA-MB-231 cells stimulated by EGF in the absence or presence of STAT3 inhibitor. Treatment of MDA-MB-231 cells with STAT3 inhibitor led to a moderate decrease in wound healing and migration capacity, whereas STAT3 inhibitor treatment of MDA-MB-231-NDRG2 cells resulted in a significant attenuation of migration in both resting and EGF-stimulated cells. Collectively, our data demonstrate that the inhibition of STAT3 signaling by NDRG2 suppresses EMT progression of EMT via the down-regulation of Snail expression. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  9. AKT/GSK-3β regulates stability and transcription of snail which is crucial for bFGF-induced epithelial-mesenchymal transition of prostate cancer cells.

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    Liu, Zong-Cai; Wang, Hong-Sheng; Zhang, Ge; Liu, Hao; Chen, Xiao-Hui; Zhang, Fan; Chen, Dan-Yang; Cai, Shao-Hui; Du, Jun

    2014-10-01

    Epithelial-mesenchymal transition (EMT) plays a pivotal role in the development of metastatic cancers. Basic fibroblast growth factor (bFGF) is significantly elevated in metastatic prostate cancers, which has been mentioned mainly to induce EMT in normal cells. However, there is no description about bFGF induced EMT and its underlying mechanism in prostate cancer cells. Western blotting, immunofluorescence and qRT-PCR assays were used to study protein or mRNA expression profiles of the EMT. Wound healing scratch, migration and invasion assays were used to test the motility of cells undergoing EMT. More methods were used to explore the underlying mechanisms. We demonstrated that bFGF promoted EMT and motility of human prostate cancer PC-3 cells. Both protein and mRNA expression of Snail were rapidly increased after bFGF treatment. Ectopic expression of Snail triggered EMT and enhanced cell motility in PC-3 cells, and knockdown of Snail almost abolished bFGF induced EMT, suggesting the critical role of Snail. Mechanistic study demonstrated that bFGF promoted the stability, nuclear localization and transcription of Snail by inhibiting the activity of glycogen synthase kinase 3 beta (GSK-3β) through phosphatidylinositide 3 kinases (PI3K)/protein kinase B (AKT) signaling pathway. It is concluded that bFGF can promote EMT and motility of PC-3 cells, and AKT/GSK-3β signaling pathway controls the stability, localization and transcription of Snail which is crucial for this bFGF induced EMT. To our knowledge, this is the first study to demonstrate that bFGF can induce EMT via AKT/GSK-3β/Snail signaling pathway in prostate cancer cells. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. miR-5003-3p promotes epithelial-mesenchymal transition in breast cancer cells through Snail stabilization and direct targeting of E-cadherin.

    Science.gov (United States)

    Kwak, Seo-Young; Yoo, Je-Ok; An, Hyun-Ju; Bae, In-Hwa; Park, Myung-Jin; Kim, Joon; Han, Young-Hoon

    2016-06-09

    One of the initial steps in metastatic dissemination is the epithelial-mesenchymal transition (EMT). Along this line, microRNAs (miRNAs) have been shown to function as important regulators of tumor progression at various stages. Therefore, we performed a functional screening for EMT-regulating miRNAs and identified several candidate miRNAs. Among these, we demonstrated that miR-5003-3p induces cellular features characteristic of EMT. miR-5003-3p induced upregulation of Snail, a key EMT-promoting transcription factor and transcriptional repressor of E-cadherin, through protein stabilization. MDM2 was identified as a direct target of miR-5003-3p, the downregulation of which induced Snail stabilization. E-cadherin was also demonstrated to be a direct target of miR-5003-3p, reinforcing the EMT-promoting function of miR-5003-3p. In situ hybridization and immunohistochemical analyses using tissue microarrays revealed that miR-5003-3p expression was higher in paired metastatic breast carcinoma tissues than in primary ductal carcinoma tissues, and was inversely correlated with the expression of MDM2 and E-cadherin. Furthermore, miR-5003-3p enhanced the formation of metastatic nodules in the lungs of mice in a tail vein injection experiment. Collectively, our results suggest that miR-5003-3p functions as a metastasis activator by promoting EMT through dual regulation of Snail stability and E-cadherin, and may therefore be a potential therapeutic target in metastatic cancers. © The Author (2016). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved.

  11. Epithelial-mesenchymal transition stimulates human cancer cells to extend microtubule-based invasive protrusions and suppresses cell growth in collagen gel.

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    Jun Oyanagi

    Full Text Available Epithelial-mesenchymal transition (EMT is a crucial event in tumor invasion and metastasis. However, most of past EMT studies have been conducted in the conventional two-dimensional (2D monolayer culture. Therefore, it remains unclear what invasive phenotypes are acquired by EMT-induced cancer cells. To address this point, we attempted to characterize EMT cells in more physiological, three-dimensional (3D collagen gel culture. EMT was induced by treating three human carcinoma cell lines (A549, Panc-1 and MKN-1 with TGF-ß. The TGF-ß treatment stimulated these cells to overexpress the invasion markers laminin γ2 and MT1-MMP in 2D culture, in addition to the induction of well-known morphological change and EMT marker expression. EMT induction enhanced cell motility and adhesiveness to fibronectin and collagen in 2D culture. Although EMT cells showed comparable cell growth to control cells in 2D culture, their growth rates were extremely suppressed in soft agar and collagen gel cultures. Most characteristically, EMT-induced cancer cells commonly and markedly extended invasive protrusions in collagen gel. These protrusions were mainly supported by microtubules rather than actin cytoskeleton. Snail-introduced, stable EMT cells showed similar protrusions in 3D conditions without TGF-ß. Moreover, these protrusions were suppressed by colchicine or inhibitors of heat shock protein 90 (HSP-90 and protein phosphatase 2A. However, MMP inhibitors did not suppress the protrusion formation. These data suggest that EMT enhances tumor cell infiltration into interstitial stroma by extending microtubule-based protrusions and suppressing cell growth. The elevated cell adhesion to fibronectin and collagen and high cell motility also seem important for the tumor invasion.

  12. Inositol-requiring protein 1 - X-box-binding protein 1 pathway promotes epithelial-mesenchymal transition via mediating snail expression in pulmonary fibrosis.

    Science.gov (United States)

    Mo, Xiao-Ting; Zhou, Wen-Cheng; Cui, Wen-Hui; Li, De-Lin; Li, Liu-Cheng; Xu, Liang; Zhao, Ping; Gao, Jian

    2015-08-01

    Epithelial-mesenchymal transition (EMT) is a complex biological program during which cells loss epithelial phenotype and acquire mesenchymal features. EMT is thought to be involved in the pathogenesis of various fibrotic diseases including pulmonary fibrosis (PF). Recent studies suggest that endoplasmic reticulum (ER) stress is associated with EMT in the progression of PF. However, the exact mechanism is unclear. Here, we developed a PF model with bleomycin (BLM) administration in rats and conducted several simulation experiments in alveolar epithelial cell (AECs) RLE-6TN to unravel the role of inositol-requiring protein 1 (IRE1) - X-box-binding protein 1 (XBP1) signal pathway in ER stress-induced EMT in PF. First, we observed that ER stress was occurred in type II AECs accompanied by EMT in BLM-induced PF. Then we explored the role of IRE1-XBP1-snail pathway in transforming growth factor (TGF)-β1/tunicamycin (TM)-induced EMT. When TGF-β1/TM was treated on AECs, IRE1 and XBP1 were overexpressed, meanwhile, snail expression was upregulated accompanied with EMT. However, when IRE1 or XBP1 was knockdown, TGF-β1/TM-induced EMT were blocked while the expression of snail was inhibited. Then we silenced snail and found that TGF-β1/TM-induced EMT were also suppressed, but it had no effect on the up-regulated expression of IRE1 and XBP1. Thus, we concluded that IRE1-XBP1 pathway promotes EMT via mediating snail expression in PF. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. Osthole inhibited TGF β-induced epithelial-mesenchymal transition (EMT) by suppressing NF-κB mediated Snail activation in lung cancer A549 cells.

    Science.gov (United States)

    Feng, Haitao; Lu, Jin-Jian; Wang, Yitao; Pei, Lixia; Chen, Xiuping

    2017-02-01

    Epithelial-mesenchymal transition (EMT), the transdifferentiation of epithelial cells into mesenchymal cells, has been implicated in the metastasis and provides novel strategies for cancer therapy. Osthole (OST), a dominant active constituent of Chinese herb Cnidium monnieri, has been reported to inhibit cancer metastasis while the mechanisms remains unclear. Here, we studied the inhibitory effect and mechanisms of OST on TGF-β1-induced EMT in A549 cells. Cells were treated with TGF-β1 in the absence and presence of OST. The morphological alterations were observed with a microscopy. The protein and mRNA expressions were determined by Western blotting and real-time PCR. The protein localization was detected with immunofluorescence. The adhesion, migration, and invasion were determined by Matrigel, wound-healing, and Transwell assays. TGF-β1 treatment induced spindle-shaped alterations of cells, upregulation of N-cadherin, Vimentin, NF-κB p65, and downregulation of E-cadherin. Dysregulated membrane expression and mRNA expression of E-cadherin and N-cadherin were observed after TGF-β1 treatment. TGF-β1 increased abilities of migration and invasion and triggered the nuclear translocation of NF-κB p65. These alterations were dramatically inhibited by OST. Furthermore, PDTC, a NF-κB inhibitor, showed similar effects. In addition, TGF-β1-induced expression of Snail was significantly inhibited by OST and silenced Snail partially reversed TGF-β1-induced EMT biomarkers without affecting NF-κB p-65. In conclusion, OST inhibited TGF-β1-induced EMT, adhesion, migration, and invasion through inactivation of NF-κB-Snail pathways in A549 cells. This study provides novel molecular mechanisms for the anti-metastatic effect of OST.

  14. Embryonic stem cells markers SOX2, OCT4 and Nanog expression and their correlations with epithelial-mesenchymal transition in nasopharyngeal carcinoma.

    Directory of Open Access Journals (Sweden)

    Weiren Luo

    Full Text Available Expression of embryonic stem cells (ESCs markers (SOX2, OCT4, Nanog and Nestin is crucial for progression of various human malignancies. The purpose of this study was to investigate the expression and prognostic impact of these molecules in nasopharyngeal carcinoma (NPC patients by immunohistochemistry and immunofluorescence. In the present study, we found that the expression levels of SOX2, OCT4 and Nanog were highly expressed in NPC compared with the non-tumorous tissues. Furthermore, these proteins correlated significantly with several clinicalpathological factors and epithelial-mesenchymal transition (EMT-associated indicators (E-cadherin/N-cadherin and Snail. In multivariate analyses, high expression of OCT4 (P = 0.013 and Nanog (P = 0.040, but not that of SOX2, was associated with worse survival and had strongly independent prognostic effects. Of note, OCT4 and Nanog were more frequently located at the invasive front of tumors, and correlated significantly with various aggressive behaviors including T classification, N classification, M classification and clinical stage. Furthermore, patients with co-expression of OCT4 and Nanog in the invasive front had significantly worse survival (P = 0.005. Interestingly, at the invasive front, these molecules correlated significantly with Nestin expression in endothelial cells (P<0.001. These findings provide evidence that ESCs biomarkers OCT4 and Nanog serves as independent prognostic factors for NPC. Additionally, cancer cells in the invasive front of NPC acquiring ESCs-like features should be maintained by vascular niches.

  15. Autocrine IL-8 and VEGF mediate epithelial-mesenchymal transition and invasiveness via p38/JNK-ATF-2 signalling in A549 lung cancer cells.

    Science.gov (United States)

    Desai, Sejal; Laskar, S; Pandey, B N

    2013-09-01

    Soluble factors in tumour microenvironment play a major role in modulating the metastatic potential of cancer cells. Herein, we investigated the effect of autocrine cytokines and growth factors in the form of self-conditioned medium (CM) on A549 lung carcinoma cells. We demonstrated that CM induced morphological and molecular changes associated with epithelial-mesenchymal transition viz change in shape from cuboidal to spindle, actin cytoskeleton remodelling, upregulation of vimentin and downregulation of E-cadherin etc. These changes were accompanied with enhanced motility, invasion, anchorage-independent growth and anoikis-resistance. Amongst the different factors of CM, IL-8 and VEGF were found to play a major role in the CM-induced motility and invasion. In the intracellular signalling cascade, CM triggered phosphorylation of JNK and p38 which was associated with the CM-enhanced invasiveness. In CM-treated cells, activated p38 and JNK further activated ATF-2 (Activating Transcription Factor-2) and knock-down of ATF-2 abrogated the CM-induced invasiveness, suggesting the signal transduction along the p38/JNK-ATF-2 axis. Furthermore, neutralising IL-8 and VEGF in CM, significantly abrogated CM-induced phosphorylation of ATF-2. Conversely, exogenous addition of these individual cytokines in plain medium, increased the activation of ATF-2 and invasiveness marginally. However, when added in combination these cytokines (IL-8 and VEGF) resulted in drastic increase in ATF-2 phosphorylation and subsequent invasiveness suggesting their synergetic interplay in the observed phenomenon. Taken together, our results identify IL-8/VEGF induced JNK/p38-ATF-2 as a novel pro-invasive pathway, which may be explored as potential therapeutic target to circumvent the invasiveness of lung malignancies. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. miR-1271 inhibits migration, invasion and epithelial-mesenchymal transition by targeting ZEB1 and TWIST1 in pancreatic cancer cells

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    Liu, Huaize [Department of Developmental Genetics, Nanjing Medical University, Nanjing 210029 (China); Wang, Han [The First Clinical Medical College of Nanjing Medical University, Nanjing 210029 (China); Liu, Xiaoxiao [Department of Biotechnology, Nanjing Medical University, Nanjing 210029 (China); Yu, Tingting, E-mail: tingting@njmu.edu.cn [Department of Developmental Genetics, Nanjing Medical University, Nanjing 210029 (China)

    2016-04-01

    Pancreatic cancer (PC) remains one of the most lethal types of cancer in adults. The purpose of this study was to determine the role of miR-1271 in regulation of epithelial mesenchymal transition (EMT) and metastasis of pancreatic cancer cells. miR-1271 was identified to be significantly down-regulated in PC tissues by miRNA array. Also, an increase of EMT-regulators ZEB1 and TWIST1 expression level is accompanied by a decrease of miR-1271. We showed that expression of miR-1271 was significantly down-regulated in PC tissues as compared with that in normal tissues. In addition, our results showed that miR-1271 expression levels were decreased while ZEB1 and TWIST1 expression levels were increased in detected PC cell lines. Moreover, ectopic expression of miR-1271 suppressed and antagomiR-1271 promoted proliferation, migration, and invasion in SW1990 and PANC-1 cells. Bioinformatics coupled with luciferase and Western blot assays also revealed that miR-1271 inhibited expression of ZEB1 and TWIST1, which are master regulators of tumor metastasis. Our study first indicates that miR-1271 functions as a suppressor in regulating of pancreatic cancer EMT by targeting ZEB1 and TWIST1, and it promise as a therapeutic target and prognostic marker for metastatic pancreatic cancer. - Highlights: • miR-1271 is downregulated in pancreatic cancer tissues and cell lines. • miR-1271 regulates cell metastasis ability and EMT marker expression. . • miR-1271 directly targets ZEB1 and TWIST1. • ZEB1 and TWIST1 are functionally related to the effects of miR-1271.

  17. Salinomycin induces cell death and differentiation in head and neck squamous cell carcinoma stem cells despite activation of epithelial-mesenchymal transition and Akt

    Directory of Open Access Journals (Sweden)

    Kuo Selena Z

    2012-11-01

    Full Text Available Abstract Background Cancer stem cells (CSC are believed to play a crucial role in cancer recurrence due to their resistance to conventional chemotherapy and capacity for self-renewal. Recent studies have reported that salinomycin, a livestock antibiotic, selectively targets breast cancer stem cells 100-fold more effectively than paclitaxel. In our study we sought to determine the effects of salinomycin on head and neck squamous cell carcinoma (HNSCC stem cells. Methods MTS and TUNEL assays were used to study cell proliferation and apoptosis as a function of salinomycin exposure in JLO-1, a putative HNSCC stem cell culture. MTS and trypan blue dye exclusion assays were performed to investigate potential drug interactions between salinomycin and cisplatin or paclitaxel. Stem cell-like phenotype was measured by mRNA expression of stem cell markers, sphere-forming capacity, and matrigel invasion assays. Immunoblotting was also used to determine expression of epithelial-mesenchymal transition (EMT markers and Akt phosphorylation. Arrays by Illumina, Inc. were used to profile microRNA expression as a function of salinomycin dose. Results In putative HNSCC stem cells, salinomycin was found to significantly inhibit cell viability, induce a 71.5% increase in levels of apoptosis, elevate the Bax/Bcl-2 ratio, and work synergistically with cisplatin and paclitaxel in inducing cell death. It was observed that salinomycin significantly inhibited sphere forming-capability and repressed the expression of CD44 and BMI-1 by 3.2-fold and 6.2-fold, respectively. Furthermore, salinomycin reduced invasion of HNSCC stem cells by 2.1 fold. Contrary to expectations, salinomycin induced the expression of EMT markers Snail, vimentin, and Zeb-1, decreased expression of E-cadherin, and also induced phosphorylation of Akt and its downstream targets GSK3-β and mTOR. Conclusions These results demonstrate that in HNSCC cancer stem cells, salinomycin can cause cell death and

  18. MiR-21-5p Links Epithelial-Mesenchymal Transition Phenotype with Stem-Like Cell Signatures via AKT Signaling in Keloid Keratinocytes

    Science.gov (United States)

    Yan, Li; Cao, Rui; Liu, Yuanbo; Wang, Lianzhao; Pan, Bo; Lv, Xiaoyan; Jiao, Hu; Zhuang, Qiang; Sun, Xuejian; Xiao, Ran

    2016-09-01

    Keloid is the abnormal wound healing puzzled by the aggressive growth and high recurrence rate due to its unrevealed key pathogenic mechanism. MicroRNAs contribute to a series of biological processes including epithelial-mesenchymal transition (EMT) and cells stemness involved in fibrotic disease. Here, using microRNAs microarray analysis we found mir-21-5p was significantly up-regulated in keloid epidermis. To investigate the role of miR-21-5p in keloid pathogenesis, we transfected miR-21-5p mimic or inhibitor in keloid keratinocytes and examined the abilities of cell proliferation, apoptosis, migration and invasion, the expressions of EMT-related markers vimentin and E-cadherin and stem-like cells-associated markers CD44 and ALDH1, and the involvement of PTEN and the signaling of AKT and ERK. Our results demonstrated that up-regulation or knockdown of miR-21-5p significantly increased or decreased the migration, invasion and sphere-forming abilities of keloid keratinocytes, and the phenotype of EMT and cells stemness were enhanced or reduced as well. Furthermore, PTEN and p-AKT were shown to participate in the regulation of miR-21-5p on EMT phenotypes and stemness signatures of keloid keratinocytes, which might account for the invasion and recurrence of keloids. This molecular mechanism of miR-21-5p on keloid keratinocytes linked EMT with cells stemness and implicated novel therapeutic targets for keloids.

  19. miR-187-3p inhibits the metastasis and epithelial-mesenchymal transition of hepatocellular carcinoma by targeting S100A4.

    Science.gov (United States)

    Dou, Changwei; Liu, Zhikui; Xu, Meng; Jia, Yuli; Wang, Yufeng; Li, Qing; Yang, Wei; Zheng, Xin; Tu, Kangsheng; Liu, Qingguang

    2016-10-28

    miR-187-3p, a novel cancer-related microRNA, was previously reported to play promoting or suppressive roles in different malignancies. However, the expression level, biological function, and underlying mechanisms of miR-187-3p in hepatocellular carcinoma (HCC) remain unknown. This study demonstrated that miR-187-3p was significantly down-regulated in HCC tissues and cell lines, and was associated with advanced TNM stage and metastasis in HCC. Functional studies confirmed that miR-187-3p could inhibit the metastasis of HCC both in vitro and in vivo. Moreover, we proved that miR-187-3p could prevent the epithelial-mesenchymal transition (EMT) of HCC cells. Mechanically, S100A4 was a direct downstream target of miR-187-3p, and mediated the functional influence of miR-187-3p in HCC. Furthermore, miR-187-3p and S100A4 expression was evidently correlated with adverse clinical features and poor prognosis of HCC. Lastly, we showed that hypoxia was responsible for the significantly decreased level of miR-187-3p in HCC, and miR-187-3p was involved in the promoting effects of hypoxia on the metastasis and EMT of HCC cells. Taken together, miR-187-3p inhibits the metastasis and EMT in HCC by targeting S100A4. miR-187-3p can serve as a prognostic indicator and a promising therapeutic target for HCC patients.

  20. Downregulation of tight junction-associated MARVEL protein marvelD3 during epithelial-mesenchymal transition in human pancreatic cancer cells.

    Science.gov (United States)

    Kojima, Takashi; Takasawa, Akira; Kyuno, Daisuke; Ito, Tatsuya; Yamaguchi, Hiroshi; Hirata, Koichi; Tsujiwaki, Mitsuhiro; Murata, Masaki; Tanaka, Satoshi; Sawada, Norimasa

    2011-10-01

    The novel tight junction protein marvelD3 contains a conserved MARVEL (MAL and related proteins for vesicle trafficking and membrane link) domain like occludin and tricellulin. However, little is yet known about the detailed role and regulation of marvelD3 in normal epithelial cells and cancer cells, including pancreatic cancer. In the present study, we investigated marvelD3 expression in well and poorly differentiated human pancreatic cancer cell lines and normal pancreatic duct epithelial cells in which the hTERT gene was introduced into human pancreatic duct epithelial cells in primary culture, and the changes of marvelD3 during Snail-induced epithelial-mesenchymal transition (EMT) under hypoxia, TGF-β treatment and knockdown of FOXA2 in well differentiated pancreatic cancer HPAC cells. MarvelD3 was transcriptionally downregulated in poorly differentiated pancreatic cancer cells and during Snail-induced EMT of pancreatic cancer cells in which Snail was highly expressed and the fence function downregulated, whereas it was maintained in well differentiated human pancreatic cancer cells and normal pancreatic duct epithelial cells. Depletion of marvelD3 by siRNAs in HPAC cells resulted in downregulation of barrier functions indicated as a decrease in transepithelial electric resistance and an increase of permeability to fluorescent dextran tracers, whereas it did not affect fence function of tight junctions. In conclusion, marvelD3 is transcriptionally downregulated in Snail-induced EMT during the progression for the pancreatic cancer. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Epidermal growth factor-like domain-containing protein 7 (EGFL7 enhances EGF receptor-AKT signaling, epithelial-mesenchymal transition, and metastasis of gastric cancer cells.

    Directory of Open Access Journals (Sweden)

    Bai-Hua Luo

    Full Text Available Epidermal growth factor-like domain-containing protein 7 (EGFL7 is upregulated in human epithelial tumors and so is a potential biomarker for malignancy. Indeed, previous studies have shown that high EGFL7 expression promotes infiltration and metastasis of gastric carcinoma. The epithelial-mesenchymal transition (EMT initiates the metastatic cascade and endows cancer cells with invasive and migratory capacity; however, it is not known if EGFL7 promotes metastasis by triggering EMT. We found that EGFL7 was overexpressed in multiple human gastric cancer (GC cell lines and that overexpression promoted cell invasion and migration as revealed by scratch wound and transwell migration assays. Conversely, shRNA-mediated EGFL7 knockdown reduced invasion and migration. Furthermore, EGFL7-overexpressing cells grew into larger tumors and were more likely to metastasize to the liver compared to underexpressing CG cells following subcutaneous injection in mice. EGFL7 overexpression protected GC cell lines against anoikis, providing a plausible mechanism for this enhanced metastatic capacity. In excised human gastric tumors, expression of EGFL7 was positively correlated with expression levels of the mesenchymal marker vimentin and the EMT-associated transcription repressor Snail, and negatively correlated with expression of the epithelial cell marker E-cadherin. In GC cell lines, EGFL7 knockdown reversed morphological signs of EMT and decreased both vimentin and Snail expression. In addition, EGFL7 overexpression promoted EGF receptor (EGFR and protein kinase B (AKT phospho-activation, effects markedly suppressed by the EGFR tyrosine kinase inhibitor AG1478. Moreover, AG1478 also reduced the elevated invasive and migratory capacity of GC cell lines overexpressing EGFL7. Collectively, these results strongly suggest that EGFL7 promotes metastasis by activating EMT through an EGFR-AKT-Snail signaling pathway. Disruption of EGFL7-EGFR-AKT-Snail signaling may a

  2. Withaferin A inhibits experimental epithelial-mesenchymal transition in MCF-10A cells and suppresses vimentin protein level in vivo in breast tumors.

    Science.gov (United States)

    Lee, Joomin; Hahm, Eun-Ryeong; Marcus, Adam I; Singh, Shivendra V

    2015-06-01

    We have shown previously that withaferin A (WA), a bioactive component of the medicinal plant Withania somnifera, inhibits growth of cultured and xenografted human breast cancer cells and prevents breast cancer development and pulmonary metastasis incidence in a transgenic mouse model. The present study was undertaken to determine if the anticancer effect of WA involved inhibition of epithelial-mesenchymal transition (EMT). Experimental EMT induced by exposure of MCF-10A cells to tumor necrosis factor-α (TNF-α) and transforming growth factor-β1 (TGF-β) was partially reversed by treatment with WA but not by its structural analogs withanone or withanolide A. Combined TNF-α and TGF-β treatments conferred partial protection against MCF-10A cell migration inhibition by WA. Inhibition of TNF-α and TGF-β-induced MCF-10A cell migration by WA exposure was modestly attenuated by knockdown of E-cadherin protein. MCF-7 and MDA-MB-231 cells exposed to WA exhibited sustained (MCF-7) or transient (MDA-MB-231) induction of E-cadherin protein. On the other hand, the level of vimentin protein was increased markedly after 24 h treatment of MDA-MB-231 cells with WA. WA-induced apoptosis was not affected by vimentin protein knockdown in MDA-MB-231 cells. Protein level of vimentin was significantly lower in the MDA-MB-231 xenografts as well as in MMTV-neu tumors from WA-treated mice compared with controls. The major conclusions of the present study are that (a) WA treatment inhibits experimental EMT in MCF-10A cells, and (b) mammary cancer growth inhibition by WA administration is associated with suppression of vimentin protein expression in vivo.

  3. Combined niclosamide with cisplatin inhibits epithelial-mesenchymal transition and tumor growth in cisplatin-resistant triple-negative breast cancer.

    Science.gov (United States)

    Liu, Junjun; Chen, Xiaosong; Ward, Toby; Pegram, Mark; Shen, Kunwei

    2016-07-01

    Women with triple-negative breast cancer have worse prognosis compared to other breast cancer subtypes. Acquired drug resistance remains to be an important reason influencing triple-negative breast cancer treatment efficacy. A prevailing theory postulates that the cancer resistance and recurrence results from a subpopulation of tumor cells with stemness program, which are often insensitive to cytotoxic drugs such as cisplatin. Recent studies suggested that niclosamide, an anti-helminthic drug, has potential therapeutic activities against breast cancer stem cells, which prompts us to determine its roles on eliminating cisplatin-resistant cancer cells. Hence, we established a stable cisplatin-resistant MDA-MB-231 cell line (231-CR) through continuously exposure to increasing concentrations of cisplatin (5-20 μmol/l). Interestingly, 231-CR exhibited properties associated to epithelial-mesenchymal transition with enhanced invasion, preserved proliferation, increased mammosphere formation, and reduced apoptosis compared to naive MDA-MB-231 sensitive cells (231-CS). Importantly, niclosamide or combination with cisplatin inhibited both 231-CS and 231-CR cell proliferation in vitro. In addition, niclosamide reversed the EMT phenotype of 231-CR by downregulation of snail and vimentin. Mechanistically, niclosamide treatment in combination with or without cisplatin significantly inhibited Akt, ERK, and Src signaling pathways. In vivo study showed that niclosamide or combination with cisplatin could repress the growth of xenografts originated from either 231-CS or 231-CR cells, with prominent suppression of Ki67 expression. These findings suggested that niclosamide might serve as a novel therapeutic strategy, either alone or in combination with cisplatin, for triple-negative breast cancer treatment, especially those resistant to cisplatin.

  4. Epithelial-mesenchymal transition and cancer stem cells, mediated by a long non-coding RNA, HOTAIR, are involved in cell malignant transformation induced by cigarette smoke extract

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    Liu, Yi; Luo, Fei; Xu, Yuan; Wang, Bairu; Zhao, Yue; Xu, Wenchao; Shi, Le; Lu, Xiaolin; Liu, Qizhan, E-mail: drqzliu@hotmail.com

    2015-01-01

    The incidence of lung diseases, including cancer, caused by cigarette smoke is increasing, but the molecular mechanisms of gene regulation induced by cigarette smoke remain unclear. This report describes a long noncoding RNA (lncRNA) that is induced by cigarette smoke extract (CSE) and experiments utilizing lncRNAs to integrate inflammation with the epithelial-mesenchymal transition (EMT) in human bronchial epithelial (HBE) cells. The present study shows that, induced by CSE, IL-6, a pro-inflammatory cytokine, leads to activation of STAT3, a transcription activator. A ChIP assay determined that the interaction of STAT3 with the promoter regions of HOX transcript antisense RNA (HOTAIR) increased levels of HOTAIR. Blocking of IL-6 with anti-IL-6 antibody, decreasing STAT3, and inhibiting STAT3 activation reduced HOTAIR expression. Moreover, for HBE cells cultured in the presence of HOTAIR siRNA for 24 h, the CSE-induced EMT, formation of cancer stem cells (CSCs), and malignant transformation were reversed. Thus, IL-6, acting on STAT3 signaling, which up-regulates HOTAIR in an autocrine manner, contributes to the EMT and to CSCs induced by CSE. These data define a link between inflammation and EMT, processes involved in the malignant transformation of cells caused by CSE. This link, mediated through lncRNAs, establishes a mechanism for CSE-induced lung carcinogenesis. - Highlights: • STAT3 directly regulates the levels of LncRNA HOTAIR. • LncRNA HOTAIR mediates the link between inflammation and EMT. • LncRNA HOTAIR is involved in the malignant transformation of cells caused by CSE.

  5. Overexpression of angiotensin II type 1 receptor in breast cancer cells induces epithelial-mesenchymal transition and promotes tumor growth and angiogenesis.

    Science.gov (United States)

    Oh, Eunhye; Kim, Ji Young; Cho, Youngkwan; An, Hyunsook; Lee, Nahyun; Jo, Hunho; Ban, Changill; Seo, Jae Hong

    2016-06-01

    The angiotensin II type I receptor (AGTR1) has been implicated in diverse aspects of human disease, from the regulation of blood pressure and cardiovascular homeostasis to cancer progression. We sought to investigate the role of AGTR1 in cell proliferation, epithelial-mesenchymal transition (EMT), migration, invasion, angiogenesis and tumor growth in the breast cancer cell line MCF7. Stable overexpression of AGTR1 was associated with accelerated cell proliferation, concomitant with increased expression of survival factors including poly(ADP-ribose) polymerase (PARP) and X-linked inhibitor of apoptosis (XIAP), as well as extracellular signal-regulated kinase (ERK) activation. AGTR1-overexpressing MCF7 cells were more aggressive than their parent line, with significantly increased activity in migration and invasion assays. These observations were associated with changes in EMT markers, including reduced E-cadherin expression and increased p-Smad3, Smad4 and Snail levels. Treatment with the AGTR1 antagonist losartan attenuated these effects. AGTR1 overexpression also accelerated tumor growth and increased Ki-67 expression in a xenograft model. This was associated with increased tumor angiogenesis, as evidenced by a significant increase in microvessels in the intratumoral and peritumoral areas, and enhanced tumor invasion, with the latter response associated with increased EMT marker expression and matrix metallopeptidase 9 (MMP-9) upregulation. In vivo administration of losartan significantly reduced both tumor growth and angiogenesis. Our findings suggest that AGTR1 plays a significant role in tumor aggressiveness, and its inhibition may have therapeutic implications. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. The T box transcription factor TBX2 promotes epithelial-mesenchymal transition and invasion of normal and malignant breast epithelial cells.

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    Bin Wang

    Full Text Available The T box transcription factor TBX2, a master regulator of organogenesis, is aberrantly amplified in aggressive human epithelial cancers. While it has been shown that overexpression of TBX2 can bypass senescence, a failsafe mechanism against cancer, its potential role in tumor invasion has remained obscure. Here we demonstrate that TBX2 is a strong cell-autonomous inducer of the epithelial-mesenchymal transition (EMT, a latent morphogenetic program that is key to tumor progression from noninvasive to invasive malignant states. Ectopic expression of TBX2 in normal HC11 and MCF10A mammary epithelial cells was sufficient to induce morphological, molecular, and behavioral changes characteristic of EMT. These changes included loss of epithelial adhesion and polarity gene (E-cadherin, ß-catenin, ZO1 expression, and abnormal gain of mesenchymal markers (N-cadherin, Vimentin, as well as increased cell motility and invasion. Conversely, abrogation of endogenous TBX2 overexpression in the malignant human breast carcinoma cell lines MDA-MB-435 and MDA-MB-157 led to a restitution of epithelial characteristics with reciprocal loss of mesenchymal markers. Importantly, TBX2 inhibition abolished tumor cell invasion and the capacity to form lung metastases in a Xenograft mouse model. Meta-analysis of gene expression in over one thousand primary human breast tumors further showed that high TBX2 expression was significantly associated with reduced metastasis-free survival in patients, and with tumor subtypes enriched in EMT gene signatures, consistent with a role of TBX2 in oncogenic EMT. ChIP analysis and cell-based reporter assays further revealed that TBX2 directly represses transcription of E-cadherin, a tumor suppressor gene, whose loss is crucial for malignant tumor progression. Collectively, our results uncover an unanticipated link between TBX2 deregulation in cancer and the acquisition of EMT and invasive features of epithelial tumor cells.

  7. 上皮向间质转化在消化道肿瘤中的研究进展%Epithelial-mesenchymal transition in digestive system tumors

    Institute of Scientific and Technical Information of China (English)

    许春红; 邹晓平

    2012-01-01

    Recent studies show that tumor cells get rid of the connections between cells,and induce tumor invasion and metastasis through epithelial-mesenchymal transition (EMT).EMT becomes an important way to invasion,metastasis and chemotherapy resistance of epithelial cell carcinoma which accounting for more than 90% of malignant carcinomas.Members of Snail family,especially Snail is regarded as important adjustment factor of EMT,which induce the transformation from epithelial cell to mesenchymal phenotype through competitive inhibition E-calcium protein gene expression.Many researches show that EMT exists widely in digestive system tumors,which is closly related to the invasion,metastasis and chemotherapy resistance of digestive system tumors.%近年研究发现,肿瘤细胞发生上皮向间质转化(EMT)摆脱细胞与细胞间连接而发生转移侵袭,已成为上皮细胞癌转移侵袭以及化疗耐药的一个重要途径.Snail家族成员,尤其是Snail被认为是肿瘤EMT发生的重要调节因子,它可以通过竞争性抑制E-钙黏蛋白基因的表达,引起上皮细胞向间质细胞表型的转变,最终引起EMT.多项研究显示,在消化系统肿瘤中普遍存在EMT现象,并与这些肿瘤的侵袭转移及化疗耐药密切相关.

  8. miR-181b-3p promotes epithelial-mesenchymal transition in breast cancer cells through Snail stabilization by directly targeting YWHAG.

    Science.gov (United States)

    Yoo, Je-Ok; Kwak, Seo-Young; An, Hyun-Ju; Bae, In-Hwa; Park, Myung-Jin; Han, Young-Hoon

    2016-07-01

    Epithelial-mesenchymal transition (EMT) is essential for increased invasion and metastasis during cancer progression. Among the candidate EMT-regulating microRNAs that we previously identified, miR-181b-3p was found to induce EMT in MCF7 breast cancer cells, as indicated by an EMT-characteristic morphological change, increased invasiveness, and altered expression of an EMT marker. Transfection with a miR-181b-3p inhibitor reduced the expression of mesenchymal markers and the migration and invasion of highly invasive breast cancer cells. miR-181b-3p induced the upregulation of Snail, a master EMT inducer and transcriptional repressor of E-cadherin, through protein stabilization. YWHAG was identified as a direct target of miR-181b-3p, downregulation of which induced Snail stabilization and EMT phenotypes. Ectopic expression of YWHAG abrogated the effect of miR-181b-3p, including Snail stabilization and the promotion of invasion. In situ hybridization and immunohistochemical analyses indicated that YWHAG expression was inversely correlated with the expression of miR-181b-3p and Snail in human breast cancer tissues. Furthermore, transfection with miR-181b-3p increased the frequency of metastatic nodule formation in the lungs of mice in experimental metastasis assays using MDA-MB-231 cells. Taken together, our data suggest that miR-181b-3p functions as a metastasis activator by promoting Snail-induced EMT, and may therefore be a therapeutic target in metastatic cancers.

  9. Hes1 triggers epithelial-mesenchymal transition (EMT)-like cellular marker alterations and promotes invasion and metastasis of nasopharyngeal carcinoma by activating the PTEN/AKT pathway.

    Science.gov (United States)

    Wang, Sheng-Chun; Lin, Xiao-Lin; Wang, Hui-Yan; Qin, Yu-Juan; Chen, Lin; Li, Jing; Jia, Jun-Shuang; Shen, Hong-Fen; Yang, Sheng; Xie, Rao-Ying; Wei, Fang; Gao, Fei; Rong, Xiao-Xiang; Yang, Jie; Zhao, Wen-Tao; Zhang, Ting-Ting; Shi, Jun-Wen; Yao, Kai-Tai; Luo, Wei-Ren; Sun, Yan; Xiao, Dong

    2015-11-03

    Overexpression of the transcriptional factor Hes1 (hairy and enhancer of split-1) has been observed in numerous cancers, but the precise roles of Hes1 in epithelial-mesenchymal transition (EMT), cancer invasion and metastasis remain unknown. Our current study firstly revealed that Hes1 upregulation in a cohort of human nasopharyngeal carcinoma (NPC) biopsies is significantly associated with the EMT, invasive and metastatic phenotypes of cancer. In the present study, we found that Hes1 overexpression triggered EMT-like cellular marker alterations of NPC cells, whereas knockdown of Hes1 through shRNA reversed the EMT-like phenotypes, as strongly supported by Hes1-mediated EMT in NPC clinical specimens described above. Gain-of-function and loss-of-function experiments demonstrated that Hes1 promoted the migration and invasion of NPC cells in vitro. In addition, exogenous expression of Hes1 significantly enhanced the metastatic ability of NPC cells in vivo. Chromatin immunoprecipitation (ChIP) assays showed that Hes1 inhibited PTEN expression in NPC cells through binding to PTEN promoter region. Increased Hes1 expression and decreased PTEN expression were also observed in a cohort of NPC biopsies. Additional studies demonstrated that Hes1-induced EMT-like molecular changes and increased motility and invasion of NPC cells were mediated by PTEN. Taken together, our results suggest, for what we believe is the first time, that Hes1 plays an important role in the invasion and metastasis of NPC through inhibiting PTEN expression to trigger EMT-like phenotypes.

  10. A CD44high/EGFRlow subpopulation within head and neck cancer cell lines shows an epithelial-mesenchymal transition phenotype and resistance to treatment.

    Directory of Open Access Journals (Sweden)

    Linnea La Fleur

    Full Text Available Mortality in head and neck squamous cell carcinoma (HNSCC is high due to emergence of therapy resistance which results in local and regional recurrences that may have their origin in resistant cancer stem cells (CSCs or cells with an epithelial-mesenchymal transition (EMT phenotype. In the present study, we investigate the possibility of using the cell surface expression of CD44 and epidermal growth factor receptor (EGFR, both of which have been used as stem cell markers, to identify subpopulations within HNSCC cell lines that differ with respect to phenotype and treatment sensitivity. Three subpopulations, consisting of CD44(high/EGFR(low, CD44(high/EGFR(high and CD44(low cells, respectively, were collected by fluorescence-activated cell sorting. The CD44(high/EGFR(low population showed a spindle-shaped EMT-like morphology, while the CD44(low population was dominated by cobblestone-shaped cells. The CD44(high/EGFR(low population was enriched with cells in G0/G1 and showed a relatively low proliferation rate and a high plating efficiency. Using a real time PCR array, 27 genes, of which 14 were related to an EMT phenotype and two with stemness, were found to be differentially expressed in CD44(high/EGFR(low cells in comparison to CD44(low cells. Moreover, CD44(high/EGFR(low cells showed a low sensitivity to radiation, cisplatin, cetuximab and gefitinib, and a high sensitivity to dasatinib relative to its CD44(high/EGFR(high and CD44(low counterparts. In conclusion, our results show that the combination of CD44 (high and EGFR (low cell surface expression can be used to identify a treatment resistant subpopulation with an EMT phenotype in HNSCC cell lines.

  11. Inhibition of SK4 Potassium Channels Suppresses Cell Proliferation, Migration and the Epithelial-Mesenchymal Transition in Triple-Negative Breast Cancer Cells.

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    Panshi Zhang

    Full Text Available Treatments for triple-negative breast cancer (TNBC are limited; intermediate-conductance calcium-activated potassium (SK4 channels are closely involved in tumor progression, but little is known about these channels in TNBC. We aimed to investigate whether SK4 channels affect TNBC. First, by immunohistochemistry (IHC and western blotting (WB, increased SK4 protein expression in breast tumor tissues was detected relative to that in non-tumor breast tissues, but there was no apparent expression difference between various subtypes of breast cancer (p>0.05. Next, functional SK4 channels were detected in the TNBC cell line MDA-MB-231 using WB, real-time PCR, immunofluorescence and patch-clamp recording. By employing SK4 specific siRNAs and blockers, including TRAM-34 and clotrimazole, in combination with an MTT assay, a colony-formation assay, flow cytometry and a cell motility assay, we found that the suppression of SK4 channels significantly inhibited cell proliferation and migration and promoted apoptosis in MDA-MB-231 cells (p<0.05. Further investigation revealed that treatment with epidermal growth factor (EGF/basic fibroblast growth factor (bFGF caused MDA-MB-231 cells to undergo the epithelial-mesenchymal transition (EMT and to show increased SK4 mRNA expression. In addition, the down-regulation of SK4 expression inhibited the EMT markers Vimentin and Snail1. Collectively, our findings suggest that SK4 channels are expressed in TNBC and are involved in the proliferation, apoptosis, migration and EMT processes of TNBC cells.

  12. MicroRNA-26a suppresses epithelial-mesenchymal transition in human hepatocellular carcinoma by repressing enhancer of zeste homolog 2

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    De-Ning Ma

    2016-01-01

    Full Text Available Abstract Background Our previous study reported that microRNA-26a (miR-26a inhibited tumor progression by inhibiting tumor angiogenesis and intratumoral macrophage infiltration in hepatocellular carcinoma (HCC. The direct roles of miR-26a on tumor cell invasion remain poorly understood. In this study, we aim to explore the mechanism of miR-26a in modulating epithelial-mesenchymal transition (EMT in HCC. Methods In vitro cell morphology and cell migration were compared between the hepatoma cell lines HCCLM3 and HepG2, which were established in the previous study. Overexpression and down-regulation of miR-26a were induced in these cell lines, and Western blot and immunofluorescence assays were used to detect the expression of EMT markers. Xenograft nude mouse models were used to observe tumor growth and pulmonary metastasis. Immunohistochemical assays were conducted to study the relationships between miR-26a expression and enhancer of zeste homolog 2 (EZH2 and E-cadherin expression in human HCC samples. Results Down-regulation of miR-26a in HCCLM3 and HepG2 cells resulted in an EMT-like cell morphology and high motility in vitro and increased in tumor growth and pulmonary metastasis in vivo. Through down-regulation of EZH2 expression and up-regulation of E-cadherin expression, miR-26a inhibited the EMT process in vitro and in vivo. Luciferase reporter assay showed that miR-26a directly interacted with EZH2 messenger RNA (mRNA. Furthermore, the expression of miR-26a was positively correlated with E-cadherin expression and inversely correlated with EZH2 expression in human HCC tissue. Conclusions miR-26a inhibited the EMT process in HCC by down-regulating EZH2 expression.

  13. Discoidin domain receptor 2 (DDR2) promotes breast cancer cell metastasis and the mechanism implicates epithelial-mesenchymal transition programme under hypoxia.

    Science.gov (United States)

    Ren, Tingting; Zhang, Wei; Liu, Xinping; Zhao, Hu; Zhang, Jian; Zhang, Jing; Li, Xia; Zhang, Yan; Bu, Xin; Shi, Man; Yao, Libo; Su, Jin

    2014-12-01

    A wide range of genes involved in breast cancer metastasis have been reported to be related to the microenvironment. We studied the role of discoidin domain receptor 2 (DDR2), a collagen-binding receptor, in breast cancer progression under hypoxic conditions. We showed that DDR2 protein expression closely correlated with the expression of hypoxic marker HIF-1α in clinical breast cancer specimens. The in vitro data demonstrated that hypoxia treatment increased the levels of both expression and phosphorylation of DDR2 in human breast cancer cell lines. In vivo, orthotopic breast tumour xenografts with DDR2 knockdown displayed reduced dissemination and significant prevention in pulmonary and lymphatic metastasis; conversely, these processes were significantly facilitated by the enforced expression of the activated form of DDR2. Further mechanism studies indicated that DDR2 plays an indispensable role in a series of hypoxia-induced behaviours of breast cancer cells, including migration, invasion, and epithelial-mesenchymal transition (EMT). The transcription factor Snail was found to mediate DDR2-induced down-regulation of the cell-cell adhesion molecule E-cadherin. It was also documented that there is a correlation between DDR2 and E-cadherin expression with the presence of lymph node metastases in 160 cases of invasive human breast carcinoma. In addition, we provided evidence that DDR2 silencing in breast cancer cells prevents the hypoxia-induced activation of ERK MAPK, suggesting its potential involvement in mediating the effect of DDR2 on hypoxia-induced signalling. Based on the results of this study, we conclude that DDR2 participates in hypoxia-induced breast cancer metastasis through the regulation of cell migration, invasion, and EMT, and thus may serve as an accessible therapeutic target for the treatment of breast cancer.

  14. Epithelial-mesenchymal transition involved in pulmonary fibrosis induced by multi-walled carbon nanotubes via TGF-beta/Smad signaling pathway.

    Science.gov (United States)

    Chen, Tian; Nie, Haiyu; Gao, Xin; Yang, Jinglin; Pu, Ji; Chen, Zhangjian; Cui, Xiaoxing; Wang, Yun; Wang, Haifang; Jia, Guang

    2014-04-21

    Multi-walled carbon nanotubes (MWCNT) are a typical nanomaterial with a wide spectrum of commercial applications. Inhalation exposure to MWCNT has been linked with lung fibrosis and mesothelioma-like lesions commonly seen with asbestos. In this study, we examined the pulmonary fibrosis response to different length of MWCNT including short MWCNT (S-MWCNT, length=350-700nm) and long MWCNT (L-MWCNT, length=5-15μm) and investigated whether the epithelial-mesenchymal transition (EMT) occurred during MWCNT-induced pulmonary fibrosis. C57Bl/6J male mice were intratracheally instilled with S-MWCNT or L-WCNT by a single dose of 60μg per mouse, and the progress of pulmonary fibrosis was evaluated at 7, 28 and 56 days post-exposure. The in vivo data showed that only L-MWCNT increased collagen deposition and pulmonary fibrosis significantly, and approximately 20% of pro-surfactant protein-C positive epithelial cells transdifferentiated to fibroblasts at 56 days, suggesting the occurrence of EMT. In order to understand the mechanism, we used human pulmonary epithelial cell line A549 to investigate the role of TGF-β/p-Smad2 signaling pathway in EMT. Our results showed that L-MWCNT downregulated E-cadherin and upregulated α-smooth muscle actin (α-SMA) protein expression in A549 cells. Taken together, both in vivo and in vitro study demonstrated that respiratory exposure to MWCNT induced length dependent pulmonary fibrosis and epithelial-derived fibroblasts via TGF-β/Smad pathway.

  15. Allelic Variants in Arhgef11 via the Rho-Rock Pathway Are Linked to Epithelial-Mesenchymal Transition and Contributes to Kidney Injury in the Dahl Salt-Sensitive Rat.

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    Zhen Jia

    Full Text Available Previously, genetic analyses identified that variants in Arhgef11 may influence kidney injury in the Dahl salt-sensitive (S rat, a model of hypertensive chronic kidney disease. To understand the potential mechanism by which altered expression and/or protein differences in Arhgef11 could play a role in kidney injury, stably transduced Arhgef11 knockdown cell lines as well as primary cultures of proximal tubule cells were studied. Genetic knockdown of Arhgef11 in HEK293 and NRK resulted in reduced RhoA activity, decreased activation of Rho-ROCK pathway, and less stress fiber formation versus control, similar to what was observed by pharmacological inhibition (fasudil. Primary proximal tubule cells (PTC cultured from the S exhibited increased expression of Arhgef11, increased RhoA activity, and up regulation of Rho-ROCK signaling compared to control (small congenic. The cells were also more prone (versus control to TGFβ-1 induced epithelial-mesenchymal transition (EMT, a hallmark feature of the development of renal interstitial fibrosis, and characterized by development of spindle shape morphology, gene expression changes in EMT markers (Col1a3, Mmp9, Bmp7, and Ocln and increased expression of N-Cadherin and Vimentin. S derived PTC demonstrated a decreased ability to uptake FITC-albumin compared to the small congenic in vitro, which was confirmed by assessment of albumin re-uptake in vivo by infusion of FITC-albumin and immunofluorescence imaging. In summary, these studies suggest that genetic variants in the S form of Arhgef11 via increased expression and/or protein activity play a role in promoting kidney injury in the S rat through changes in cell morphology (Rho-Rock and/or EMT that impact the function of tubule cells.

  16. The Genetics and Biophysics of the Epithelial-Mesenchymal Transition (EMT): Can Theoretical Physics Help Cancer Biology

    Science.gov (United States)

    Levine, Herbert

    In order to spread from the primary tumor to distant sites, cancer cells must undergo a coordinated change in their phenotypic properties referred to as the ''epithelial-to-mesenchymal'' transition. We have studied the nonlinear genetic circuits that are responsible for this cellular decision-making progress and propose that the transition actually goes through a series of intermediate states. At the same time, we have formulated motility models which allow for the correlation of the state of this network and the cell's biophysical capabilities. Hopefully, these thereby efforts will help us better understand the transition to metastatic disease and possible treatments thereof.

  17. Development of a Novel Method to Detect Prostate Cancer Circulating Tumor Cells (CTCs) Based on Epithelial-Mesenchymal Transition Biology

    Science.gov (United States)

    2015-12-01

    during systemic therapies. Individual CTC profiling offers the ability to reconstruct complex evolution- ary trees of tumor molecular changes from the...of these methods is the need for adequate internal controls including matched leukocytes to determine the somatic nature of the genomic changes versus... Polyak , K., Brisken, C., Yang, J., & Weinberg, R. A. (2008). The epithelial–mesenchymal transition generates cells with properties of stem cells. Cell

  18. Investigating the link between molecular subtypes of glioblastoma, epithelial-mesenchymal transition, and CD133 cell surface protein.

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    Hadi Zarkoob

    Full Text Available In this manuscript, we use genetic data to provide a three-faceted analysis on the links between molecular subclasses of glioblastoma, epithelial-to-mesenchymal transition (EMT and CD133 cell surface protein. The contribution of this paper is three-fold: First, we use a newly identified signature for epithelial-to-mesenchymal transition in human mammary epithelial cells, and demonstrate that genes in this signature have significant overlap with genes differentially expressed in all known GBM subtypes. However, the overlap between genes up regulated in the mesenchymal subtype of GBM and in the EMT signature was more significant than other GBM subtypes. Second, we provide evidence that there is a negative correlation between the genetic signature of EMT and that of CD133 cell surface protein, a putative marker for neural stem cells. Third, we study the correlation between GBM molecular subtypes and the genetic signature of CD133 cell surface protein. We demonstrate that the mesenchymal and neural subtypes of GBM have the strongest correlations with the CD133 genetic signature. While the mesenchymal subtype of GBM displays similarity with the signatures of both EMT and CD133, it also exhibits some differences with each of these signatures that are partly due to the fact that the signatures of EMT and CD133 are inversely related to each other. Taken together these data shed light on the role of the mesenchymal transition and neural stem cells, and their mutual interaction, in molecular subtypes of glioblastoma multiforme.

  19. Phosphoglucose isomerase/autocrine motility factor mediates epithelial-mesenchymal transition regulated by miR-200 in breast cancer cells.

    Science.gov (United States)

    Ahmad, Aamir; Aboukameel, Amro; Kong, Dejuan; Wang, Zhiwei; Sethi, Seema; Chen, Wei; Sarkar, Fazlul H; Raz, Avraham

    2011-05-01

    Phosphoglucose isomerase/autocrine motility factor (PGI/AMF) plays an important role in glycolysis and gluconeogenesis and is associated with invasion and metastasis of cancer cells. We have previously shown its role in the induction of epithelial-mesenchymal transition (EMT) in breast cancer cells, which led to increased aggressiveness; however, the molecular mechanism by which PGI/AMF regulates EMT is not known. Here we show, for the first time, that PGI/AMF overexpression led to an increase in the DNA-binding activity of NF-κB, which, in turn, led to increased expression of ZEB1/ZEB2. The microRNA-200s (miR-200s) miR-200a, miR-200b, and miR-200c are known to negatively regulate the expression of ZEB1/ZEB2, and we found that the expression of miR-200s was lost in PGI/AMF overexpressing MCF-10A cells and in highly invasive MDA-MB-231 cells, which was consistent with increased expression of ZEB1/ZEB2. Moreover, silencing of PGI/AMF expression in MDA-MB-231 cells led to overexpression of miR-200s, which was associated with reversal of EMT phenotype (i.e., mesenchymal-epithelial transition), and these findings were consistent with alterations in the relative expression of epithelial (E-cadherin) and mesenchymal (vimentin, ZEB1, ZEB2) markers and decreased aggressiveness as judged by clonogenic, motility, and invasion assays. Moreover, either reexpression of miR-200 or silencing of PGI/AMF suppressed pulmonary metastases of MDA-MB-231 cells in vivo, and anti-miR-200 treatment in vivo resulted in increased metastases. Collectively, these results suggest a role of miR-200s in PGI/AMF-induced EMT and thus approaches for upregulation of miR-200s could be a novel therapeutic strategy for the treatment of highly invasive breast cancer.

  20. N1-guanyl-1,7-diaminoheptane sensitizes bladder cancer cells to doxorubicin by preventing epithelial-mesenchymal transition through inhibition of eukaryotic translation initiation factor 5A2 activation.

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    Yang, Jinsong; Yu, Haogang; Shen, Mo; Wei, Wei; Xia, Lihong; Zhao, Peng

    2014-02-01

    Drug resistance greatly reduces the efficacy of doxorubicin-based chemotherapy in bladder cancer treatment; however, the underlying mechanisms are poorly understood. We aimed to investigate whether N1-guanyl-1,7-diaminoheptane (GC7), which inhibits eukaryotic translation initiation factor 5A2 (eIF5A2) activation, exerts synergistic cytotoxicity with doxorubicin in bladder cancer, and whether eIF5A2 is involved in chemoresistance to doxorubicin-based bladder cancer treatment. BIU-87, J82, and UM-UC-3 bladder cancer cells were transfected with eIF5A2 siRNA or negative control siRNA before incubation with doxorubicin alone or doxorubicin plus GC7 for 48 h. Doxorubicin cytotoxicity was enhanced by GC7 in BIU-87, J82, and UM-UC-3 cells. It significantly inhibited activity of eIF5A2, suppressed doxorubicin-induced epithelial-mesenchymal transition in BIU-87 cells, and promoted mesenchymal-epithelial transition in J82 and UM-UC-3 cells. Knockdown of eIF5A2 sensitized bladder cancer cells to doxorubicin, prevented doxorubicin-induced EMT in BIU-87 cells, and encouraged mesenchymal-epithelial transition in J82 and UM-UC-3 cells. Combination therapy with GC7 may enhance the therapeutic efficacy of doxorubicin in bladder cancer by inhibiting eIF5A2 activation and preventing epithelial-mesenchymal transition.

  1. Increase of α-SMA(+) and CK (+) cells as an early sign of epithelial-mesenchymal transition during colorectal carcinogenesis.

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    Valcz, Gábor; Sipos, Ferenc; Krenács, Tibor; Molnár, Jeannette; Patai, Arpád V; Leiszter, Katalin; Tóth, Kinga; Wichmann, Barna; Molnár, Béla; Tulassay, Zsolt

    2012-04-01

    Our aim was to examine cell transition events by detecting the frequency of intrapithelial α-smooth muscle actin (SMA)(+)/cytokeratin (CK)(+) cells during colorectal adenoma-carcinoma sequence, in relation to E-cadherin expression. Our further aim was to determine the proliferative activity of intraepithelial α-SMA(+) cells. Histologically healthy, adenoma, and colorectal cancer (CRC) biopsy samples were taken during routine colonoscopy and were included into tissue microarrays (TMAs). Slides immunostained for Ki-67, α-SMA, E-cadherin and pan-cytokeratin were digitalized and analyzed by using a digital microscope software. The proportion of α-SMA(+)/CK(+) cells was significantly higher in CRC samples (3.34 ± 1.01%) compared to healthy (1.94 ± 0.69%) or adenoma (1.62 ± 0.78%) samples (p SMA(+) cells. The majority of intraepithelial α-SMA(+) cells were in the proliferative phase. During tumor progression, the appearance of dot-like α-SMA staining in CK positive cells may indicate the initial phase of the epithelial-to-mesenchymal transition (EMT). The high proportion of intraepithelial α-SMA(+) proliferating cells may refer to their increased plasticity compared to differentiated cells. The negative correlation between E-cadherin and intraepithelial α-SMA expression suggests that EMT is facilitated by a loss of epithelial cell contact.

  2. Effect of Rapamycin on TGF-β1-induced epithelial-mesenchymal transition in LoVo colonic adenocarcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Renhu Sun; Jiang Li; Jing Cui; Qing Lv; Xinghua Liu; Guobin Wang

    2009-01-01

    Objective:To investigate the effect of Rapamycin on epithelial-mesenchyrnal transition(EMT) of LoVo colonic adenocarcinoma cells in vitro.Methods:Cultured LoVo colonic adenocarcinoma cells were divided into three groups: negative control group,EMT-inducing group(TGF-β1) and EMT-interfering group(TGF-β1 plus Rapamycin).E-cadherin expression in LoVo cells was detected by Western Blot,while the expression of vimentin was evaluated through immunocytochemistry.The Snail mRNA in LoVo cells was examined by RT-PCR.Results:TGF-β1 induced LoVo cell switching from polygonal to spindle-shaped.TGF-β1 enhanced the expression of vimentin,but lowered the level of E-cadherin.In contrast,Rapamycin impaired the transition induced by TGF-β1.Rapamycin dramatically abrogated TGF-β1-induced vimentin expression and restored E-cadherin expression in LoVo cells.Rapamycin significantly repressed the up-regulation of Snail mRNA expression induced by TGF-β1.Conclusion:Rapamycin dramatically abrogated TGF-β1 induced Snail mRNA expression in LoVo cells,hence inhibiting EMT of these cells in vitro.

  3. MicroRNA-503 represses epithelial-mesenchymal transition and inhibits metastasis of osteosarcoma by targeting c-myb.

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    Guo, Xinzhen; Zhang, Jie; Pang, Jianfeng; He, Sheng; Li, Guojun; Chong, Yang; Li, Chao; Jiao, Zhijian; Zhang, Shiqian; Shao, Ming

    2016-07-01

    Deregulated expression of miRNAs contributes to the development of osteosarcoma. Our previous study has showed that miR-503 was downregulated in osteosarcoma tissues. However, the mechanism of the miR-503 in osteosarcoma development still remains largely undefined. In our study, we found that miR-503 overexpression suppressed cell invasion and migration and inhibited epithelial-to-mesenchymal transition (EMT) of MG-63. Furthermore, we identified that c-myb, an oncogene, was a direct target of miR-503. Moreover, overexpression of c-myb could rescue miR-503-suppressed invasion and EMT. The expression of c-myb was upregulated in osteosarcoma cell lines. Therefore, we conclude that high miR-503 expression suppressed osteosarcoma cell mobility and EMT through targeting c-myb, and this may serve as a therapeutic target.

  4. Notch-1-mediated esophageal carcinoma EC-9706 cell invasion and metastasis by inducing epithelial-mesenchymal transition through Snail.

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    Wang, Tao; Xuan, Xiaoyan; Pian, Linping; Gao, Ping; Hu, Hong; Zheng, Yuling; Zang, Wenqiao; Zhao, Guoqiang

    2014-02-01

    Notch has recently been shown to promote epithelial-to-mesenchymal transition (EMT) by involving in the EMT process that occurs during tumor progression and converts polarized epithelial cells into motile, invasive cells. However, it is still unclear whether the Notch signaling pathway is associated with the regulation of EMT in esophageal carcinoma. The present study explored Notch-1-mediated esophageal carcinoma EC-9706 cell invasion and metastasis by inducing epithelial–mesenchymal transition through Snail. The results demonstrated that the inhibition of Notch-1 expression in the esophageal carcinoma cell line EC-9706 could suppress the occurrence of EMT and at the same time could decrease the invasion and metastasis ability of the EC-9706 cells, indicative of its role in EMT. Snail is a transcriptional repressor of E-cadherin. We found that with the inhibition of Notch-1 expression in the esophageal carcinoma cell line EC-9706, the expression of Snail also decreased. Mechanistic studies showed that the up-expression of Snail in the EC-9706 cells restored the suppression of EMT regulated by Notch-1 inhibition, suggesting the role of Snail in Notch-1-mediated EMT. At the same time, the up-expression of Snail in the EC-9706 cells could also rescue the invasion and metastasis ability inhibited by Notch-1 siRNA. Taken together, our results had revealed that Notch-1 could participate in the invasion and metastasis of esophageal carcinoma through EMT via Snail. This study indicated that Notch-1 might be a useful target for esophageal carcinoma prevention and therapy.

  5. FHOD1, a formin upregulated in epithelial-mesenchymal transition, participates in cancer cell migration and invasion.

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    Maria Gardberg

    Full Text Available Cancer cells can obtain their ability to invade and metastasise by undergoing epithelial-to-mesenchymal transition (EMT. Exploiting this mechanism of cellular plasticity, malignant cells can remodel their actin cytoskeleton and down-regulate proteins needed for cell-cell contacts. The mechanisms of cytoskeletal reorganisation resulting in mesenchymal morphology and increased invasive potential are poorly understood. Actin nucleating formins have been implicated as key players in EMT. Here, we analysed which formins are altered in squamous cell carcinoma related EMT. FHOD1, a poorly studied formin, appeared to be markedly upregulated upon EMT. In human tissues FHOD1 was primarily expressed in mesenchymal cells, with little expression in epithelia. However, specimens from oral squamous cell cancers demonstrated consistent FHOD1 upregulation in mesenchymally transformed cells at the invasive edge. This upregulation was confirmed in an oral squamous carcinoma model, where FHOD1 expression was markedly increased upon EMT in a PI3K signalling dependent manner. In the EMT cells FHOD1 contributed to the spindle-shaped morphology and mesenchymal F-actin organization. Furthermore, functional assays demonstrated that FHOD1 contributes to cell migration and invasion. Finally, FHOD1 depletion reduced the ability of EMT cancer cells to form invadopodia and to degrade extracellular matrix. Our results indicate that FHOD1 participates in cytoskeletal changes in EMT. In addition, we show that FHOD1 upregulation occurs during cancer cell EMT in vivo, which indicates that FHOD1 may contribute to tumour progression.

  6. FHOD1, a Formin Upregulated in Epithelial-Mesenchymal Transition, Participates in Cancer Cell Migration and Invasion

    Science.gov (United States)

    Gardberg, Maria; Kaipio, Katja; Lehtinen, Laura; Mikkonen, Piia; Heuser, Vanina D.; Talvinen, Kati; Iljin, Kristiina; Kampf, Caroline; Uhlen, Mathias; Grénman, Reidar; Koivisto, Mari; Carpén, Olli

    2013-01-01

    Cancer cells can obtain their ability to invade and metastasise by undergoing epithelial-to-mesenchymal transition (EMT). Exploiting this mechanism of cellular plasticity, malignant cells can remodel their actin cytoskeleton and down-regulate proteins needed for cell-cell contacts. The mechanisms of cytoskeletal reorganisation resulting in mesenchymal morphology and increased invasive potential are poorly understood. Actin nucleating formins have been implicated as key players in EMT. Here, we analysed which formins are altered in squamous cell carcinoma related EMT. FHOD1, a poorly studied formin, appeared to be markedly upregulated upon EMT. In human tissues FHOD1 was primarily expressed in mesenchymal cells, with little expression in epithelia. However, specimens from oral squamous cell cancers demonstrated consistent FHOD1 upregulation in mesenchymally transformed cells at the invasive edge. This upregulation was confirmed in an oral squamous carcinoma model, where FHOD1 expression was markedly increased upon EMT in a PI3K signalling dependent manner. In the EMT cells FHOD1 contributed to the spindle-shaped morphology and mesenchymal F-actin organization. Furthermore, functional assays demonstrated that FHOD1 contributes to cell migration and invasion. Finally, FHOD1 depletion reduced the ability of EMT cancer cells to form invadopodia and to degrade extracellular matrix. Our results indicate that FHOD1 participates in cytoskeletal changes in EMT. In addition, we show that FHOD1 upregulation occurs during cancer cell EMT in vivo, which indicates that FHOD1 may contribute to tumour progression. PMID:24086398

  7. Integrin β4 promotes cell invasion and epithelial-mesenchymal transition through the modulation of Slug expression in hepatocellular carcinoma

    Science.gov (United States)

    Li, Xiao-Long; Liu, Lin; Li, Dan-Dan; He, Ya-Ping; Guo, Le-Hang; Sun, Li-Ping; Liu, Lin-Na; Xu, Hui-Xiong; Zhang, Xiao-Ping

    2017-01-01

    Integrin β4 (ITGB4) is a transmembrane receptor involved in tumorigenesis and the invasiveness of many cancers. However, its role in hepatocellular carcinoma (HCC), one of the most prevalent human cancers worldwide, remains unclear. Here, we examined the involvement of ITGB4 in HCC and explored the underlying mechanisms. Real-time PCR and immunohistochemical analyses of tissues from 82 patients with HCC and four HCC cell lines showed higher ITGB4 levels in tumor than in adjacent non-tumor tissues and in HCC than in normal hepatic cells. Silencing of ITGB4 repressed cell proliferation, colony forming ability and cell invasiveness, whereas ectopic expression of ITGB4 promoted the proliferation and invasion of HCC cells and induced epithelial to mesenchymal transition (EMT) in parallel with the upregulation of Slug, as shown by transwell assays, WB and immunocytochemistry. Knockdown of Slug reduced cell viability inhibited invasion and reversed the effects of ITBG4 overexpression on promoting EMT, and AKT/Sox2-Nanog may also be involved. In a xenograft tumor model induced by injection of ITGB4-overexpressing cells into nude mice, ITGB4 promoted tumor growth and metastasis to the lungs. Taken together, our results indicate that ITGB4 plays a tumorigenic and pro-metastatic role mediated by Slug and suggest IGTB4 could be a prognostic indicator or a therapeutic target in patients with HCC. PMID:28084395

  8. NEDD9 is a positive regulator of epithelial-mesenchymal transition and promotes invasion in aggressive breast cancer.

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    Chenfei Kong

    Full Text Available Epithelial to mesenchymal transition (EMT plays an important role in many biological processes. The latest studies revealed that aggressive breast cancer, especially the triple-negative breast cancer (TNBC subtype was frequently associated with apparent EMT, but the mechanisms are still unclear. NEDD9/HEF1/Cas-L is a member of the Cas protein family and was identified as a metastasis marker in multiple cancer types. In this study, we wished to discern the role of NEDD9 in breast cancer progression and to investigate the molecular mechanism by which NEDD9 regulates EMT and promotes invasion in triple-negative breast cancer. We showed that expression of NEDD9 was frequently upregulated in TNBC cell lines, and in aggressive breast tumors, especially in TNBC subtype. Knockdown of endogenous NEDD9 reduced the migration, invasion and proliferation of TNBC cells. Moreover, ectopic overexpression of NEDD9 in mammary epithelial cells led to a string of events including the trigger of EMT, activation of ERK signaling, increase of several EMT-inducing transcription factors and promotion of their interactions with the E-cadherin promoter. Data presented in this report contribute to the understanding of the mechanisms by which NEDD9 promotes EMT, and provide useful clues to the evaluation of the potential of NEDD9 as a responsive molecular target for TNBC chemotherapy.

  9. Canine Mammary Cancer Stem Cells are Radio- and Chemo- Resistant and Exhibit an Epithelial-Mesenchymal Transition Phenotype.

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    Pang, Lisa Y; Cervantes-Arias, Alejandro; Else, Rod W; Argyle, David J

    2011-03-30

    Canine mammary carcinoma is the most common cancer among female dogs and is often fatal due to the development of distant metastases. In humans, solid tumors are made up of heterogeneous cell populations, which perform different roles in the tumor economy. A small subset of tumor cells can hold or acquire stem cell characteristics, enabling them to drive tumor growth, recurrence and metastasis. In veterinary medicine, the molecular drivers of canine mammary carcinoma are as yet undefined. Here we report that putative cancer stem cells (CSCs) can be isolated form a canine mammary carcinoma cell line, REM134. We show that these cells have an increased ability to form tumorspheres, a characteristic of stem cells, and that they express embryonic stem cell markers associated with pluripotency. Moreover, canine CSCs are relatively resistant to the cytotoxic effects of common chemotherapeutic drugs and ionizing radiation, indicating that failure of clinical therapy to eradicate canine mammary cancer may be due to the survival of CSCs. The epithelial to mesenchymal transition (EMT) has been associated with cancer invasion, metastasis, and the acquisition of stem cell characteristics. Our results show that canine CSCs predominantly express mesenchymal markers and are more invasive than parental cells, indicating that these cells have a mesenchymal phenotype. Furthermore, we show that canine mammary cancer cells can be induced to undergo EMT by TGFβ and that these cells have an increased ability to form tumorspheres. Our findings indicate that EMT induction can enrich for cells with CSC properties, and provide further insight into canine CSC biology.

  10. Canine Mammary Cancer Stem Cells are Radio- and Chemo-Resistant and Exhibit an Epithelial-Mesenchymal Transition Phenotype

    Energy Technology Data Exchange (ETDEWEB)

    Pang, Lisa Y., E-mail: lisa.pang@ed.ac.uk; Cervantes-Arias, Alejandro; Else, Rod W.; Argyle, David J. [Royal (Dick) School of Veterinary Studies and Roslin Institute, The University of Edinburgh, Easter Bush, Midlothian, EH25 9RG (United Kingdom)

    2011-03-30

    Canine mammary carcinoma is the most common cancer among female dogs and is often fatal due to the development of distant metastases. In humans, solid tumors are made up of heterogeneous cell populations, which perform different roles in the tumor economy. A small subset of tumor cells can hold or acquire stem cell characteristics, enabling them to drive tumor growth, recurrence and metastasis. In veterinary medicine, the molecular drivers of canine mammary carcinoma are as yet undefined. Here we report that putative cancer stem cells (CSCs) can be isolated form a canine mammary carcinoma cell line, REM134. We show that these cells have an increased ability to form tumorspheres, a characteristic of stem cells, and that they express embryonic stem cell markers associated with pluripotency. Moreover, canine CSCs are relatively resistant to the cytotoxic effects of common chemotherapeutic drugs and ionizing radiation, indicating that failure of clinical therapy to eradicate canine mammary cancer may be due to the survival of CSCs. The epithelial to mesenchymal transition (EMT) has been associated with cancer invasion, metastasis, and the acquisition of stem cell characteristics. Our results show that canine CSCs predominantly express mesenchymal markers and are more invasive than parental cells, indicating that these cells have a mesenchymal phenotype. Furthermore, we show that canine mammary cancer cells can be induced to undergo EMT by TGFβ and that these cells have an increased ability to form tumorspheres. Our findings indicate that EMT induction can enrich for cells with CSC properties, and provide further insight into canine CSC biology.

  11. Treatment of Human Placental Choriocarcinoma Cells with Formaldehyde and Benzene Induced Growth and Epithelial Mesenchymal Transition via Induction of an Antioxidant Effect

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    Hae-Miru Lee

    2017-07-01

    Full Text Available Cigarette smoke (CS causes about 480,000 deaths each year worldwide, and it is well-known to have harmful effects on the human body, leading to heart disease, stroke, lung cancer, and cardiovascular problems. In this study, the effects of formaldehyde (FA and benzene (Bz, the main components of CS, on cell proliferation and epithelial mesenchymal transition (EMT of JEG-3 human choriocarcinoma cells were examined to confirm the relationship between CS components and placenta carcinoma. Upon MTT assay, FA (10−8 M to 10−5 M and Bz (10−11 M to 10−8 M increased JEG-3 cell proliferation. Western blot assay revealed that the protein expression of cyclin D1 and E1 increased, while the levels of p21 and p27 were reduced following treatment. In Scratch assay, FA (10−8 M and 10−5 M and Bz (10−11 M and 10−8 M increased migration of JEG-3 cells at 24 h and 48 h compared with that at 0 h. In addition, the expression of the epithelial marker, E-cadherin, was significantly decreased, while the expression of the mesenchymal marker, N-cadherin, was significantly increased by FA (10−8 M and 10−5 M and Bz (10−11 M and 10−8 M. snail and slug transcriptional factors were associated with EMT, which were also up-regulated by FA and Bz, indicating that FA and Bz lead to an increase in the EMT process in JEG-3 choriocarcinoma cells. We further evaluated reactive oxygen species (ROS and activation of antioxidant effect using dichlorofluorescin diacetate (DCFH-DA and Western blot assay. FA and Bz increased the ROS production and an antioxidant related marker, Nrf2, in JEG-3 cells. However, eIF2α levels were reduced by FA and Bz via activation of the antioxidant reaction. Taken together, these results indicated that FA and Bz induce the growth and migration of human choriocarcinoma cells via regulation of the cell cycle and EMT and activation of ROS and antioxidant related markers.

  12. Inhibitor of growth 4 suppresses colorectal cancer growth and invasion by inducing G1 arrest, inhibiting tumor angiogenesis and reversing epithelial-mesenchymal transition.

    Science.gov (United States)

    Qu, Hui; Yin, Hong; Yan, Su; Tao, Min; Xie, Yufeng; Chen, Weichang

    2016-05-01

    Snail1 epithelial-mesenchymal transition (EMT)-inducing transcription factor (EMT-TF).

  13. Paraquat induces epithelial-mesenchymal transition-like cellular response resulting in fibrogenesis and the prevention of apoptosis in human pulmonary epithelial cells.

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    Atsushi Yamada

    Full Text Available The aim of this study is to investigate the molecular mechanisms underlying delayed progressive pulmonary fibrosis, a characteristic of subacute paraquat (PQ poisoning. Epithelial-mesenchymal transition (EMT has been proposed as a cause of organ fibrosis, and transforming growth factor-β (TGF-β is suggested to be a powerful mediator of EMT. We thus examined the possibility that EMT is involved in pulmonary fibrosis during PQ poisoning using A549 human alveolar epithelial cells in vitro. The cells were treated with various concentrations of PQ (0-500 μM for 2-12 days. Short-term (2 days high-dose (>100 μM treatments with PQ induced cell death accompanied by the activation of caspase9 as well as a decrease in E-cadherin (an epithelial cell marker, suggesting apoptotic cell death with the features of anoikis (cell death due to the loss of cell-cell adhesion. In contrast, long-term (6-12 days low-dose (30 μM treatments with PQ resulted in a transformation into spindle-shaped mesenchymal-like cells with a decrease of E-cadherin as well as an increase of α-smooth muscle actin (α-SMA. The mesenchymal-like cells also secreted the extracellular matrix (ECM protein fibronectin into the culture medium. The administration of a TGF-β1 receptor antagonist, SB431542, almost completely attenuated the mesenchymal transformation as well as fibronectin secretion, suggesting a crucial role of TGF-β1 in EMT-like cellular response and subsequent fibrogenesis. It is noteworthy that despite the suppression of EMT-fibrogenesis, apoptotic death was observed in cells treated with PQ+SB431542. EMT-like cellular response and subsequent fibrogenesis were also observed in normal human bronchial epithelial (NHBE cells exposed to PQ in a TGF-β1-dependent manner. Taken together, our experimental model reflects well the etiology of PQ poisoning in human and shows the involvement of EMT-like cellular response in both fibrogenesis and resistance to cell death during

  14. Autocrine/Paracrine Human Growth Hormone-stimulated MicroRNA 96-182-183 Cluster Promotes Epithelial-Mesenchymal Transition and Invasion in Breast Cancer.

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    Zhang, Weijie; Qian, Pengxu; Zhang, Xiao; Zhang, Min; Wang, Hong; Wu, Mingming; Kong, Xiangjun; Tan, Sheng; Ding, Keshuo; Perry, Jo K; Wu, Zhengsheng; Cao, Yuan; Lobie, Peter E; Zhu, Tao

    2015-05-29

    Human growth hormone (hGH) plays critical roles in pubertal mammary gland growth, development, and sexual maturation. Accumulated studies have reported that autocrine/paracrine hGH is an orthotopically expressed oncoprotein that promotes normal mammary epithelial cell oncogenic transformation. Autocrine/paracrine hGH has also been reported to promote mammary epithelial cell epithelial-mesenchymal transition (EMT) and invasion. However, the underlying mechanism remains largely obscure. MicroRNAs (miRNAs) are reported to be involved in regulation of multiple cellular functions of cancer. To determine whether autocrine/paracrine hGH promotes EMT and invasion through modulation of miRNA expression, we performed microarray profiling using MCF-7 cells stably expressing wild type or a translation-deficient hGH gene and identified miR-96-182-183 as an autocrine/paracrine hGH-regulated miRNA cluster. Forced expression of miR-96-182-183 conferred on epithelioid MCF-7 cells a mesenchymal phenotype and promoted invasive behavior in vitro and dissemination in vivo. Moreover, we observed that miR-96-182-183 promoted EMT and invasion by directly and simultaneously suppressing BRMS1L (breast cancer metastasis suppressor 1-like) gene expression. miR-96 and miR-182 also targeted GHR, providing a potential negative feedback loop in the hGH-GHR signaling pathway. We further demonstrated that autocrine/paracrine hGH stimulated miR-96-182-183 expression and facilitated EMT and invasion via STAT3 and STAT5 signaling. Consistent with elevated expression of autocrine/paracrine hGH in metastatic breast cancer tissue, miR-96-182-183 expression was also remarkably enhanced. Hence, we delineate the roles of the miRNA-96-182-183 cluster and elucidate a novel hGH-GHR-STAT3/STAT5-miR-96-182-183-BRMS1L-ZEB1/E47-EMT/invasion axis, which provides further understanding of the mechanism of autocrine/paracrine hGH-stimulated EMT and invasion in breast cancer.

  15. Epithelial mesenchymal transition status is associated with anti-cancer responses towards receptor tyrosine-kinase inhibition by dovitinib in human bladder cancer cells.

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    Hänze, Jörg; Henrici, Marcus; Hegele, Axel; Hofmann, Rainer; Olbert, Peter J

    2013-12-11

    Dovitinib (TKI-258) is a receptor tyrosine kinase (RTK) inhibitor targeting fibroblast growth factor receptor (FGFR) and further related RTKs. TKI-258 is under investigation as anticancer drug for the treatment of various cancers including bladder cancer with aberrant RTK signaling. Here, we analyzed the responses of ten human bladder cancer cell lines towards TKI-258 treatment in relation to the epithelial mesenchymal transition (EMT) status of the cells. Expression of epithelial marker E-cadherin as well as mesenchymal markers N-cadherin and vimentin was determined by quantitative RT-PCR and Western-blot in RNA and protein extracts from the cultured cell lines. The cell responses were analyzed upon addition of TKI-258 by viability/proliferation (XTT assay) and colony formation assay for measurement of cell contact independent growth. The investigated bladder cancer cell lines turned out to display quite different EMT patterns as indicated by the abundance of E-cadherin or N-cadherin and vimentin. Protein and mRNA levels of the respective components strongly correlated. Based on E-cadherin and N-cadherin mRNA levels that were expressed approximately mutual exclusively, an EMT-score was calculated for each cell line. A high EMT-score indicated mesenchymal-like cells and a low EMT-score epithelial-like cells. Then, we determined the IC₅₀ values for TKI-258 by dose response curves (0-12 μM TKI-258) in XTT assays for each cell line. Also, we measured the clonogenic survival fraction after adding TKI-258 (1 μM) by colony formation assay. We observed significant correlations between EMT-score and IC₅₀ values (r = 0.637, p = 0.0474) and between EMT-score and clonogenic survival fraction (r = 0.635, p = 0.0483) as analyzed by linear regression analyses. In sum, we demonstrated that the EMT status based on E-cadherin and N-cadherin mRNA levels may be useful to predict responses towards TKI-258 treatment in bladder cancer.

  16. [The expression of transcription factors Snail and Slug in epithelial-mesenchymal transition of human lens epithelial cells induced by transforming growth factor-β2].

    Science.gov (United States)

    Wang, Y N; Pei, C; Qin, L; Li, J M; Yi, J L; Chen, L

    2016-04-11

    To investigate the expression of transcription factors snail and slug in epithelial mesenchymal transition (EMT) of human lens epithelial cells (HLEC) induced by transforming growth factor-β2 (TGF-β2). Experimental research. HLEC were treated with different concentrations of TGF-β2 (1.0 and 10.0 μg/L) for different time. The morphological changes were observed under inverted microscope. The expression and cellular localization of snail and slug were evaluated by immunofluorescence. Expressions of snail, slug, E-Cadherin and α-SMA were further determined by Western blot analysis. Single factor analysis of variance, rank sum test and Pearson correlation were used for statistical analysis. Cultured HLEC were polygonal monolayer cells with tight intercellular adhesion closely and patchy distribution. After treatment of different doses of TGF-β2 for 24 h, HLEC became isolated, exhibited long spindle-like shape as fibroblastic phenotype. The immunofluorescence staining indicated that snail and slug were localized in the nuclei. The expressions of snail and slug appeared to be positive correlative to TGF-β2 dose (snail protein expression: 0.74±0.16, 1.13±0.03, 1.54±0.18 and slug protein expression: 1.96±0.02, 3.12±0.09, 4.07±0.12 in HLEC treated with 0.1, 1.0 and 10 μg/L TGF-β2 respectively) (χ(2)=9.62,P=0.022;F=241.10,Psnail and slug in HLEC were also increased with extending duration of TGF-β2 (1.0 μg/L). The expression levels of both proteins were modestly up-regulated at 8 hours, robustly increased at 24 h, reached peak at 48h and began to decline at 72 h (snail protein expression: 0.90±0.13, 1.43±0.14, 1.96±0.27, 1.57±0.16 and slug protein expression: 0.91±0.36, 1.24±0.16, 2.44±0.26, 1.43±0.16 in HLEC treated with 1.0 μg/L TGF-β2 for 8 h, 24 h, 48 h and 72 h respectively) (F=12.49,P=0.001;F=14.03,Psnail and slug might be time and dose-dependently involved in in-vitro TGF-β2-induced EMT of HLEC. (Chin J Ophthalmol, 2016, 52: 285-290).

  17. Impact of p120-catenin isoforms 1A and 3A on epithelial mesenchymal transition of lung cancer cells expressing E-cadherin in different subcellular locations.

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    Yijun Zhang

    Full Text Available The epithelial mesenchymal transition (EMT is an important process in tumor development. Despite previous investigations, it remains unclear how p120-catenin (p120ctn isoforms 1A and 3A affect the EMT of tumor cells. Here we investigated expression of p120ctn, E-cadherin and vimentin in 78 human non-small cell lung cancer (NSCLC samples by immunohistochemistry and found that p120ctn membrane expression positively correlated with E-cadherin expression (P<0.001 and negatively correlated with vimentin expression and lymph node metastasis (P<0.05. Meanwhile, p120ctn cytoplasmic expression negatively correlated with E-cadherin expression (P<0.001 and positively correlated with vimentin expression and lymph node metastasis (P<0.05. Cells expressing high (H460 and SPC and low (H1299 and LK2 levels of p120ctn were screen to investigate its impact on EMT. E-cadherin was restricted to the cell membrane in H460 and H1299 cells, whereas it was expressed in the cytoplasm of SPC and LK2 cells. Ablation of endogenous p120ctn isoform 1A in cells expressing high levels of the protein resulted in decreased E-cadherin expression, increased N-cadherin, vimentin and snail expression and enhanced invasiveness in H460 cells. Meanwhile, completely opposite results were observed in SPC cells. Furthermore, transfection of in H1299 cells expressing low p120ctn levels with the p120ctn isoform 1A plasmid resulted in increased E-cadherin expression, decreased N-cadherin, vimentin and snail expression and weakened invasiveness, while LK2 cells showed completely opposite results. Both cell lines expressing low p120ctn levels and transfected with the p120ctn isoform 3A plasmid appeared to have increased E-cadherin expression, decreased N-cadherin, vimentin and snail expression and weakened invasiveness. In conclusion, in cells with membrane E-cadherin, both p120ctn isoforms 1A and 3A inhibited EMT and decreased cell invasiveness. In cells with cytoplasmic E-cadherin, p120ctn

  18. Posttranslational modification of E-cadherin by core fucosylation regulates Src activation and induces epithelial-mesenchymal transition-like process in lung cancer cells.

    Science.gov (United States)

    Shao, Kang; Chen, Zhong Yi; Gautam, Suraj; Deng, Nian Hui; Zhou, You; Wu, Xing Zhong

    2016-02-01

    E-cadherin is often dysregulated in aggressive lung cancer, the mechanism of which cannot always be explained at the level of transcription. In 66 patients with lung cancer, immunohistochemical staining demonstrated that co-localization of E-cadherin and core fucose by Lens culinaris agglutinin was significantly less extensive in tumor than in nontumor tissue. Through gain and loss of fucosylation experiments in the giant lung carcinoma cell lines 95C and 95D, our results revealed that E-cadherin core fucosylation in 95C cells overexpressing α-1, 6-fucosyltransferase (Fut8) inhibited Fut8-95C cell migration, whereas knockdown of Fut8 in 95D cells enhanced migration of short-interfering RNA-targeting Fut8 (siFut8)-95D cells. The level of active Src (phosphorylated Src [Y416]) was significantly reduced in Fut8-95C cells, but elevated in siFut8-95D cells. In protein complexes immunoprecipitated from Fut8-95C cell lysates with anti-E-cadherin, less phosphorylated Src (Y416) and more β-catenin were observed, but immunoprecipitates from siFut8-95D cells, containing less core fucosylated E-cadherin, contained an elevated level of phospho-Src Y416. In Fut8-95C cells, phosphorylation of Akt (Y315, Y326) and GSK-3β (S9) was significantly reduced, but β-catenin (S37) phosphorylation was enhanced. Expression of N-cadherin and Snail1 was also reduced in Fut8-95C cells, but significantly increased in siFut8-95D cells. Intriguingly, when Src kinase activity was inhibited by treatment of cells with PP2 and SU6656, regulation of N-cadherin, Snail1 and cell migration by E-cadherin core fucosylation was abrogated in both Fut8-95C and siFut8-95D cells. Therefore, posttranslational modification of E-cadherin by less core fucosylation recruited and activated Src, and induced an epithelial-mesenchymal transition-like process in lung cancer cells.

  19. Antagonism of miR-21 reverses epithelial-mesenchymal transition and cancer stem cell phenotype through AKT/ERK1/2 inactivation by targeting PTEN.

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    Mingli Han

    Full Text Available BACKGROUND: Accumulating evidence suggested that epithelial-mesenchymal transition (EMT and cancer stem cell (CSC characteristics, both of which contribute to tumor invasion and metastasis, are interrelated with miR-21. MiR-21 is one of the important microRNAs associated with tumor progression and metastasis, but the molecular mechanisms underlying EMT and CSC phenotype during miR-21 contributes to migration and invasion of breast cancer cells remain to be elucidated. METHODOLOGY/PRINCIPAL FINDINGS: In this study, MDA-MB-231/anti-miR-21 cells were established by transfected hsa-miR-21 antagomir into breast cancer MDA-MB-231 cells. EMT was evaluated by the changes of mesenchymal cell markers (N-cadherin, Vimentin, and alpha-SMA, epithelial cell marker (E-cadherin, as well as capacities of cell migration and invasion; CSC phenotype was measured using the changes of CSC surface markers (ALDH1 and CD44, and the capacity of sphereforming (mammospheres. We found that antagonism of miR-21 reversed EMT and CSC phenotype, accompanied with PTEN up-regulation and AKT/ERK1/2 inactivation. Interestingly, down-regulation of PTEN by siPTEN suppressed the effects of miR-21 antagomir on EMT and CSC phenotype, confirming that PTEN is a target of miR-21 in reversing EMT and CSC phenotype. The inhibitors of PI3K-AKT and ERK1/2 pathways, LY294002 and U0126, both significantly suppressed EMT and CSC phenotype, indicating that AKT and ERK1/2 pathways are required for miR-21 mediating EMT and CSC phenotype. CONCLUSIONS/SIGNIFICANCE: In conclusion, our results demonstrated that antagonism of miR-21 reverses EMT and CSC phenotype through targeting PTEN, via inactivation of AKT and ERK1/2 pathways, and showed a novel mechanism of which might relieve the malignant biological behaviors of breast cancer.

  20. Na+/H+ exchanger NHE1 regulation modulates metastatic potential and epithelial-mesenchymal transition of triple-negative breast cancer cells.

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    Amith, Schammim Ray; Wilkinson, Jodi Marie; Fliegel, Larry

    2016-04-19

    In triple-negative breast cancer (TNBC), the high recurrence rate, increased invasion and aggressive metastatic formation dictate patient survival. We previously demonstrated a critical role for the Na+/H+ exchanger isoform 1 (NHE1) in controlling metastasis of triple-negative cells. Here, we investigated the effect of changes to three regulatory loci of NHE1. Two via the Ras/Raf/ERK/p90RSK pathway: p90RSK/14-3-3 (S703A) and ERK1/2 (S766,770,771A, SSSA) and a third via a calmodulin-binding domain (K641,R643,645,647E, 1K3R4E). MDA-MB-231 cells with a mutation at the p90RSK site (S703A-NHE1) changed from a wild-type mesenchymal morphology to a smaller epithelial-like phenotype with a loss of expression of mesenchymal marker vimentin. S703A cells also had reduced metastatic potential and markedly decreased rates of migration, invasion, spheroid growth, anchorage-dependent and soft agar colony formation. Similarly, BI-D1870, a specific inhibitor of p90RSK, significantly inhibited the metastatic potential of highly invasive MDA-MB-231 and moderately invasive MDA-MB-468 TNBC cells, but was minimally effective in non-invasive Hs578T TNBC cells. In contrast, invasion and spheroid growth were unaffected in cells containing NHE1 with mutations interfering with its activation by ERK1/2 (SSSA), though rates of migration and colony formation were reduced. Cells with a constitutive activation of NHE1 via the 1K3R4E mutation exhibited higher rates of migration, invasion, and spheroid growth. Taken together, our data demonstrate the critical role of NHE1 in metastasis, and suggest a novel link between NHE1 and the expression and cytosolic organization of vimentin, a key factor in epithelial-mesenchymal transition, that is dependent on p90RSK/14-3-3-mediated activation of the exchanger.

  1. Epithelial-mesenchymal transition delayed by E-cad to promote tissue formation in hepatic differentiation of mouse embryonic stem cells in vitro.

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    Hu, Anbin; Shang, Changzhen; Li, Qiang; Sun, Nianfeng; Wu, Linwei; Ma, Yi; Jiao, Xingyuan; Min, Jun; Zeng, Gucheng; He, Xiaoshun

    2014-04-15

    Hepatic differentiation of embryonic stem cells (ESCs) usually results in a single cell lineage, and the formation of liver tissues remains difficult. Here, we examine the role of epithelial-mesenchymal transition (EMT) that is regulated by epithelial cadherin (E-cad) expression in hepatic tissue formation from ESCs. E-cad was transfected into mouse ESCs to enable a stable expression of E-cad. Hepatic differentiation of ESCs was then induced by hepatic growth factors. Wnt/β-catenin signaling and EMT speed were examined to determine the differentiation process. Hepatic and angiogenesis markers, as well as differentiated cell-adhesive force were also examined to identify the hepatic tissue differentiation. In our results, E-cad expression gradually decreased in normal ESC (N-ESC) differentiation, but remained stable in the E-cad transfected ESC (EC-ESC) group. In EC-ESC differentiation, expressions of cytoplastic β-catenin and EMT were much lower and significantly prolonged. Angiogenesis markers vascular endothelial growth factor receptor-1 (VEGFR-1) and CD31/PECAM-1 were expressed only on day 5-13 in N-ESC differentiation, whereas VEGFR-1 and CD31/PECAM-1 were expressed prolonged on day 5-17 in the EC-ESC group and were coincident with the expression of hepatic markers. Finally, EC-ESC differentiation maintained multilayer-growth patterns, and abundant vascular network structures appeared and migrated in albumin-positive cell areas. The cellular adhesion forces between embryonic body cells in EC-ESC differentiation during day 13-17 were similar to those of mouse liver tissue. In conclusion, accelerated EMT due to the decreased E-cad expression may partially contribute to the failure of hepatic tissue formation in N-ESC differentiation. E-cad can act in synergy with hepatic growth factors and facilitate the early-stage formation of hepatic tissues through down-regulating Wnt/β-catenin signaling and delaying EMT. This work provides a new insight into hepatic tissue

  2. Mechanism of c-Met and EGFR tyrosine kinase inhibitor resistance through epithelial mesenchymal transition in non-small cell lung cancer

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    Rastogi, Ichwaku; Rajanna, Supriya; Webb, Andrew; Chhabra, Gagan; Foster, Brad [Department of Biomedical Sciences, University of Illinois College of Medicine at Rockford, Illinois (United States); Webb, Brian [Thermo Fisher Scientific, Rockford, Illinois (United States); Puri, Neelu, E-mail: neelupur@uic.edu [Department of Biomedical Sciences, University of Illinois College of Medicine at Rockford, Illinois (United States)

    2016-09-02

    According to currently available estimates from Cancer Research UK, 14.1 million new lung cancer cases were diagnosed and a staggering 8.2 million people worldwide died from lung cancer in 2012. EGFR and c-Met are two tyrosine kinase receptors most commonly overexpressed or mutated in Non-small Cell Lung Cancer (NSCLC) resulting in increased proliferation and survival of lung cancer cells. Tyrosine kinase inhibitors (TKIs), such as erlotinib, approved by the FDA as first/second line therapy for NSCLC patients have limited clinical efficacy due to acquired resistance. In this manuscript, we investigate and discuss the role of epithelial mesenchymal transition (EMT) in the development of resistance against EGFR and c-Met TKIs in NSCLC. Our findings show that Zeb-1, a transcriptional repressor of E-Cadherin, is upregulated in TKI-resistant cells causing EMT. We observed that TKI-resistant cells have increased gene and protein expression of EMT related proteins such as Vimentin, N-Cadherin, β-Catenin and Zeb-1, while expression of E-Cadherin, an important cell adhesion molecule, was suppressed. We also confirmed that TKI-resistant cells display mesenchymal cell type morphology, and have upregulation of β-Catenin which may regulate expression of Zeb-1, a transcriptional repressor of E-Cadherin in TKI-resistant NSCLC cells. Finally, we show that down-regulating Zeb-1 by inducing miR-200a or β-Catenin siRNA can increase drug sensitivity of TKI-resistant cells. - Highlights: • Resistance to TKIs in NSCLC cells is mediated via modulation in EMT related proteins. • EMT may induce c-Met mediated TKI resistance, similar to EGFR TKI resistance. • Role of β-catenin and cadherins in TKI resistance was validated by FACS and qPCR. • Knockdown of β-catenin or Zeb-1 can increase TKI sensitivity in TKI-resistant cells. • Targeting key EMT related proteins may overcome TKI resistance in NSCLC.

  3. Oncogenic H-Ras Reprograms Madin-Darby Canine Kidney (MDCK) Cell-derived Exosomal Proteins Following Epithelial-Mesenchymal Transition*

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    Tauro, Bow J.; Mathias, Rommel A.; Greening, David W.; Gopal, Shashi K.; Ji, Hong; Kapp, Eugene A.; Coleman, Bradley M.; Hill, Andrew F.; Kusebauch, Ulrike; Hallows, Janice L.; Shteynberg, David; Moritz, Robert L.; Zhu, Hong-Jian; Simpson, Richard J.

    2013-01-01

    Epithelial-mesenchymal transition (EMT) is a highly conserved morphogenic process defined by the loss of epithelial characteristics and the acquisition of a mesenchymal phenotype. EMT is associated with increased aggressiveness, invasiveness, and metastatic potential in carcinoma cells. To assess the contribution of extracellular vesicles following EMT, we conducted a proteomic analysis of exosomes released from Madin-Darby canine kidney (MDCK) cells, and MDCK cells transformed with oncogenic H-Ras (21D1 cells). Exosomes are 40–100 nm membranous vesicles originating from the inward budding of late endosomes and multivesicular bodies and are released from cells on fusion of multivesicular bodies with the plasma membrane. Exosomes from MDCK cells (MDCK-Exos) and 21D1 cells (21D1-Exos) were purified from cell culture media using density gradient centrifugation (OptiPrep™), and protein content identified by GeLC-MS/MS proteomic profiling. Both MDCK- and 21D1-Exos populations were morphologically similar by cryo-electron microscopy and contained stereotypical exosome marker proteins such as TSG101, Alix, and CD63. In this study we show that the expression levels of typical EMT hallmark proteins seen in whole cells correlate with those observed in MDCK- and 21D1-Exos, i.e. reduction of characteristic inhibitor of angiogenesis, thrombospondin-1, and epithelial markers E-cadherin, and EpCAM, with a concomitant up-regulation of mesenchymal makers such as vimentin. Further, we reveal that 21D1-Exos are enriched with several proteases (e.g. MMP-1, -14, -19, ADAM-10, and ADAMTS1), and integrins (e.g. ITGB1, ITGA3, and ITGA6) that have been recently implicated in regulating the tumor microenvironment to promote metastatic progression. A salient finding of this study was the unique presence of key transcriptional regulators (e.g. the master transcriptional regulator YBX1) and core splicing complex components (e.g. SF3B1, SF3B3, and SFRS1) in mesenchymal 21D1-Exos. Taken

  4. Resveratrol reverses Doxorubicin resistance by inhibiting epithelial-mesenchymal transition (EMT) through modulating PTEN/Akt signaling pathway in gastric cancer.

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    Xu, Jiahui; Liu, Deying; Niu, Huilin; Zhu, Guifang; Xu, Yangwei; Ye, Danli; Li, Jian; Zhang, Qingling

    2017-01-26

    Gastric cancer is one of the major causes of cancer-related mortality worldwide. Most of patients presenting with inoperable gastric cancers rely on systemic chemotherapy for prolongation of survival. Doxorubicin (DOX) is one of the important agents against gastric cancer. Acquired DOX-resistance severely impedes the chemotherapeutic effect, invariably leading to poor prognosis. Resveratrol (RES) as a kind of phytoalexin has demonstrated anti-tumor functions in breast cancer and myeloid leukemia, but its function and mechanism are still unknown in gastric cancer treatment. CCK8 assay was used to detect the cytotoxicity of DOX and RES to gastric cancer cells. DOX-resistant subclone cell line (SGC7901/DOX) was derived from SGC7901 cells exposed to stepwise increasing concentrations of DOX treatment. We measured the migratory capabilities of SGC7901/DOX cells by Cell scratch test and Transwell assay. SGC7901/DOX cells were treated with DOX, RES, neither or both. Then we analyzed cell survival by CCK8 assay, colony formation by Colony-forming assay, cell apoptosis by Annexin-V-FITC and PI dual staining assay and cell migration by Cell scratch test and Transwell assay. Western blotting was conducted to detect the protein expressions of PTEN/Akt signaling pathway and EMT-related markers. Immunofluorescence was performed to confirm the EMT-related markers expressions. The xenograft model was used to assess the effect of DOX and RES in vivo. The key molecules associated with proliferation, apoptosis and EMT were evaluated by immunohistochemistry in tumor specimens. SGC7901/DOX cells acquired drug resistance and enhancive migratory capability. RES enabled SGC7901/DOX cells to regain DOX sensitivity, mitigated the aggressive biological features, promoted cell apoptosis in vitro and inhibited tumor growth in vivo. Mechanistic studies revealed that SGC7901/DOX cells underwent epithelial-mesenchymal transition (EMT) which was induced by Akt activation, and through activating

  5. Functional relevance of a six mesenchymal gene signature in epithelial-mesenchymal transition (EMT) reversal by the triple angiokinase inhibitor, nintedanib (BIBF1120).

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    Huang, Ruby Yun-Ju; Kuay, Kuee Theng; Tan, Tuan Zea; Asad, Mohammad; Tang, Hei Mui; Ng, Aloysius Hsien Chun; Ye, Jieru; Chung, Vin Yee; Thiery, Jean Paul

    2015-09-08

    Epithelial-mesenchymal transition (EMT), a crucial mechanism in carcinoma progression, describes the process whereby epithelial cells lose their apico-basal polarity and junctional complexes and acquire a mesenchymal-like morphology. Several markers are considered to be authentic indicators of an epithelial or mesenchymal status; however, there is currently no comprehensive or systematic method with which to determine their functional relevance. Previously, we identified a 33-gene EMT signature comprising 25 epithelial and 6 mesenchymal genes that best describe this concept of the EMT spectrum. Here, we designed small-scale siRNA screens targeting these six mesenchymal signature genes (CD99L2, EMP3, ITGA5, SYDE1, VIM, ZEB1) to explore their functional relevance and their roles during EMT reversal by nintedanib (BIBF1120) in a mesenchymal-like SKOV3 ovarian cancer cell line. We found that neither cell proliferation nor cytotoxicity was affected by silencing any of these genes. SKOV3 cells expressing siRNA against mesenchymal genes (ZEB1, EMP3, CD99L2, ITGA5, and SYDE1) showed enhanced colony compaction (reduced inter-nuclear distance). Inductions of E-cadherin expression were only observed in SYDE1- and ZEB1-silenced SKOV3 cells. In addition, only SYDE1-silenced SKOV3 cells showed increased anoikis. Finally, we identified that SYDE1 and ZEB1 were down-regulated in nintedanib-treated SKOV3 cells and SYDE1- and ZEB1-silenced SKOV3 cells showed enhanced nintedanib-induced up-regulation of E-cadherin. Nintedanib-treated SKOV3 cells also showed colony compaction and decreases in EMT scores both in vitro and in vivo. We conclude that SYDE1 and ZEB1 are functionally relevant in EMT reversal. This study thus provides a proof-of-concept for the use of in vitro siRNA screening to explore the EMT-related functions of selected genes and their potential relevance in the discovery of EMT reversing drugs.

  6. Partially Evoked Epithelial-Mesenchymal Transition (EMT Is Associated with Increased TGFβ Signaling within Lesional Scleroderma Skin.

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    Joanna Nikitorowicz-Buniak

    Full Text Available The origin of myofibroblasts in fibrotic conditions remains unknown and in systemic sclerosis (SSc it has been proposed that activation of local fibroblasts, trans-differentiation of perivascular or vascular cells, recruitment of fibrocyte progenitors, or epithelial to mesenchymal transition (EMT could be contributing. Data from our laboratory indicate that the epidermis in scleroderma is activated with the keratinocytes exhibiting a phenotype normally associated with tissue repair, including phosphorylation profiles indicative of TGFβ signaling. Since TGFβ is a known inducer of EMT, we investigated if there is evidence of this process in the SSc epidermis. In order to validate antibodies and primers, EMT was modeled in HaCaT cells cultured in the presence of TGFβ1. Skin sections were stained with phosho-SMAD2/3, as well as with epithelial and mesenchymal markers. Moreover, mRNA levels of transcription factors associated with EMT were studied in epidermal blister sheets. We observed critical changes in the scleroderma epidermis; showing significantly increased nuclear translocation of phosphorylated Smad2/3, consistent with active TGFβ signaling in SSc keratinocytes. While profound EMT could be induced in keratinocytes in vitro with the appearance of SNAI1/2 and FSP-1, and an accompanying loss of E-cadherin, in the scleroderma skin active TGFβ signaling was accompanied by only partial EMT-like changes characterised by induction of SNAI1 alone and with no loss of E-cadherin. Together, our findings support a model of altered differentiation and TGFβ dependent activation of scleroderma epithelial cells leading to a partially evoked EMT like process in the fibrotic skin.

  7. Dioscin suppresses TGF-β1-induced epithelial-mesenchymal transition and suppresses A549 lung cancer migration and invasion.

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    Lim, Won-Chul; Kim, Hyunhee; Kim, Young-Joo; Choi, Kyung-Chul; Lee, In Ho; Lee, Ki Heon; Kim, Mi Kyung; Ko, Hyeonseok

    2017-08-01

    Epithelial-to-mesenchymal transition (EMT), an important cellular process, occurs during cancer development and progression, has a crucial role in metastasis by enhancing the motility of tumor cells. Dioscin is a polyphenolic component isolated from Phyllanthus amarus, which exhibits a wide range of pharmacological and physiological activities, such as anti-tumor, anti-inflammatory, anti-obesity, anti-fungal, and anti-viral activities. However, the possible role of dioscin in the EMT is unclear. We investigated the suppressive effect of dioscin on the EMT. Transforming growth factor-beta 1 (TGF-β1) is known to induce EMT in a number of cancer cell types and promote lung adenocarcinoma migration and invasion. To verify the inhibitory role of dioscin in lung cancer migration and invasion, we investigated the use of dioscin as inhibitors of TGF-β1-induced EMT in A549 lung cancer cells in vitro. Here, we found that dioscin prominently increased expression of the epithelial marker E-cadherin and expression of the mesenchymal marker N-cadherin and Snail during the TGF-β1-induced EMT. In addition, dioscin inhibited the TGF-β1-induced increase in cell migration and invasion of A549 lung cancer cells. Also, dioscin remarkably inhibited TGF-β1-regulated activation of MMP-2/9, Smad2, and p38. Taken together, our findings provide new evidence that dioscin suppresses lung cancer migration, and invasion in vitro by inhibiting the TGF-β1-induced EMT. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Snail Promotes Epithelial Mesenchymal Transition in Breast Cancer Cells in Part via Activation of Nuclear ERK2

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    Smith, Bethany N.; Burton, Liza J.; Henderson, Veronica; Randle, Diandra D.; Morton, Derrick J.; Smith, Basil A.; Taliaferro-Smith, Latonia; Nagappan, Peri; Yates, Clayton; Zayzafoon, Majd; Chung, Leland W. K.; Odero-Marah, Valerie A.

    2014-01-01

    Snail transcription factor is up-regulated in several cancers and associated with increased tumor migration and invasion via induction of epithelial-to-mesenchymal transition (EMT). MAPK (ERK1/2) signaling regulates cellular processes including cell motility, adhesion, and invasion. We investigated the regulation of ERK1/2 by Snail in breast cancer cells. ERK1/2 activity (p-ERK) was higher in breast cancer patient tissue as compared to normal tissue. Snail and p-ERK were increased in several breast cancer cell lines as compared to normal mammary epithelial cells. Snail knockdown in MDA-MB-231 and T47-D breast cancer cells decreased or re-localized p-ERK from the nuclear compartment to the cytoplasm. Snail overexpression in MCF-7 breast cancer cells induced EMT, increased cell migration, decreased cell adhesion and also increased tumorigenicity. Snail induced nuclear translocation of p-ERK, and the activation of its subcellular downstream effector, Elk-1. Inhibiting MAPK activity with UO126 or knockdown of ERK2 isoform with siRNA in MCF-7 Snail cells reverted EMT induced by Snail as shown by decreased Snail and vimentin expression, decreased cell migration and increased cell adhesion. Overall, our data suggest that ERK2 isoform activation by Snail in aggressive breast cancer cells leads to EMT associated with increased cell migration and decreased cell adhesion. This regulation is enhanced by positive feedback regulation of Snail by ERK2. Therefore, therapeutic targeting of ERK2 isoform may be beneficial for breast cancer. PMID:25122124

  9. Toll-like receptor 4 signaling promotes epithelial-mesenchymal transition in human hepatocellular carcinoma induced by lipopolysaccharide

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    Jing Ying-Ying

    2012-08-01

    Full Text Available Abstract Background The endotoxin level in the portal and peripheral veins of hepatocellular carcinoma (HCC patients is higher and lipopolysaccharide (LPS, a cell wall constituent of gram-negative bacteria, has been reported to inhibit tumor growth. However, in this study, we found that LPS-induced toll-like receptor 4 (TLR4 signaling was involved in tumor invasion and survival, and the molecular mechanism was investigated, Methods Four HCC cell lines and a splenic vein metastasis of the nude mouse model were used to study the invasion ability of LPS-induced HCC cells and the epithelia-mesenchymal transition (EMT in vitro and in vivo. A total of 106 clinical samples from HCC patients were used to evaluate TLR4 expression and analyze its association with clinicopathological characteristics Results The in vitro and in vivo experiments demonstrated that LPS could significantly enhance the invasive potential and induce EMT in HCC cells with TLR4 dependent. Further studies showed that LPS could directly activate nuclear factor kappa B (NF-κB signaling through TLR4 in HCC cells. Interestingly, blocking NF-κB signaling significantly inhibited transcription factor Snail expression and thereby inhibited EMT occurrence. High expression of TLR4 in HCC tissues was strongly associated with both poor cancer-free survival and overall survival in patients. Conclusions Our results indicate that TLR4 signaling is required for LPS-induced EMT, tumor cell invasion and metastasis, which provide molecular insights for LPS-related pathogenesis and a basis for developing new strategies against metastasis in HCC.

  10. Snail promotes epithelial mesenchymal transition in breast cancer cells in part via activation of nuclear ERK2.

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    Bethany N Smith

    Full Text Available Snail transcription factor is up-regulated in several cancers and associated with increased tumor migration and invasion via induction of epithelial-to-mesenchymal transition (EMT. MAPK (ERK1/2 signaling regulates cellular processes including cell motility, adhesion, and invasion. We investigated the regulation of ERK1/2 by Snail in breast cancer cells. ERK1/2 activity (p-ERK was higher in breast cancer patient tissue as compared to normal tissue. Snail and p-ERK were increased in several breast cancer cell lines as compared to normal mammary epithelial cells. Snail knockdown in MDA-MB-231 and T47-D breast cancer cells decreased or re-localized p-ERK from the nuclear compartment to the cytoplasm. Snail overexpression in MCF-7 breast cancer cells induced EMT, increased cell migration, decreased cell adhesion and also increased tumorigenicity. Snail induced nuclear translocation of p-ERK, and the activation of its subcellular downstream effector, Elk-1. Inhibiting MAPK activity with UO126 or knockdown of ERK2 isoform with siRNA in MCF-7 Snail cells reverted EMT induced by Snail as shown by decreased Snail and vimentin expression, decreased cell migration and increased cell adhesion. Overall, our data suggest that ERK2 isoform activation by Snail in aggressive breast cancer cells leads to EMT associated with increased cell migration and decreased cell adhesion. This regulation is enhanced by positive feedback regulation of Snail by ERK2. Therefore, therapeutic targeting of ERK2 isoform may be beneficial for breast cancer.

  11. Canine Mammary Cancer Stem Cells are Radio- and Chemo- Resistant and Exhibit an Epithelial-Mesenchymal Transition Phenotype

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    David J. Argyle

    2011-03-01

    Full Text Available Canine mammary carcinoma is the most common cancer among female dogs and is often fatal due to the development of distant metastases. In humans, solid tumors are made up of heterogeneous cell populations, which perform different roles in the tumor economy. A small subset of tumor cells can hold or acquire stem cell characteristics, enabling them to drive tumor growth, recurrence and metastasis. In veterinary medicine, the molecular drivers of canine mammary carcinoma are as yet undefined. Here we report that putative cancer stem cells (CSCs can be isolated form a canine mammary carcinoma cell line, REM134. We show that these cells have an increased ability to form tumorspheres, a characteristic of stem cells, and that they express embryonic stem cell markers associated with pluripotency. Moreover, canine CSCs are relatively resistant to the cytotoxic effects of common chemotherapeutic drugs and ionizing radiation, indicating that failure of clinical therapy to eradicate canine mammary cancer may be due to the survival of CSCs. The epithelial to mesenchymal transition (EMT has been associated with cancer invasion, metastasis, and the acquisition of stem cell characteristics. Our results show that canine CSCs predominantly express mesenchymal markers and are more invasive than parental cells, indicating that these cells have a mesenchymal phenotype. Furthermore, we show that canine mammary cancer cells can be induced to undergo EMT by TGFβ and that these cells have an increased ability to form tumorspheres. Our findings indicate that EMT induction can enrich for cells with CSC properties, and provide further insight into canine CSC biology.

  12. 6-OH-BDE-47 promotes human lung cancer cells epithelial mesenchymal transition via the AKT/Snail signal pathway.

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    Qu, Bao-Lin; Yu, Wei; Huang, Yu-Rong; Cai, Bo-Ning; Du, Le-Hui; Liu, Fang

    2015-01-01

    Hydroxylated polybrominated diphenyl ethers (OH-PBDEs) have been detected in the various human tissues. The OH-PBDEs are suggested to be stronger endocrine-disrupting compounds than PBDEs, therefore the toxicological effects of OH-PBDEs had received lots of attention. However, there is no study about the carcinogenic effect of OH-PBDEs and their estrogen potencies on the tumorigenesis and development of cancer. In the present study, we found that 6-hydroxy-2,2',4',4'-tetrabromodiphenyl ether (6-OH-BDE-47), the most abundant OH-PBDE congeners in human serum, promoted the in vitro migration of lung cancer A549 and H358 cells by induction of epithelial to mesenchymal transition (EMT). This was confirmed by that 6-OH-BDE-47 significantly down regulated the expression of epithelial markers E-cadherin (E-Cad) and zona occludin-1 (ZO-1) while up regulated the mesenchymal markers vimentin (Vim) and N-cadherin (N-Cad). 6-OH-BDE-47 up regulated the protein while not mRNA levels of Snail, which was the key transcription factor of EMT. Silencing of Snail by use of siRNA attenuated the 6-OH-BDE-47 induced EMT. This suggested that the stabilization of Snail was essential for 6-OH-BDE-47 induced EMT. Further, the treatment of 6-OH-BDE-47 increased the phosphorylation of AKT and ERK in A549 cells. Only PI3K/AKT inhibitor (LY294002), but not ERK inhibitor (PD98059), completely blocked the 6-OH-BDE-47 induced up regulation of Snail and down regulation of E-Cad, suggesting that PI3K/AKT pathway is important for 6-OH-BDE-47-mediated Snail stabilization and EMT in A549 cells. Generally, our results revealed for the first time that 6-OH-BDE-47 promoted the EMT of lung cancer cells via AKT/Snail signals. This suggested that more attention should be paid to the effects of OH-PBDEs on tumorigenesis and development of lung cancer. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Cancer stem-like cells enriched with CD29 and CD44 markers exhibit molecular characteristics with epithelial-mesenchymal transition in squamous cell carcinoma.

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    Geng, Songmei; Guo, Yuanyuan; Wang, Qianqian; Li, Lan; Wang, Jianli

    2013-01-01

    Increasing evidences have indicated that only a phenotypic subset of cancer cells, termed as the cancer stem cells (CSCs), is capable of initiating tumor growth and provide a reservoir of cells that cause tumor recurrence after therapy. Epithelial-mesenchymal transition (EMT), a cell type change from an epithelial cobblestone phenotype to an elongated fibroblastic phenotype, plays a critical role not only in tumor metastasis but also in tumor recurrence and contributes to drug resistance. Accumulating evidence has shown that cells with an EMT phenotype are rich sources for CSCs, suggesting a biological link between EMT and CSCs; thus study on the link will help understand the cellular and molecular mechanisms of tumor metastasis and drug resistance. CD29 is involved in EMT through cross-talk with cadherins and CD44 has been reported as a successful used marker for CSCs. Here, we try to address whether combination of CD29 and CD44 could be used to identify cancer stem-like cells undergoing EMT in squamous cell carcinoma (SCC) and compare the molecular differences between CD29high/CD44high and CD29low/CD44low cells in SCC. Expression pattern of CD29 and CD44 was analyzed in tissues of skin SCC and cultured A431 cells by immunostaining. Subtype cells of CD29high/CD44high and CD29low/CD44low A431 were sorted by fluorescence-activated cell sorting and proliferating abilities were assayed by cell counting, colony forming and tumorigenicity in NOD/SCID mice. Finally, to probe more deeply into the molecular differences between CD29high/CD44high and CD29low/CD44low A431 cells, gene microarray analysis was applied to compare gene expression profiling. Staining of CD29 and CD44 showed similar heterogeneous expression pattern with positive cells located in the invasion front of SCC tissue as well as in cultured A431 cells. Sorted CD29high/CD44high A431 cells had higher proliferating ability in vitro and in NOD/SCID mice as compared with CD29low/CD44low cells. Gene profiling

  14. Epithelial-mesenchymal transition in primary human bronchial epithelial cells is Smad-dependent and enhanced by fibronectin and TNF-α

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    Câmara Joana

    2010-01-01

    Full Text Available Abstract Background Defective epithelial repair, excess fibroblasts and myofibroblasts, collagen overproduction and fibrosis occur in a number of respiratory diseases such as asthma, chronic obstructive pulmonary disease (COPD and pulmonary fibrosis. Pathological conversion of epithelial cells into fibroblasts (epithelial-mesenchymal transition, EMT has been proposed as a mechanism for the increased fibroblast numbers and has been demonstrated to occur in lung alveolar epithelial cells. Whether other airway cell types also have the capability to undergo EMT has been less explored so far. A better understanding of the full extent of EMT in airways, and the underlying mechanisms, can provide important insights into airway disease pathology and enable the development of new therapies. The main aim of this study was to test whether primary human bronchial epithelial cells are able to undergo EMT in vitro and to investigate the effect of various profibrotic factors in the process. Results Our data demonstrate that primary human bronchial epithelial cells (HBECs are able to undergo EMT in response to transforming growth factor-beta 1 (TGF-β1, as revealed by typical morphological alterations and EMT marker progression at the RNA level by real-time quantitative polymerase chain reaction and, at the protein level, by western blot. By using pharmacological inhibitors we show that this is a Smad-dependent mechanism and is independent of extracellular signal-related kinase pathway activation. Additional cytokines and growth factors such as tumour necrosis factor-alpha (TNF-α, interleukin-1 beta (IL1β and connective tissue growth factor (CTGF were also tested, alone or in combination with TGF-β1. TNF-α markedly enhances the effect of TGF-β1 on EMT, whereas IL1β shows only a very weak effect and CTGF has no significant effect. We have also found that cell-matrix contact, in particular to fibronectin, an ECM component upregulated in fibrotic lesions

  15. Leptin-induced Epithelial-Mesenchymal Transition in Breast Cancer Cells Requires β-Catenin Activation via Akt/GSK3- and MTA1/Wnt1 Protein-dependent Pathways*

    Science.gov (United States)

    Yan, Dan; Avtanski, Dimiter; Saxena, Neeraj K.; Sharma, Dipali

    2012-01-01

    Perturbations in the adipocytokine profile, especially higher levels of leptin, are a major cause of breast tumor progression and metastasis; the underlying mechanisms, however, are not well understood. In particular, it remains elusive whether leptin is involved in epithelial-mesenchymal transition (EMT). Here, we provide molecular evidence that leptin induces breast cancer cells to undergo a transition from epithelial to spindle-like mesenchymal morphology. Investigating the downstream mediator(s) that may direct leptin-induced EMT, we found functional interactions between leptin, metastasis-associated protein 1 (MTA1), and Wnt1 signaling components. Leptin increases accumulation and nuclear translocation of β-catenin leading to increased promoter recruitment. Silencing of β-catenin or treatment with the small molecule inhibitor, ICG-001, inhibits leptin-induced EMT, invasion, and tumorsphere formation. Mechanistically, leptin stimulates phosphorylation of glycogen synthase kinase 3β (GSK3β) via Akt activation resulting in a substantial decrease in the formation of the GSK3β-LKB1-Axin complex that leads to increased accumulation of β-catenin. Leptin treatment also increases Wnt1 expression that contributes to GSK3β phosphorylation. Inhibition of Wnt1 abrogates leptin-stimulated GSK3β phosphorylation. We also discovered that leptin increases the expression of an important modifier of Wnt1 signaling, MTA1, which is integral to leptin-mediated regulation of the Wnt/β-catenin pathway as silencing of MTA1 inhibits leptin-induced Wnt1 expression, GSK3β phosphorylation, and β-catenin activation. Furthermore, analysis of leptin-treated breast tumors shows increased expression of Wnt1, pGSK3β, and vimentin along with higher nuclear accumulation of β-catenin and reduced E-cadherin expression providing in vivo evidence for a previously unrecognized cross-talk between leptin and MTA1/Wnt signaling in epithelial-mesenchymal transition of breast cancer cells. PMID

  16. Suppression of LPS-induced epithelial-mesenchymal transition by aqueous extracts of Prunella vulgaris through inhibition of the NF-κB/Snail signaling pathway and regulation of EMT-related protein expression.

    Science.gov (United States)

    Cho, In-Hye; Jang, Eun Hyang; Hong, Darong; Jung, Bom; Park, Min-Ju; Kim, Jong-Ho

    2015-11-01

    Epithelial-mesenchymal transition (EMT) is a pivotal event in the invasion and metastasis of cancer cells. Prunella vulgaris (PV) inhibits the proliferation of various cancer cells; however, its possible role in EMT has not been demonstrated. In the present study, we explored the effect of PV aqueous extract (PVAE), a typical medicine for decoction, on EMT. Lipopolysaccharide (LPS) induced EMT-like phenotype changes in cancer cell lines that enhanced cell migration and invasion. PVAE markedly inhibited these effects and produced accompanying changes in the expression of EMT markers, including decreased expression of N-cadherin and vimentin, and increased expression of β-catenin. We found that PVAE effects on LPS-induced EMT were mediated by inhibition of the NF-κB/Snail signaling pathway. Our findings provide new evidence that PVAE suppresses cancer invasion and migration by inhibiting EMT. Therefore, we suggest that PVAE is an effective dietary chemopreventive agent with antimetastatic activity against malignant tumors.

  17. 放疗诱导人肝细胞癌上皮-间质转化%Induction of epithelial-mesenchymal transition(EMT) in human hepatocellular carcinoma after radiotherapy

    Institute of Scientific and Technical Information of China (English)

    Ximing Xu; Junjian Deng; Guangjin Yuan; Miao Xiang; Biao Chen; Jiao Yang; Yiqiao Zhang; Lei Shi; Zuguo Li

    2012-01-01

    Objective: Epithelial-mesenchymal transition (EMT) is a critical early event for the invasion and metastasis of many carcinomas. In the present study, we examined EMT markers in the residual cancer cells of hepatocellular carcinoma (HCC) after radiotherapy. Methods: Eight patients with large HCC who underwent hepatectomy with preoperative radiotherapy were studied. The expressions of E-cadherin and vimentin were determined immunohistochemically in the residual cancer cells of HCC following radiotherapy, and also in the pre-radiotherapy biopsy cancer cells. Results: Histological analysis showed that some residual cancer cells of HCC displayed an elongated spindle or fibroblast-like shape. The expression of Ecadherin was markedly reduced or negative in the spindle residual cancer cells, but the expression of vimentin significantly induced.However, the above changes were not found in the pre-radiotherapy biopsy cancer cells. Conclusion: EMT is induced in the residual cancer cells of HCC following radiotherapy, which may facilitate the systemic dissemination of cancer cells.

  18. The Epigenetic Regulation of Epithelial-Mesenchymal Transition in Cancer Progression%EMT的表观遗传调控在癌症进程中的研究进展

    Institute of Scientific and Technical Information of China (English)

    张建超; 李红昌

    2015-01-01

    Epithelial-mesenchymal transition (EMT), a morphologic program in which cells convert from the epithelial to the mesenchymal state, plays a pivotal role during malignant tumor invasion-metastasis cascade. During the cancer progression, tumor cells undergo a series of dynamic and reversible cell phenotypic states transitions. Phenotypic plasticity of EMT program implies that epigenetic regulators play crucial roles in this process. Several EMT transcription factors can modulate EMT through regulating expression of the key target genes. These master EMT inducers orchestrate EMT program depending on complex epigenetic regulatory mechanisms. Therefore, understanding of epigenetic mechanisms controlling EMT will provide critical insights into the fundamental mechanisms underlying cancer metastasis, and new therapeutic targets for the treatment of malignant tumor.%上皮-间质转化(Epithelial-Mesenchymal Transition,EMT)是上皮细胞通过特定程序转变为间充质细胞的形态学过程,在癌症侵袭-转移级联过程中发挥着重要的作用。在癌症进程中,肿瘤细胞会经过一系列动态和可逆的细胞表型变化。EMT 程序的这种可塑性提示表观遗传调控在这一过程中发挥着重要的作用。EMT 相关的转录因子能够通过调控关键靶基因的表达,从而调节EMT程序。这些主要的EMT诱导因子依赖于表观遗传调控机制,从而调节EMT过程中基因表达变化。因此理解EMT调控的表观遗传机制有助于我们更好地了解肿瘤转移的分子机制,为恶性肿瘤的治疗提供新的靶点和思路。

  19. Chlamydia trachomatis Infection Is Associated with E-Cadherin Promoter Methylation, Downregulation of E-Cadherin Expression, and Increased Expression of Fibronectin and α-SMA—Implications for Epithelial-Mesenchymal Transition

    Directory of Open Access Journals (Sweden)

    Jovana Rajić

    2017-06-01

    Full Text Available Chlamydia trachomatis (Ct can induce scarring disease of the ocular mucosa, known as trachoma, the most common infectious cause of blindness worldwide. We hypothesized that epithelial-mesenchymal transition (EMT contributes to the fibrotic process in trachomatous scarring. Infection of human conjunctival epithelial cells (HCjE with Ct activated signaling pathways involved in EMT induction, which was correlated with decreased expression of E-cadherin, guardian of the epithelial phenotype. In addition, Ct infection was associated with increased expression of two mesenchymal cell markers: fibronectin and α-SMA. The DNA methylation statuses of selected regions of E-cadherin, fibronectin, and α-SMA genes revealed that Ct infection was accompanied with changes in DNA methylation of the E-cadherin promoter, while the expression of the two mesenchymal markers was not related with this epigenetic event. Our data suggest that Ct infection of conjunctival epithelial cells induces EMT-like changes that go along with modification of the methylation profile of the E-cadherin promoter and could, as one of the earliest events, contribute to processes triggering conjunctival scarring.

  20. The mechanisms for epithelial-mesenchymal transition in malignant cells%肿瘤细胞发生细胞上皮-间质转变机制的研究

    Institute of Scientific and Technical Information of China (English)

    赵俊卿; 李云峰; 杨之斌

    2010-01-01

    细胞上皮-间质转变(epithelial-mesenchymal transition,EMT),与肿瘤的局部浸润以及远处转移有着非常密切的关系.肿瘤细胞EMT的发生包括诸多因素,主要有转录因子表达上调以及信号通路[如磷脂酰肌醇3-激酶(phosphatidyl inositol-3 kinase,PI3K)/蛋白激酶B(protein kinase B, PKB,又称AKT)通路和Ras/丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)通路等]开启等,从而促使细胞间黏附力下降,失去了上皮细胞的顶底极性,并伴随细胞形态的改变.其中E-钙黏着糖蛋白(E-cadherins,E-cad)的表达下调或沉默是EMT发生的重要标志.本文就肿瘤细胞EMT发生机制的最新研究进展作一综述.

  1. Acquisition of epithelial-mesenchymal transition phenotype and cancer stem cell-like properties in cisplatin-resistant lung cancer cells through AKT/β-catenin/Snail signaling pathway.

    Science.gov (United States)

    Wang, Hao; Zhang, Ge; Zhang, Huan; Zhang, Fan; Zhou, Binhua; Ning, Fen; Wang, Hong-Sheng; Cai, Shao-Hui; Du, Jun

    2014-01-15

    Cisplatin is a first-line chemotherapeutic agent in the treatment of non-small cell lung cancer (NSCLC), but the therapeutic effect is disappointing, partly due to drug resistance. Emerging evidence showed that chemoresistance associates with acquisition of epithelial-mesenchymal transition (EMT) phenotype and cancer stem cell-like properties. However, the underlying mechanism is not entirely clear. In this study, we showed that cisplatin-resistant A549 cells (A549/CDDP) acquire EMT phenotype associated with migratory and invasive capability. A549/CDDP cells also displayed enhanced cancer stem cell-like properties. Increased expression of transcription factor Snail, but not ZEB1, Slug and Twist, was observed in A549/CDDP cells. Knockdown of Snail reversed EMT and significantly attenuated migration, invasion and cancer stem cell-like properties of A549/CDDP cells. Conversely, overexpressed Snail in A549 cells induced EMT and cancer stem cell-like properties. Finally, we demonstrated that activated AKT signal leads to increased β-catenin expression and subsequently up-regulates Snail in A549/CDDP cells. Taken together, these results revealed that AKT/β-catenin/Snail signaling pathway is mechanistically associated with cancer stem cell-like properties and EMT features of A549/CDDP cells, and thus, this pathway could be a novel target for the treatment of NSCLC. © 2013 Published by Elsevier B.V.

  2. Hepatitis C virus core protein increases Snail expression and induces epithelial-mesenchymal transition through the signal transducer and activator of transcription 3 pathway in hepatoma cells.

    Science.gov (United States)

    Zhou, Jia-Jia; Meng, Zhe; He, Xiao-Yu; Cheng, Di; Ye, Hui-Lin; Deng, Xiao-Geng; Chen, Ru-Fu

    2017-05-01

    Aberrant expression of Snail, a mediator of epithelial-mesenchymal transition (EMT), is crucial for cancer invasiveness and metastasis. Although hepatitis C virus (HCV) core protein has been implicated in hepatocarcinogenesis, the relationship between HCV core and Snail expression has not been clarified. HepG2 and Huh7 stable cell lines were established by transfection with pcDNA-HCVc. HepG2-HCVc and Huh7-HCVc cells were co-administered with AG490. Cell migration and invasiveness were tested. STAT3 and Snail expression was analyzed by Real-time PCR and Western blot. We found that HCV core is capable of increasing Snail expression and inducing EMT in hepatoma cells. HCV core-induced Snail expression was accompanied by activation of signal transducer and activator of transcription 3 (STAT3), inhibition of STAT3 abrogated HCV core-induced Snail expression and EMT. Furthermore, chromatin immunoprecipitation showed that phosphorylated STAT3 directly binds to the Snail promoter. Collectively, these results suggest that HCV core would play a role in hepatocellular carcinoma invasiveness and metastasis by activating the STAT3 pathway, increasing Snail expression and subsequently triggering EMT. These findings would advance the understanding of HCV-mediated invasiveness and metastasis, and might provide a new potential therapeutic target for HCV-related hepatocellular carcinoma. © 2016 The Japan Society of Hepatology.

  3. Role of RbBP5 and H3K4me3 in the vicinity of Snail transcription start site during epithelial-mesenchymal transition in prostate cancer cell.

    Science.gov (United States)

    Li, Dong; Sun, Hui; Sun, Wen-Jing; Bao, Hong-Bo; Si, Shu-Han; Fan, Jia-Lin; Lin, Ping; Cui, Rong-Jun; Pan, Yu-Jia; Wen, Si-Min; Zheng, Xiu-Lan; Yu, Xiao-Guang

    2016-10-04

    EMT (epithelial-mesenchymal transition) occurs in a wide range of tumor types, and has been shown to be crucial for metastasis. Epigenetic modifications of histones contribute to chromatin structure and result in the alterations in gene expression. Tri-methylation of histone H3 lysine 4 (H3K4me3) is associated with the promoters of actively transcribed genes and can serve as a transcriptional on/off switch. RbBP5 is a component of the COMPASS/ -like complex, which catalyzes H3K4me3 formation. In this study, we found that in the process of TGF-Beta1 induced EMT in the prostate cancer cell line DU145, H3K4me3 enrichment and RbBP5 binding increased in the vicinity of Snail (SNAI1) transcription start site. Knocking-down of RbBP5 notably decreased Snail expression and EMT. Recruitment of RbBP5 and formation of H3K4me3 at Snail TSS during EMT depend on binding of SMAD2/3 and CBP at Snail TSS. This study links the SMAD2/3 signal with Snail transcription via a histone modification - H3K4me3. Furthermore, our research also demonstrates that RbBP5 and even WRAD may be a promising therapeutic candidates in treating prostate cancer metastasis, and that DU145 cells maintain their incomplete mesenchymal state in an auto/ paracrine manner.

  4. Lectin-like oxidized low-density lipoprotein receptor-1 facilitates metastasis of gastric cancer through driving epithelial-mesenchymal transition and PI3K/Akt/GSK3β activation

    Science.gov (United States)

    Li, Can; Zhang, Jie; Wu, Hao; Li, Lili; Yang, Caiting; Song, Shushu; Peng, Peike; Shao, Miaomiao; Zhang, Mingming; Zhao, Junjie; Zhao, Ran; Wu, Weicheng; Ruan, Yuanyuan; Wang, Lan; Gu, Jianxin

    2017-01-01

    Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is a pattern recognition receptor that plays a critical role in vascular diseases and host immune response. Recently, our research discovered that LOX-1 could facilitate the uptake of dying cells and cross-presentation of cellular antigen via binding with heat shock proteins, which have a close relationship with gastric neoplasia. Therefore, we speculated that LOX-1 may serve as an oncogene in gastric cancer (GC) development and progression. In this study, through immunohistochemistry staining assay and cancer-related databases, we found that LOX-1 expression was up-regulated in GC tissues and correlated with a poor prognosis in GC patients. The expression of LOX-1 was an independent prognostic factor for OS in GC patients, and the incorporation of LOX-1 with TNM stage is more accurate for predicting prognosis. Additionally, in vitro study by transwell assay and western blot analysis confirmed that LOX-1 could promote the migration and invasion of GC cells by driving epithelial-mesenchymal transition and PI3K/Akt/GSK3β activation. Taken together, we first explored the expression profiles, clinical significance and biological function of LOX-1 in GC, and these data suggest that LOX-1 may represent a promising prognostic biomarker for GC and offer a novel molecular target for GC therapies. PMID:28345638

  5. miR-203 inhibits arecoline-induced epithelial-mesenchymal transition by regulating secreted frizzled-related protein 4 and transmembrane-4 L six family member 1 in oral submucous fibrosis.

    Science.gov (United States)

    Zheng, Lian; Jian, Xinchun; Guo, Feng; Li, Ning; Jiang, Canhua; Yin, Ping; Min, An-Jie; Huang, Long

    2015-06-01

    Oral submucous fibrosis (OSF) is a potentially malignant disease predominantly found in Asian people. The areca nut has been implicated in this disease. Arecoline, one of the areca alkaloids, induces epithelial-mesenchymal transition (EMT)-related factors in primary human buccal mucosal fibroblasts. Yet, the mechanisms of the underlying arecoline-induced EMT in OSF remain unknown. In the present study, we aimed to investigate the role of microRNAs (miRNAs) in arecoline-induced EMT in HaCaT cells. We found that miR-203 was significantly downregulated in OSF tissues compared to that in normal buccal mucosa tissues, and that miR-203 negatively regulated secreted frizzled-related protein 4 (SFRP4) and positively regulated transmembrane-4 L six family member 1 (TM4SF1). We observed that upregulation of miR-203 significantly decreased the cell proliferation of HaCaT cells, and significantly upregulated the expression of cytokeratin 19 (CK19) and E-cadherin proteins, whereas it significantly downregulated the expression of N-cadherin and vimentin compared to these levels in the vehicle control cells. Thus, we provide evidence to illustrate that miR-203 plays a role in the pathogenesis of OSF, which may be a target for OSF management.

  6. Copper depletion inhibits CoCl2-induced aggressive phenotype of MCF-7 cells via downregulation of HIF-1 and inhibition of Snail/Twist-mediated epithelial-mesenchymal transition.

    Science.gov (United States)

    Li, Shun; Zhang, Jing; Yang, Hong; Wu, Chunhui; Dang, Xitong; Liu, Yiyao

    2015-07-15

    Copper, a strictly regulated trace element, is essential for many physiological processes including angiogenesis. Dysregulated angiogenesis has been associated with increased copper in tumors, and thus copper chelators have been used to inhibit tumor angiogenesis. However, it remains unclear whether copper has any effect on epithelial-mesenchymal transition (EMT). Using CoCl2-induced EMT of human breast carcinoma MCF-7 cells, we found that TEPA, a copper chelator, inhibited EMT-like cell morphology and cytoskeleton arrangement triggered by CoCl2; decreased the expression of vimentin and fibronectin, markers typical of EMT; inhibited HIF-1 activation and HIF1-α accumulation in nuclear; and down-regulated the expression of hypoxia-associated transcription factors, Snail and Twist1. Moreover, knockdown copper transport protein, Ctr1, also inhibited CoCl2-induced EMT and reversed the mesenchymal phenotype. In EMT6 xenograft mouse models, TEPA administration inhibited the tumor growth and increased mice survival. Immunohistochemical analysis of the xenograft further demonstrated that TEPA administration significantly inhibited tumor angiogenesis, down-regulated hypoxia-induced transcription factors, Snail and Twist1, leading to decreased transactivation of EMT-associated marker genes, vimentin and fibronectin. These results indicate that TEPA inhibits CoCl2-induced EMT most likely via HIF1-α-Snail/Twist signaling pathway, and copper depletion may be exploited as a therapeutic for breast cancer.

  7. Embelin Inhibits Invasion and Migration of MDA-MB-231 Breast Cancer Cells by Suppression of CXC Chemokine Receptor 4, Matrix Metalloproteinases-9/2, and Epithelial-Mesenchymal Transition.

    Science.gov (United States)

    Lee, Hanwool; Ko, Jeong-Hyeon; Baek, Seung Ho; Nam, Dongwoo; Lee, Seok Geun; Lee, Junhee; Yang, Woong Mo; Um, Jae-Young; Kim, Sung-Hoon; Shim, Bum Sang; Ahn, Kwang Seok

    2016-06-01

    Embelin (EB) is a benzoquinone derivative isolated from Embelia ribes Burm plant. Recent scientific evidence shows that EB induces apoptosis and inhibits migration and invasion in highly metastatic human breast cancer cells. However, the exact mechanisms of EB in tumor metastasis and invasion have not been fully elucidated. Here, we investigated the underlying mechanisms of antimetastatic activities of EB in breast cancer cells. The EB downregulated the chemokine receptor 4 (CXCR4) as well as matrix metalloproteinase (MMP)-9/2 expression and upregulated the tissue inhibitor of metalloproteinase 1 expression in MDA-MB-231 cells under noncytotoxic concentrations but not in MCF-7 cells. Additionally, EB inhibited the CXC motif chemokine ligand 12 induced invasion and migration activities of MDA-MB-231 cells. A detailed study of underlying mechanisms revealed that the regulation of the downregulation of CXCR4 was at the transcriptional level, as indicated by the downregulation of mRNA expression and suppression of nuclear factor-kappa B (NF-κB) activation. It further reduced the binding of NF-κB to the CXCR4 promoter. Besides, EB downregulated mesenchymal marker proteins (neural cadherin and vimentin) and concurrently upregulated epithelial markers (epithelial cadherin and occludin). Overall, these findings suggest that EB can abrogate breast cancer cell invasion and metastasis by suppression of CXCR4, MMP-9/2 expressions, and inhibition of epithelial-mesenchymal transition and thus may have a great potential to suppress metastasis of breast cancer. Copyright © 2016 John Wiley & Sons, Ltd.

  8. 1,25(OH)2D3 attenuates TGF-β1/β2-induced increased migration and invasion via inhibiting epithelial-mesenchymal transition in colon cancer cells.

    Science.gov (United States)

    Chen, Shanwen; Zhu, Jing; Zuo, Shuai; Ma, Ju; Zhang, Junling; Chen, Guowei; Wang, Xin; Pan, Yisheng; Liu, Yucun; Wang, Pengyuan

    1,25-Dihydroxyvitamin D3 (1,25(OH)2D3) has been reported to inhibit proliferation and migration of multiple types of cancer cells. However, the mechanism underlying its anti-metastasis effect is not fully illustrated. In this study, the effect of 1,25(OH)2D3 on TGF-β1/β2-induced epithelial-mesenchymal transition (EMT) is tested in colon cancer cells. The results suggest that 1,25(OH)2D3 inhibited TGF-β1/β2-induced increased invasion and migration of in SW-480 and HT-29 cells. 1,25(OH)2D3 also inhibited the cadherin switch in SW-480 and HT-29 cells. TGF-β1/β2-induced increased expression of EMT-related transcription factors was also inhibited by 1,25(OH)2D3. 1,25(OH)2D3 also inhibited the secretion of MMP-2 and MMP-9 and increased expression of F-actin induced by TGF-β1/β2 in SW-480 cells. Taken together, this study suggests that the suppression of EMT might be one of the mechanisms underlying the anti-metastasis effect of 1,25(OH)2D3 in colon cancer cells.

  9. N1-Guanyl-1,7-Diaminoheptane Sensitizes Estrogen Receptor Negative Breast Cancer Cells to Doxorubicin by Preventing Epithelial-Mesenchymal Transition through Inhibition of Eukaryotic Translation Initiation Factor 5A2 Activation

    Directory of Open Access Journals (Sweden)

    Yu Liu

    2015-08-01

    Full Text Available Background: Approximately 30% of breast cancer does not express the estrogen receptor (ER, which is necessary for endocrine-based therapy approaches. Many studies demonstrated that eukaryotic translation initiation factor 5A2 (eIF5A2 serves as a proliferation-related oncogene in tumorigenic processes. Methods: The present study used cell viability assays, EdU incorporation assays, western blot, and immunofluorescence to explore whether N1-guanyl-1,7-diaminoheptane (GC7, which inhibits eIF5A2 activation, exerts synergistic cytotoxicity with doxorubicin in breast cancer. Results: We found that GC7 enhanced doxorubicin cytotoxicity in ER-negative HCC1937 cells but had little effect in ER-positive MCF-7 and Bcap-37 cells. Administration of GC7 reversed the doxorubicin-induced epithelial-mesenchymal transition (EMT in ER-negative breast cancer cells. Knockdown of eIF5A2 by siRNA inhibited the doxorubicin-induced EMT in ER-negative HCC1937 cells. Conclusion: These data demonstrated that GC7 combination therapy may enhance the therapeutic efficacy of doxorubicin in estrogen negative breast cancer cells by preventing EMT through inhibition of eIF5A2 activation.

  10. 上皮细胞间质转化在大肠癌发生发展中的研究进展%Progress in understanding the role of epithelial-mesenchymal transition in the pathogenesis of colorectal tumors

    Institute of Scientific and Technical Information of China (English)

    朱庆超; 秦环龙

    2012-01-01

    Epithelial-mesenchymal transition is a well established biological event that plays an important role not only in the normal development of tissues and organs but also in the pathogenesis of many diseases. Increasing evidence has established its presence in the human colon during colorectal carcinogenesis and cancer invasion, chronic inflammation-related fibrosis, and mu-cosal healing. A large body of evidence supports the role of transforming growth factor-p and its downstream Smad signaling, the phosphati-dylinositol 3'-kinase/Akt/mTOR axis, the Ras-mitogen-activated protein kinase/Snail/Slug and FOXC2 pathway, and Hedgehog signaling and microRNAs in epithelial-mesenchymal transition in the development of colorectal cancers. Here we discuss the role of these pathways in the initiation and development of the transition events. A better understanding of their induction and regulation may lead to the identification of pathways and factors that could be potent therapeutic targets.%上皮间质转化是一种已知的分子事件,他不仅在正常组织器官发育过程中起着重要的作用,而且在疾病状态也发挥着重要的作用.越来越多的证据显示在人类大肠癌形成和肿瘤侵袭过程中发生了上皮间质转化,而且其也参与了慢性炎症相关性纤维化和黏膜的修复过程.在大肠癌发生发展过程中越来越多的证据支持转化生长因子β(transforming growth factor-β,TGF-β)和他下游的Smad信号传导、磷酸酰肌醇3’激酶/Akt/mTOR轴、Ras丝裂原活化蛋白激酶/Snail/Slug/FOXC2途径、Hedgehog信号通路和microRNAs等介导的上皮细胞间质转化(epithelial-to-mesenchymal transition,EMT)所发挥的重要作用.现将对这些途径在大肠癌上皮细胞间质转化启动和发展过程中的作用进行综述.对EMT在大肠癌发生发展过程中所起到的诱导和调控作用进行深入了解,将会促进对相关信号通路和潜在治疗靶点分子的认识.

  11. 非可控性炎症和肿瘤细胞上皮间质转化关系的研究进展%Relationship between nonresolving inflammation and epithelial-mesenchymal transition in tumor

    Institute of Scientific and Technical Information of China (English)

    倪建波; 郭传勇; 王兴鹏

    2011-01-01

    OBJECTIVE: To summarize the research advancement of the relationship between nonresolving inflammation and epithelial-mesenchymal transition in tumor, and discuss the role and probable mechanism of nonresolving inflammation in malignant transition of tumor cells. METHODS. By PubMed and CNK1 retrieval system, with "epithelial-mesenchymal transition (EMT), nonresolving inflammation and tumor microenvironment" as key words, papers from 2005-01 to 2010-12 were obtained. Twenty-six papers of literature are selected and analyzed according to the inclusion criteria as follows: Dthe relationship between nonresolving inflammation and tumor; 2)The relationship between nonresolving inflammation and EMT; 3) Mechanisms of EMT regulated by nonresolving inflammation. RESULTS: In nonresolving inflammatory status, inflammatory cytokines such as TGF-f), TNF-a and Ils can initiate and maintain the EMT process of tumor cells, which enhances their invasive and migratory ability, by activating the transcription factors associated with EMT through NF-kB signaling pathway. Furthermore, EMT promotes construction of the inflammatory tumor microenvironment. CONCLUSIONS: Nonresolving inflammation and EMT are tightly related in tumor, and inhibition of EMT through targeting the key components of the Nonresolving Inflammation-EMT signaling network probably be novel strategies for the prevention and treatment of tumor in the future.%目的:总结国内外关于非可控性炎症和肿瘤细胞上皮间质转化关系的研究进展,探讨非可控性炎症在肿瘤细胞恶性转化中的作用及可能机制.方法:应用PubMed及CNKI期刊全文数据库系统,以“上皮间质转化( Epithelial-mesenchymaltransition,EMT)、非可控性炎症及肿瘤微环境”为关键词,检索2005-01-2010-12的相关文献.纳入标准:1)非可控性炎症与肿瘤的联系;2) EMT与肿瘤的关系;3)非可控性炎症对EMT的调控机制.根据纳入标准分析26篇文献.结果:在非可控性

  12. c-Src activation promotes nasopharyngeal carcinoma metastasis by inducing the epithelial-mesenchymal transition via PI3K/Akt signaling pathway: a new and promising target for NPC

    Science.gov (United States)

    Lu, Jinping; Xia, Weixiong; Yu, Yahui; Peng, Yongjian; Wang, Li; Wang, Gang; Ye, Yanfang; Yang, Jing; Liang, Hu; Kang, Tiebang; Lv, Xing

    2016-01-01

    Aberrant activation of cellular Src (c-Src), a non-receptor tyrosine kinase, could promote cancer progression through activating its downstream signaling pathways. However, the roles of c-Src and phosphorylated-Src (p-Src) in nasopharyngeal carcinoma (NPC) progression are rarely investigated. Herein, we have identified high c-Src concentrations in the serum of NPC patients with distant metastasis using high-throughput protein microarrays. Levels of c-Src in serum and p-Src in human primary NPC samples were unfavorable independent prognostic factors for cancer-specific survival, disease-free survival, and distant metastasis-free survival. Depletion or inactivation of c-Src in NPC cells using sgRNA with CRISPR/Cas9 system or PP2 decreased cell viability, colony formation, migration and invasion in vitro and metastasis in vivo. In contrast, these malignancies could be up-regulated by overexpressed c-Src in a NPC cell line with low-metastasis potential. Furthermore, p-Src was involved in promoting NPC cell metastasis by inducing the epithelial-mesenchymal transition (EMT) process via activating the PI3K/Akt pathway and cytoskeleton remodeling. The p-Src-induced EMT process could be retarded by PP2, which mediated by down-regulating the PI3K/Akt pathway. In conclusion, elevated levels of c-Src in serum and p-Src in primary NPC tissue correlated with poor outcomes of NPC patients. And aberrant activation of c-Src facilitated NPC cells with malignant potential, especially metastasis ability, which mediated by the PI3K/Akt pathway activation and sequentially induced the EMT process. These findings unveiled a promising approach for targeted therapy of advanced NPC. PMID:27078847

  13. Inhibition of Nonsmall Cell Lung Cancer Cell Migration by Protein Arginine Methyltransferase 1-small Hairpin RNA Through Inhibiting Epithelial-mesenchymal Transition,Extracellular Matrix Degradation, and Src Phosphorylation In Vitro

    Institute of Scientific and Technical Information of China (English)

    Ting Zhang; Ge Cui; Yun-Liang Yao; Yue Guo; Qi-Chun Wang; Xi-Ning Li; Wen-Ming Feng

    2015-01-01

    Background:Protein arginine methyltransferases 1 (PRMT1) is over-expressed in a variety of cancers,including lung cancer,and is correlated with a poor prognosis of tumor development.This study aimed to investigate the role of PRMT1 in nonsmall cell lung cancer (NSCLC) migration in vitro.Methods:In this study,PRMT1 expression in the NSCLC cell line A549 was silenced using lentiviral vector-mediated short hairpin RNAs.Cell migration was measured using both scratch wound healing and transwell cell migration assays.The mRNA expression levels of matrix metalloproteinase 2 (MMP-2) and tissue inhibitor ofmetalloproteinase 1,2 (TIMP l,2) were measured using quantitative real-time reverse transcription-polymerase chain reaction.The expression levels of protein markers for epithelial-mesenchymal transition (EMT) (E-cadherin,N-cadherin),focal adhesion kinase (FAK),Src,AKT,and their corresponding phosphorylated states were detected by Western blot.Results:Cell migration was significantly inhibited in the PRMT1 silenced group compared to the control group.The mRNA expression of MMP-2 decreased while TIMP 1 and TIMP2 increased significantly.E-cadherin mRNA expression also increased while N-cadherin decreased.Only phosphorylated Src levels decreased in the silenced group while FAK or AKT remained unchanged.Conclusions:PRMT1-small hairpin RNA inhibits the migration abilities of NSCLC A549 cells by inhibiting EMT,extracellular matrix degradation,and Src phosphorylation in vitro.

  14. A comprehensive comparative analysis of the histomorphological features of ALK-rearranged lung adenocarcinoma based on driver oncogene mutations: frequent expression of epithelial-mesenchymal transition markers than other genotype.

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    Hyojin Kim

    Full Text Available Molecular classification of lung cancer correlates well with histomorphological features. However, specific histomorphological features that differentiate anaplastic lymphoma kinase (ALK-rearranged tumors from ALK-negative tumors have not been fully evaluated. Eighty ALK-rearranged and 213 ALK-negative (91 epidermal growth factor receptor-mutated; 29 K-ras-mutated; 93 triple-negative resected lung adenocarcinomas were analyzed for several histomorphological parameters and histological subtype. ALK-rearranged tumors were associated with younger age at presentation, frequent nodal metastasis, and higher stage of disease at diagnosis. ALK-rearranged tumors were more likely to show a solid predominant pattern than ALK-negative tumors (43.8%; 35/80; p<0.001. Unlike ALK-negative tumors, a lepidic predominant pattern was not observed in ALK-rearranged tumors (p<0.001. In multivariate analysis, the most significant morphological features that distinguished ALK-rearranged tumors from ALK-negative tumors were cribriform formation (odds ratio [OR], 3.253; p = 0.028, presence of mucin-containing cells (OR, 4.899; p = 0.008, close relationship to adjacent bronchioles (OR, 5.361; p = 0.001, presence of psammoma bodies (OR, 4.026; p = 0.002, and a solid predominant pattern (OR, 13.685; p = 0.023. ALK-rearranged tumors exhibited invasive histomorphological features, aggressive behavior and frequent expression of epithelial-mesenchymal transition markers (loss of E-cadherin and expression of vimentin compared with other genotype (p = 0.015. Spatial proximity between bronchus and ALK-rearranged tumors and frequent solid histologic subtype with p63 expression may cause diagnostic difficulties to differentiate squamous cell carcinoma in the small biopsy, whereas p40 was rarely expressed in ALK-rearranged adenocarcinoma. Knowledge of these features may improve the diagnostic accuracy and lead to a better understanding of the characteristic behavior of ALK

  15. The complex interplay between ERK1/2, TGFβ/Smad, and Jagged/Notch signaling pathways in the regulation of epithelial-mesenchymal transition in retinal pigment epithelium cells.

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    Xiaoyun Chen

    Full Text Available Epithelial-mesenchymal transition (EMT of retinal pigment epithelium (RPE cells is a major pathologic change in the development of proliferative vitreoretinopathy (PVR, which leads to severe visual impairment. ERK1/2 pathway has been reported to play a key role in the carcinogenesis, cancer metastasis, and multiple fibrotic diseases. We hypothesized that ERK1/2 signaling could cross-interact with transforming growth factor β2 (TGFβ2/Smad and Notch signaling pathways in the regulation of EMT in RPE cells. Here, we demonstrated that ERK1/2 signaling was activated in TGFβ2-induced EMT in human RPE cells, while blockade of the canonical TGFβ2/Smad2/3 signaling with SB431542 could not inhibit TGFβ2-induced the activation of ERK1/2. Meanwhile, blockade of ERK1/2 signaling with a specific MEK/ERK1/2 inhibitor U0126 strongly prevented TGFβ2-induced the downregulation of P-cadherin, and the upregulation of α-SMA, collagen type IV, N-cadherin and fibronectin in RPE cells. In addition, we also identified that blockade of ERK1/2 signaling could inhibit not only the canonical TGFβ/Smad signaling, but also the Jagged/Notch pathway. Finally, we found that blockade of Notch pathway with a specific inhibitor DAPT could inhibit TGFβ2-induced the activation of ERK1/2 pathway conversely. Therefore, our study provides evidence that ERK1/2 signaling can cross-interact with the canonical TGFβ/Smad and the Jagged/Notch signaling pathways in RPE cells EMT. ERK1/2 inhibitor may have therapeutic value in the prevention and treatment of PVR and other fibrotic diseases.

  16. Targeting the Nuclear Cathepsin L CCAAT Displacement Protein/Cut Homeobox Transcription Factor-Epithelial Mesenchymal Transition Pathway in Prostate and Breast Cancer Cells with the Z-FY-CHO Inhibitor

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    Burton, Liza J.; Dougan, Jodi; Jones, Jasmine; Smith, Bethany N.; Randle, Diandra; Henderson, Veronica

    2016-01-01

    ABSTRACT The epithelial mesenchymal transition (EMT) promotes tumor migration and invasion by downregulating epithelial markers such as E-cadherin and upregulating mesenchymal markers such as vimentin. Cathepsin L (Cat L) is a cysteine protease that can proteolytically activate CCAAT displacement protein/cut homeobox transcription factor (CUX1). We hypothesized that nuclear Cat L may promote EMT via CUX1 and that this could be antagonized with the Cat L-specific inhibitor Z-FY-CHO. Mesenchymal prostate (ARCaP-M and ARCaP-E overexpressing Snail) and breast (MDA-MB-468, MDA-MB-231, and MCF-7 overexpressing Snail) cancer cells expressed lower E-cadherin activity, higher Snail, vimentin, and Cat L activity, and a p110/p90 active CUX1 form, compared to epithelial prostate (ARCaP-E and ARCaP-Neo) and breast (MCF-7 and MCF-7 Neo) cancer cells. There was increased binding of CUX1 to Snail and the E-cadherin promoter in mesenchymal cells compared to epithelial prostate and breast cells. Treatment of mesenchymal cells with the Cat L inhibitor Z-FY-CHO led to nuclear-to-cytoplasmic relocalization of Cat L, decreased binding of CUX1 to Snail and the E-cadherin promoter, reversed EMT, and decreased cell migration/invasion. Overall, our novel data suggest that a positive feedback loop between Snail-nuclear Cat L-CUX1 drives EMT, which can be antagonized by Z-FY-CHO. Therefore, Z-FY-CHO may be an important therapeutic tool to antagonize EMT and cancer progression. PMID:27956696

  17. Targeting the Nuclear Cathepsin L CCAAT Displacement Protein/Cut Homeobox Transcription Factor-Epithelial Mesenchymal Transition Pathway in Prostate and Breast Cancer Cells with the Z-FY-CHO Inhibitor.

    Science.gov (United States)

    Burton, Liza J; Dougan, Jodi; Jones, Jasmine; Smith, Bethany N; Randle, Diandra; Henderson, Veronica; Odero-Marah, Valerie A

    2017-03-01

    The epithelial mesenchymal transition (EMT) promotes tumor migration and invasion by downregulating epithelial markers such as E-cadherin and upregulating mesenchymal markers such as vimentin. Cathepsin L (Cat L) is a cysteine protease that can proteolytically activate CCAAT displacement protein/cut homeobox transcription factor (CUX1). We hypothesized that nuclear Cat L may promote EMT via CUX1 and that this could be antagonized with the Cat L-specific inhibitor Z-FY-CHO. Mesenchymal prostate (ARCaP-M and ARCaP-E overexpressing Snail) and breast (MDA-MB-468, MDA-MB-231, and MCF-7 overexpressing Snail) cancer cells expressed lower E-cadherin activity, higher Snail, vimentin, and Cat L activity, and a p110/p90 active CUX1 form, compared to epithelial prostate (ARCaP-E and ARCaP-Neo) and breast (MCF-7 and MCF-7 Neo) cancer cells. There was increased binding of CUX1 to Snail and the E-cadherin promoter in mesenchymal cells compared to epithelial prostate and breast cells. Treatment of mesenchymal cells with the Cat L inhibitor Z-FY-CHO led to nuclear-to-cytoplasmic relocalization of Cat L, decreased binding of CUX1 to Snail and the E-cadherin promoter, reversed EMT, and decreased cell migration/invasion. Overall, our novel data suggest that a positive feedback loop between Snail-nuclear Cat L-CUX1 drives EMT, which can be antagonized by Z-FY-CHO. Therefore, Z-FY-CHO may be an important therapeutic tool to antagonize EMT and cancer progression.

  18. Cancer-associated fibroblasts in a human HEp-2 established laryngeal xenografted tumor are not derived from cancer cells through epithelial-mesenchymal transition, phenotypically activated but karyotypically normal.

    Science.gov (United States)

    Wang, Mei; Wu, Chun-Ping; Pan, Jun-Yan; Zheng, Wen-Wei; Cao, Xiao-Juan; Fan, Guo-Kang

    2015-01-01

    Cancer-associated fibroblasts (CAFs) play a crucial role in cancer progression and even initiation. However, the origins of CAFs in various cancer types remain controversial, and one of the important hypothesized origins is through epithelial-mesenchymal transition (EMT) from cancer cells. In this study, we investigated whether the HEp-2 laryngeal cancer cells are able to generate CAFs via EMT during tumor formation, which is now still unknown. The laryngeal xenografted tumor model was established by inoculating the HEp-2 laryngeal cancer cell line in nude mice. Primary cultured CAFs from the tumor nodules and matched normal fibroblasts (NFs) from the adjacent connective tissues were subcultured, purified, and verified by immunofluorescence. Migration, invasion, and proliferation potentials were compared between the CAFs and NFs. A co-culture of CAFs with HEp-2 cells and a co-injection of CAFs with HEp-2 cells in nude mice were performed to examine the cancer-promoting potential of CAFs to further verify their identity. Karyotypic analyses of the CAFs, NFs, and HEp-2 cells were conducted. A co-culture of NFs with HEp-2 cells was also performed to examine the expression of activated markers of CAFs. A pathological examination confirmed that the laryngeal xenografted tumor model was successfully established, containing abundant CAFs. Immunocytochemical staining verified the purities and identities of the CAFs and NFs. Although the CAFs manifested higher migration, invasion, proliferation, and cancer-promoting capacities compared with the NFs, an analysis of chromosomes revealed that both the CAFs and NFs showed typical normal mouse karyotypes. In addition, the NFs co-cultured with HEp-2 cells did not show induced expressions of activated markers of CAFs. Our findings reveal that the CAFs in the HEp-2 established laryngeal xenografted tumor are not of laryngeal cancer origin but of mouse origin, indicating that the HEp-2 laryngeal cancer cells cannot generate their

  19. Clustering and jamming in epithelial-mesenchymal co-cultures.

    Science.gov (United States)

    Gamboa Castro, Marielena; Leggett, Susan E; Wong, Ian Y

    2016-10-12

    Collective behaviors emerge from coordinated cell-cell interactions during the morphogenesis of tissues and tumors. For instance, cells may display density-dependent phase transitions from a fluid-like "unjammed" phase to a solid-like "jammed" phase, while different cell types can "self-sort". Here, we comprehensively track single cell dynamics in mixtures of sheet-forming epithelial cells and dispersed mesenchymal cells. We find that proliferating epithelial cells nucleate multicellular clusters that coarsen at a critical density, arresting migration and strengthening spatial velocity correlations. The addition of mesenchymal cells can slow cluster formation and coarsening, resulting in more dispersed individual cells with weak spatial velocity correlations. These behaviors have analogies with a jamming-unjamming transition, where the control parameters are cell density and mesenchymal fraction. This complex interplay of proliferation, clustering and correlated migration may have physical implications for understanding epithelial-mesenchymal interactions in development and disease.

  20. Cyr61 promotes epithelial-mesenchymal transition and tumor metastasis of osteosarcoma by Raf-1/MEK/ERK/Elk-1/TWIST-1 signaling pathway.

    Science.gov (United States)

    Hou, Chun-Han; Lin, Feng-Ling; Hou, Sheng-Mon; Liu, Ju-Fang

    2014-10-19

    Osteosarcoma is the most common primary malignant tumor in children and young adults, and its treatment requires effective therapeutic approaches because of a high mortality rate for lung metastasis. Epithelial to mesenchymal transition (EMT) has received considerable attention as a conceptual paradigm for explaining the invasive and metastatic behavior during cancer progression. The cysteine-rich angiogenic inducer 61 (Cyr61) gene, a member of the CCN gene family, is responsible for the secretion of Cyr61, a matrix-associated protein that is involved in several cellular functions. A previous study showed that Cyr61 expression is related to osteosarcoma progression. In addition, Cyr61 could promote cell migration and metastasis in osteosarcoma. However, discussions on the molecular mechanism involved in Cyr61-regulated metastasis in osteosarcoma is poorly discussed. We determined that the expression level of Cyr61 induced cell migration ability in osteosarcoma cells. The Cyr61 protein promoted the mesenchymal transition of osteosarcoma cells by upregulating mesenchymal markers (TWIST-1 and N-cadherin) and inhibiting the epithelial marker (E-cadherin). Moreover, the Cyr61-induced cell migration was mediated by EMT. The Cyr61 protein elicited a signaling cascade that included αvβ5 integrin, Raf-1, mitogen-activated protein kinase (MEK), extracellular signal-regulated kinase (ERK), and Elk-1. The reagent or gene knockdown of these signaling proteins could inhibit Cyr61-promoted EMT in osteosarcoma. Finally, the knockdown of Cyr61 expression obviously inhibited cell migration and repressed mesenchymal phenotypes, reducing lung metastasis. Our results indicate that Cyr61 promotes the EMT of osteosarcoma cells by regulating EMT markers via a signal transduction pathway that involves αvβ5 integrin, Raf-1, MEK, ERK, and Elk-1.

  1. Proteomic analysis of ovarian cancer cells during epithelial-mesenchymal transition (EMT) induced by epidermal growth factor (EGF) reveals mechanisms of cell cycle control.

    Science.gov (United States)

    Grassi, Mariana Lopes; Palma, Camila de Souza; Thomé, Carolina Hassibe; Lanfredi, Guilherme Pauperio; Poersch, Aline; Faça, Vitor Marcel

    2017-01-16

    Epithelial to mesenchymal transition (EMT) is a well-orchestrated process that culminates with loss of epithelial phenotype and gain of a mesenchymal and migratory phenotype. EMT enhances cancer cell invasiveness and drug resistance, favoring metastasis. Dysregulation of transcription factors, signaling pathways, miRNAs and growth factors including EGF, TGF-beta and HGF can trigger EMT. In ovarian cancer, overexpression of the EGFR family is associated with more aggressive clinical behavior. Here, the ovarian adenocarcinoma cell line Caov-3 was induced to EMT with EGF in order to identify specific mechanisms controlled by this process. Caov-3 cells induced to EMT were thoroughly validated and a combination of subcellular proteome enrichment, GEL-LC-MS/MS and SILAC strategy allowed consistent proteome identification and quantitation. Protein network analysis of differentially expressed proteins highlighted regulation of metabolism and cell cycle. Activation of relevant signaling pathways, such as PI3K/Akt/mTOR and Ras/Erk MAPK, in response to EGF-induced EMT was validated. Also, EMT did not affected the proliferation rate of Caov-3 cells, but led to cell cycle arrest in G1 phase regulated by increased levels of p21Waf1/Cip1, independently of p53. Furthermore, a decrease in G1 and G2 checkpoint proteins was observed, supporting the involvement of EGF-induced EMT in cell cycle control. Cancer is a complex multistep process characterized by accumulation of several hallmarks including epithelial to mesenchymal transition (EMT), which promotes cellular and microenvironmental changes resulting in invasion and migration to distant sites, favoring metastasis. EMT can be triggered by different extracellular stimuli, including growth factors such as EGF. In ovarian cancer, the most lethal gynecological cancer, overexpression of the EGFR family is associated with more aggressive clinical behavior, increasing mortality rate caused by metastasis. Our proteomic data, together

  2. Melanoma cell-derived exosomes promote epithelial-mesenchymal transition in primary melanocytes through paracrine/autocrine signaling in the tumor microenvironment.

    Science.gov (United States)

    Xiao, Deyi; Barry, Samantha; Kmetz, Daniel; Egger, Michael; Pan, Jianmin; Rai, Shesh N; Qu, Jifu; McMasters, Kelly M; Hao, Hongying

    2016-07-01

    The tumor microenvironment is abundant with exosomes that are secreted by the cancer cells themselves. Exosomes are nanosized, organelle-like membranous structures that are increasingly being recognized as major contributors in the progression of malignant neoplasms. A critical element in melanoma progression is its propensity to metastasize, but little is known about how melanoma cell-derived exosomes modulate the microenvironment to optimize conditions for tumor progression and metastasis. Here, we provide evidence that melanoma cell-derived exosomes promote phenotype switching in primary melanocytes through paracrine/autocrine signaling. We found that the mitogen-activated protein kinase (MAPK) signaling pathway was activated during the exosome-mediated epithelial-to-mesenchymal transition (EMT)-resembling process, which promotes metastasis. Let-7i, an miRNA modulator of EMT, was also involved in this process. We further defined two other miRNA modulators of EMT (miR-191 and let-7a) in serum exosomes for differentiating stage I melanoma patients from non-melanoma subjects. These results provide the first strong molecular evidence that melanoma cell-derived exosomes promote the EMT-resembling process in the tumor microenvironment. Thus, novel strategies targeting EMT and modulating the tumor microenvironment may emerge as important approaches for the treatment of metastatic melanoma. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  3. RNA Interference Targeting Snail Inhibits the Transforming Growth Factor β 2-Induced Epithelial-Mesenchymal Transition in Human Lens Epithelial Cells.

    Science.gov (United States)

    Li, Ping; Jing, Jiaona; Hu, Jianyan; Li, Tiejun; Sun, Yuncheng; Guan, Huaijin

    2013-01-01

    Epithelial-msenchymal transition (EMT) contributes to posterior capsule opacification (PCO) type of cataract. Transcription factors Snail is a key trigger of EMT activated by transforming growth factor β (TGF β ). This study was done to investigate the effect of Snail targeting siRNA on TGF β 2-induced EMT in human lens epithelial cells. TGF β 2 treatment of cultured human epithelial cell line (HLEB3) upregulated the expression of Snail and the EMT relevant molecules such as vimentin and α -SMA but downregulated the expression of keratin and E-cadherin. After the stimulation of TGF β 2, the HLEB3 cells became fibroblast-like in morphology, and the junctions of cell-cell disappeared. TGF β 2 treatment also enhanced migration ability of HLEB3 cells. TGF β 2-induced Snail expression and EMT were significantly inhibited by Snail siRNA. By analyzing the response characteristics of HLEB3 in TGF β 2-induced EMT model with/without Snail-specific siRNA, we concluded that Snail is an element in the EMT of HLEB3 cells induced by TGF β 2. Snail siRNA targeting can block the induced EMT and therefore has the potential to suppress the development of PCO.

  4. microRNA-181a has a critical role in ovarian cancer progression through the regulation of the epithelial-mesenchymal transition

    Science.gov (United States)

    Parikh, Aditya; Lee, Christine; Joseph, Peronne; Marchini, Sergio; Baccarini, Alessia; Kolev, Valentin; Romualdi, Chiara; Fruscio, Robert; Shah, Hardik; Wang, Feng; Mullokandov, Gavriel; Fishman, David; D'Incalci, Maurizio; Rahaman, Jamal; Kalir, Tamara; Redline, Raymond W.; Brown, Brian D.; Narla, Goutham; Difeo, Analisa

    2014-01-01

    Ovarian cancer is a leading cause of cancer deaths among women. Effective targets to treat advanced epithelial ovarian cancer (EOC) and biomarkers to predict treatment response are still lacking because of the complexity of pathways involved in ovarian cancer progression. Here we show that miR-181a promotes TGF-β-mediated epithelial-to-mesenchymal transition via repression of its functional target, Smad7. miR-181a and phosphorylated Smad2 are enriched in recurrent compared with matched-primary ovarian tumours and their expression is associated with shorter time to recurrence and poor outcome in patients with EOC. Furthermore, ectopic expression of miR-181a results in increased cellular survival, migration, invasion, drug resistance and in vivo tumour burden and dissemination. In contrast, miR-181a inhibition via decoy vector suppression and Smad7 re-expression results in significant reversion of these phenotypes. Combined, our findings highlight an unappreciated role for miR-181a, Smad7, and the TGF-β signalling pathway in high-grade serous ovarian cancer.

  5. RNA Interference Targeting Snail Inhibits the Transforming Growth Factor β2-Induced Epithelial-Mesenchymal Transition in Human Lens Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Ping Li

    2013-01-01

    Full Text Available Epithelial-msenchymal transition (EMT contributes to posterior capsule opacification (PCO type of cataract. Transcription factors Snail is a key trigger of EMT activated by transforming growth factor β (TGFβ. This study was done to investigate the effect of Snail targeting siRNA on TGFβ2-induced EMT in human lens epithelial cells. TGFβ2 treatment of cultured human epithelial cell line (HLEB3 upregulated the expression of Snail and the EMT relevant molecules such as vimentin and α-SMA but downregulated the expression of keratin and E-cadherin. After the stimulation of TGFβ2, the HLEB3 cells became fibroblast-like in morphology, and the junctions of cell-cell disappeared. TGFβ2 treatment also enhanced migration ability of HLEB3 cells. TGFβ2-induced Snail expression and EMT were significantly inhibited by Snail siRNA. By analyzing the response characteristics of HLEB3 in TGFβ2-induced EMT model with/without Snail-specific siRNA, we concluded that Snail is an element in the EMT of HLEB3 cells induced by TGFβ2. Snail siRNA targeting can block the induced EMT and therefore has the potential to suppress the development of PCO.

  6. A novel spontaneous model of epithelial-mesenchymal transition (EMT) using a primary prostate cancer derived cell line demonstrating distinct stem-like characteristics.

    Science.gov (United States)

    Harner-Foreman, Naomi; Vadakekolathu, Jayakumar; Laversin, Stéphanie A; Mathieu, Morgan G; Reeder, Stephen; Pockley, A Graham; Rees, Robert C; Boocock, David J

    2017-01-17

    Cells acquire the invasive and migratory properties necessary for the invasion-metastasis cascade and the establishment of aggressive, metastatic disease by reactivating a latent embryonic programme: epithelial-to-mesenchymal transition (EMT). Herein, we report the development of a new, spontaneous model of EMT which involves four phenotypically distinct clones derived from a primary tumour-derived human prostate cancer cell line (OPCT-1), and its use to explore relationships between EMT and the generation of cancer stem cells (CSCs) in prostate cancer. Expression of epithelial (E-cadherin) and mesenchymal markers (vimentin, fibronectin) revealed that two of the four clones were incapable of spontaneously activating EMT, whereas the others contained large populations of EMT-derived, vimentin-positive cells having spindle-like morphology. One of the two EMT-positive clones exhibited aggressive and stem cell-like characteristics, whereas the other was non-aggressive and showed no stem cell phenotype. One of the two EMT-negative clones exhibited aggressive stem cell-like properties, whereas the other was the least aggressive of all clones. These findings demonstrate the existence of distinct, aggressive CSC-like populations in prostate cancer, but, importantly, that not all cells having a potential for EMT exhibit stem cell-like properties. This unique model can be used to further interrogate the biology of EMT in prostate cancer.

  7. Liver cancer-derived hepatitis C virus core proteins shift TGF-beta responses from tumor suppression to epithelial-mesenchymal transition.

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    Serena Battaglia

    Full Text Available BACKGROUND: Chronic hepatitis C virus (HCV infection and associated liver cirrhosis represent a major risk factor for hepatocellular carcinoma (HCC development. TGF-beta is an important driver of liver fibrogenesis and cancer; however, its actual impact in human cancer progression is still poorly known. The aim of this study was to investigate the role of HCC-derived HCV core natural variants on cancer progression through their impact on TGF-beta signaling. PRINCIPAL FINDINGS: We provide evidence that HCC-derived core protein expression in primary human or mouse hepatocyte alleviates TGF-beta responses in terms or growth inhibition or apoptosis. Instead, in these hepatocytes TGF-beta was still able to induce an epithelial to mesenchymal transition (EMT, a process that contributes to the promotion of cell invasion and metastasis. Moreover, we demonstrate that different thresholds of Smad3 activation dictate the TGF-beta responses in hepatic cells and that HCV core protein, by decreasing Smad3 activation, may switch TGF-beta growth inhibitory effects to tumor promoting responses. CONCLUSION/SIGNIFICANCE: Our data illustrate the capacity of hepatocytes to develop EMT and plasticity under TGF-beta, emphasize the role of HCV core protein in the dynamic of these effects and provide evidence for a paradigm whereby a viral protein implicated in oncogenesis is capable to shift TGF-beta responses from cytostatic effects to EMT development.

  8. Long-Term Safety Issues of iPSC-Based Cell Therapy in a Spinal Cord Injury Model: Oncogenic Transformation with Epithelial-Mesenchymal Transition

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    Satoshi Nori

    2015-03-01

    Full Text Available Previously, we described the safety and therapeutic potential of neurospheres (NSs derived from a human induced pluripotent stem cell (iPSC clone, 201B7, in a spinal cord injury (SCI mouse model. However, several safety issues concerning iPSC-based cell therapy remain unresolved. Here, we investigated another iPSC clone, 253G1, that we established by transducing OCT4, SOX2, and KLF4 into adult human dermal fibroblasts collected from the same donor who provided the 201B7 clone. The grafted 253G1-NSs survived, differentiated into three neural lineages, and promoted functional recovery accompanied by stimulated synapse formation 47 days after transplantation. However, long-term observation (for up to 103 days revealed deteriorated motor function accompanied by tumor formation. The tumors consisted of Nestin+ undifferentiated neural cells and exhibited activation of the OCT4 transgene. Transcriptome analysis revealed that a heightened mesenchymal transition may have contributed to the progression of tumors derived from grafted cells.

  9. Overexpression of p16(INK4a in urothelial carcinoma in situ is a marker for MAPK-mediated epithelial-mesenchymal transition but is not related to human papillomavirus infection.

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    Julie Steinestel

    Full Text Available BACKGROUND: The role of human papillomavirus (HPV in bladder carcinogenesis remains controversial. Overexpression of p16(INK4a, a surrogate marker for infection with oncogenic HPV in other tumours, has been described for urothelial carcinoma in situ (UCIS. Our goal was therefore to evaluate whether overexpression of p16(INK4a is associated with HPV infection and to identify mechanisms of p16(INK4a upregulation in UCIS. MATERIALS AND METHODS: In 60 tissue specimens from a total of 45 patients (UCIS and controls, we performed p16(INK4a immunohistochemistry followed by detection and subclassification of HPV DNA. In a subset of samples, we tested for gene amplification of p16(INK4a applying fluorescence in situ hybridization (FISH. RAS/MAPK signalling and epithelial-mesenchymal transition (EMT was assessed using immunohistochemistry. Finally, we transfected urothelial carcinoma cells with KRAS and examined the expression of p16(INK4a as well as markers of EMT. RESULTS: We found overexpression of p16(INK4a in 92.6% of UCIS and in all cervical intraepithelial neoplasia (CIN controls. In contrast, we detected high-risk HPV DNA in 80% of CIN, but none in UCIS. There was no gene amplification of p16(INK4a. High levels of phosphorylated kinases and urokinase plasminogen activator (uPA and loss of membraneous E-cadherin were detected in UCIS. KRAS transfection of urothelial carcinoma cells led to upregulation of p16(INK4a and uPA accompanied by loss of E-cadherin that could be inhibited by application of the kinase-inhibitor Sorafenib. CONCLUSIONS: Our results show that overexpression of p16(INK4a in UCIS is neither associated with HPV infection nor p16(INK4a gene amplification but is a consequence of enhanced RAS/MAPK signalling that promotes EMT, possibly due to Sorafenib-sensitive paracrine secretion of the EMT activator uPA. These findings might open a novel therapeutic option for localized but aggressive urothelial cancer.

  10. Synergistic effects of CD44 and TGF-β1 through AKT/GSK-3β/β-catenin signaling during epithelial-mesenchymal transition in liver cancer cells.

    Science.gov (United States)

    Park, Na Ri; Cha, Jung Hoon; Jang, Jeong Won; Bae, Si Hyun; Jang, Bohyun; Kim, Jung-Hee; Hur, Wonhee; Choi, Jong Young; Yoon, Seung Kew

    2016-09-01

    Cancer metastasis is strongly correlated with epithelial-mesenchymal transition (EMT), in which transforming growth factor-β (TGF-β) signaling plays a central role. CD44 has emerged as a cancer stem cell (CSC) marker that strongly induces EMT together with TGF-β1. This study aimed to investigate the link between high CD44 and TGF-β1 levels during EMT in HCC cell lines. FACS analysis showed high expression of CD44 in TGF-β1-positive SNU-368 cells and TGF-β1-negative SNU-354 cells. SNU-368 CD44(+) cells showed EMT through up-regulation of the AKT/GSK-3β/β-catenin pathway. By comparison, SNU-354 CD44(+) cells showed only increased N-cadherin expression, which was not accompanied by a decrease in E-cadherin expression, and also down-regulated the AKT/GSK-3β/β-catenin pathway. However, TGF-β1-stimulated SNU-354 cells (CD44/TGF-β1(+)) exhibited lower E-cadherin and higher N-cadherin expression with increased AKT/GSK-3β/β-catenin pathway activity. CD44/TGF-β1(+) SNU-354 cells also showed enhanced migration and formed larger spheres, while the TGF-β1-induced stem cell properties returned to their original state with the TGF-β1 inhibitor SB431542. SB431542-treated SNU-368 (CD44/TGF-β1(-)) cells also showed diminished N-cadherin and AKT/GSK-3β/β-catenin pathway activity and further decreased cell motility in a wound healing assay. However, CD44 knockdown in SNU-354 cells did not induce EMT even after treatment with TGF-β1. Finally, double inhibition of both CD44 and TGF-β1 further decreased migration and sphere formation more strongly than a single inhibition in SNU-368 cells. In conclusion, the current study demonstrated the synergistic interactions between CD44 and TGF-β1 in EMT induction and CSC properties through the AKT/GSK-3β/β-catenin pathway in HCC cells.

  11. Epithelial-mesenchymal transition process during human induced pluripotent stem cells differentiation%人诱导多能干细胞自然分化过程中EMT变化研究

    Institute of Scientific and Technical Information of China (English)

    林媛媛; 娄远蕾; 梁昌达; 谢淑佩; 谢安

    2013-01-01

    Objective To investigate whether the differentiation of induced pluripotent stem cells (iPS) is associated with the presence of epithelial-mesenchymal transition (EMT). Methods Human iPS cells were maintained in an undifferentiated state by culture in iPS cells medium in the presence of a fibroblast feeder layer, and were differentiated in EB cells medium. IPS cells dif-ferentiated for 0, 7, 14, 21, and 28 d were collected, and detected EMT associated genes and proteins by RT-RCR and western blot analysis. Results E-cadherin gene and protein were detected in undifferentiated human iPS cells, lower levels protein was ob-served following induction of differentiation. In contrast, N-cadherin gene and protein were absent in undifferentiated human iPS cells and subsequently detected following differentiation of the cells. Undifferentiated human iPS cells lacked transcripts encoding Slug and exhibited low levels of Snail transcripts. Upon differentiation both Snail and Slug transcripts were increased. Conclusion EMT event occurs during Human iPS cells differentiation.%目的:研究人诱导多能干细胞(induced pluripotent stem cells,iPS)自然分化过程中上皮-间质转化(epithelial-mes-enchymal transition,EMT)的变化过程。方法按常规方法iPS细胞接种于单层饲养层细胞上培养并传代,通过EB的方式令iPS细胞自然分化,收集未分化的iPS细胞以及分化7d、14d、21d和28d的细胞,采用RT-PCR和Western Blot方法检测EMT相关基因及蛋白的表达水平。结果 RT-PCR和Western Blot结果表明,未分化人iPS细胞可见E-cadherin基因和蛋白的表达,分化后E-cadherin的表达明显减少,而N-cadherin基因和蛋白在细胞未分化时均未见表达,发生分化后表达明显上调;在人iPS细胞分化过程中,Snail和Slug转录因子以及蛋白都明显发生上调。结论人iPS细胞自然分化与EMT,即E-cadherin和N-cadherin转化有关。

  12. Progress of Ovarian Cancer Stem Cells and Epithelial-mesenchymal Transition%卵巢癌干细胞与上皮-间质转化的研究进展

    Institute of Scientific and Technical Information of China (English)

    尧晓芬; 赖东梅

    2014-01-01

    近年有研究指出肿瘤可能是肿瘤干细胞性疾病。随着各种肿瘤干细胞不断被分离与鉴定出来,卵巢癌干细胞的分离与鉴定也取得了进展。卵巢癌干细胞是卵巢癌中具有自我更新和多向分化能力的细胞。越来越多的证据表明,肿瘤干细胞可能是介导肿瘤转移和耐药的关键细胞。同时,研究发现上皮-间质转化(EMT)在肿瘤转移中发挥着重要作用,且EMT可能介导肿瘤细胞获得干细胞特性。综述卵巢癌干细胞与EMT之间的关系,进一步探讨卵巢癌的潜在治疗策略。%Cancer as a disease driven by cancer stem cells is a concept that has emerged over the last few years. Along with varieties of cancer stem cells (CSCs) have been identified,the isolation and identification of ovarian cancer stem cells(OCSCs) developes quickly. OCSCs is a self-renewal and differentiation of cells. Re-cently,increasing evidences suggest that CSCs are potential candidates for mediating metastatic progression and chemoresistance. Studies have also indicated the process of epithelial-mesenchymal transition (EMT) plays an important role in regulating cancer metastasis. Meanwhile ,more and more findings illustrate a direct link between the EMT and the gain of stem-cell properties. The purpose of this review is to discuss the relationship between OCSCs and EMT,and to explore the potential therapeutic strategies of ovarian cancer.

  13. Multi-walled carbon nanotubes directly induce epithelial-mesenchymal transition in human bronchial epithelial cells via the TGF-β-mediated Akt/GSK-3β/SNAIL-1 signalling pathway.

    Science.gov (United States)

    Polimeni, Manuela; Gulino, Giulia Rossana; Gazzano, Elena; Kopecka, Joanna; Marucco, Arianna; Fenoglio, Ivana; Cesano, Federico; Campagnolo, Luisa; Magrini, Andrea; Pietroiusti, Antonio; Ghigo, Dario; Aldieri, Elisabetta

    2016-06-01

    Multi-walled carbon nanotubes (MWCNT) are currently under intense toxicological investigation due to concern on their potential health effects. Current in vitro and in vivo data indicate that MWCNT exposure is strongly associated with lung toxicity (inflammation, fibrosis, granuloma, cancer and airway injury) and their effects might be comparable to asbestos-induced carcinogenesis. Although fibrosis is a multi-origin disease, epithelial-mesenchymal transition (EMT) is recently recognized as an important pathway in cell transformation. It is known that MWCNT exposure induces EMT through the activation of the TGF-β/Smad signalling pathway thus promoting pulmonary fibrosis, but the molecular mechanisms involved are not fully understood. In the present work we propose a new mechanism involving a TGF-β-mediated signalling pathway. Human bronchial epithelial cells were incubated with two different MWCNT samples at various concentrations for up to 96 h and several markers of EMT were investigated. Quantitative real time PCR, western blot, immunofluorescent staining and gelatin zymographies were performed to detect the marker protein alterations. ELISA was performed to evaluate TGF-β production. Experiments with neutralizing anti-TGF-β antibody, specific inhibitors of GSK-3β and Akt and siRNA were carried out in order to confirm their involvement in MWCNT-induced EMT. In vivo experiments of pharyngeal aspiration in C57BL/6 mice were also performed. Data were analyzed by a one-way ANOVA with Tukey's post-hoc test. Fully characterized MWCNT (mean length EMT in an in vitro human model (BEAS-2B cells) after long-term incubation at sub-cytotoxic concentrations. MWCNT stimulate TGF-β secretion, Akt activation and GSK-3β inhibition, which induces nuclear accumulation of SNAIL-1 and its transcriptional activity, thus contributing to switch on the EMT program. Moreover, a significant increment of nuclear β-catenin - due to E-cadherin repression and following

  14. Response gene to complement-32 enhances metastatic phenotype by mediating transforming growth factor beta-induced epithelial-mesenchymal transition in human pancreatic cancer cell line BxPC-3

    Directory of Open Access Journals (Sweden)

    Zhu Liang

    2012-03-01

    Full Text Available Abstract Background Response gene to complement-32 (RGC-32 is comprehensively expressed in many kinds of tissues and has been reported to be expressed abnormally in different kinds of human tumors. However, the role of RGC-32 in cancer remains controversial and no reports have described the effect of RGC-32 in pancreatic cancer. The present study investigated the expression of RGC-32 in pancreatic cancer tissues and explored the role of RGC-32 in transforming growth factor-beta (TGF-β-induced epithelial-mesenchymal transition (EMT in human pancreatic cancer cell line BxPC-3. Methods Immunohistochemical staining of RGC-32 and E-cadherin was performed on specimens from 42 patients with pancreatic cancer, 12 with chronic pancreatitis and 8 with normal pancreas. To evaluate the role of RGC-32 in TGF-β-induced EMT in pancreatic cancer cells, BxPC-3 cells were treated with TGF-β1, and RGC-32 siRNA silencing and gene overexpression were performed as well. The mRNA expression and protein expression of RGC-32 and EMT markers such E-cadherin and vimentin were determined by quantitative reverse transcription-PCR (qRT-PCR and western blot respectively. Finally, migration ability of BxPC-3 cells treated with TGF-β and RGC-32 siRNA transfection was examined by transwell cell migration assay. Results We found stronger expression of RGC-32 and higher abnormal expression rate of E-cadherin in pancreatic cancer tissues than those in chronic pancreatitis tissues and normal pancreatic tissues. Immunohistochemical analysis revealed that both RGC-32 positive expression and E-cadherin abnormal expression in pancreatic cancer were correlated with lymph node metastasis and TNM staging. In addition, a significant and positive correlation was found between positive expression of RGC-32 and abnormal expression of E-cadherin. Furthermore, in vitro, we found sustained TGF-β stimuli induced EMT and up-regulated RGC-32 expression in BxPC-3 cells. By means of si

  15. 沉默Sam68基因表达抑制乳腺癌细胞的上皮-间质转化%Sam68 gene-silencing inhibits epithelial-mesenchymal transition of breast cancer cells

    Institute of Scientific and Technical Information of China (English)

    邝紫桥; 章乐虹; 潘小梅; 刘毅俊; 邹颖; 陈欣欣

    2016-01-01

    目的:探讨沉默Sam68 (Src-associated substrated during mitosis of 68KD)基因表达对乳腺癌MDA-MB-231细胞上皮-间质转化(epithelial-mesenchymal transition,EMT)的影响.方法:采用实时荧光定量PCR和蛋白质印迹法检测具有不同侵袭能力的乳腺癌MDA-MB-231和MCF-7细胞中Sam68 mRNA及蛋白的表达情况;选择Sam68高表达的MDA-MB-231细胞,利用siRNA技术沉默细胞中内源性Sam68蛋白的表达.随后,采用实时荧光定量PCR和蛋白质印迹法检测EMT标志物E-钙黏蛋白(E-cadherin)和波形蛋白(vimentin) mRNA及蛋白的表达;采用CCK-8法、Transwell小室迁移和侵袭实验检测沉默Sam68基因表达后对乳腺癌MDA-MB-231细胞增殖,迁移和侵袭的影响.结果:Sam68蛋白在高侵袭性的间质型乳腺癌MDA-MB-231细胞中高表达(P<0.05);采用siRNA沉默MDA-MB-231细胞内源性Sam68蛋白的表达后,E-cadherin的表达水平明显上调(P<0.05),而vimentin的表达水平明显下调(P<0.05);MDA-MB-231细胞的增殖能力以及细胞的迁移和侵袭能力明显降低(P值均< 0.05),细胞可能发生间质-上皮转化.结论:Sam68在高侵袭性乳腺癌细胞中高表达,沉默Sam68基因的表达能抑制EMT的发生,并降低细胞增殖、侵袭和迁移能力.

  16. RELATIONSHIP BETWEEN EPITHELIAL-MESENCHYMAL TRANSITION AND BIOLOGICAL BEHAVIOR OF PANCREATIC CANCER%上皮-间质转化与胰腺癌生物学行为的关系

    Institute of Scientific and Technical Information of China (English)

    张红梅; 王骥平; 陈德利

    2011-01-01

    目的 探讨上皮-间质转化(EMT)在胰腺癌浸润转移中的作用.方法 采用免疫组织化学PV6000法检测47例胰腺癌组织上皮-钙黏蛋白(E-cad)和波形蛋白(Vim)的表达.把E-cad阴性-Vim阳性表达作为癌细胞发生EMT的免疫表型.结果 47例胰腺癌组织中E-cad和Vim阳性率分别为46.8%、23.4%,其中E-cad阴性-Vim阳性表达7例,EMT发生率为19.1%.EMT发生率与胰腺癌的分化程度和转移有关(x2=6.73、4.06,P<0.05),与临床分期和1年生存率无关(x2=3.56、2.72,P>0.05).结论 EMT在胰腺癌的分化、浸润和转移过程中起重要作用.%Objective To assess the role of epithelial-mesenchymal transition (EMT) in infiltration and metastasis of pancreatic cancer. Methods Immunohistochemical PV 6000 technique was used to detect the expressions of E-cadherin (E-cad) and Vimentin (Vim) in 47 cases of pancreatic cancer tissue. Results The positive rates of E-cad and Vim were 46. 8% and 23. 4%, respectively, in 47 cases of pancreatic cancer tissue, of which, the E-cad negative-Vim positive expression seven cases, EMT incidence was 19.1%. The EMT was associated with the cancer cell differentiation and metastasis (χ2 = 6.73,4.06; P0. 05). Conclusion The EMT plays an important role in cell differentiation, invasion and metastasis of pancreatic carcinoma.

  17. DA-Raf-Mediated Suppression of the Ras--ERK Pathway Is Essential for TGF-β1-Induced Epithelial-Mesenchymal Transition in Alveolar Epithelial Type 2 Cells.

    Science.gov (United States)

    Watanabe-Takano, Haruko; Takano, Kazunori; Hatano, Masahiko; Tokuhisa, Takeshi; Endo, Takeshi

    2015-01-01

    Myofibroblasts play critical roles in the development of idiopathic pulmonary fibrosis by depositing components of extracellular matrix. One source of lung myofibroblasts is thought to be alveolar epithelial type 2 cells that undergo epithelial-mesenchymal transition (EMT). Rat RLE-6TN alveolar epithelial type 2 cells treated with transforming growth factor-β1 (TGF-β1) are converted into myofibroblasts through EMT. TGF-β induces both canonical Smad signaling and non-canonical signaling, including the Ras-induced ERK pathway (Raf-MEK-ERK). However, the signaling mechanisms regulating TGF-β1-induced EMT are not fully understood. Here, we show that the Ras-ERK pathway negatively regulates TGF-β1-induced EMT in RLE-6TN cells and that DA-Raf1 (DA-Raf), a splicing isoform of A-Raf and a dominant-negative antagonist of the Ras-ERK pathway, plays an essential role in EMT. Stimulation of the cells with fibroblast growth factor 2 (FGF2), which activated the ERK pathway, prominently suppressed TGF-β1-induced EMT. An inhibitor of MEK, but not an inhibitor of phosphatidylinositol 3-kinase, rescued the TGF-β1-treated cells from the suppression of EMT by FGF2. Overexpression of a constitutively active mutant of a component of the Ras-ERK pathway, i.e., H-Ras, B-Raf, or MEK1, interfered with EMT. Knockdown of DA-Raf expression with siRNAs facilitated the activity of MEK and ERK, which were only weakly and transiently activated by TGF-β1. Although DA-Raf knockdown abrogated TGF-β1-induced EMT, the abrogation of EMT was reversed by the addition of the MEK inhibitor. Furthermore, DA-Raf knockdown impaired the TGF-β1-induced nuclear translocation of Smad2, which mediates the transcription required for EMT. These results imply that intrinsic DA-Raf exerts essential functions for EMT by antagonizing the TGF-β1-induced Ras-ERK pathway in RLE-6TN cells.

  18. DNA methylation status of epithelial-mesenchymal transition (EMT--related genes is associated with severe clinical phenotypes in ulcerative colitis (UC.

    Directory of Open Access Journals (Sweden)

    Tomomitsu Tahara

    Full Text Available BACKGROUND: Epithelial-to-mesenchymal transition (EMT is a phenomenon that allows the conversion of adherent epithelial cells to a mesenchymal cell phenotype, which enhances migratory capacity and invasiveness. Recent studies have suggested that EMT contributes to the pathogenesis of ulcerative colitis (UC. We investigated the promoter DNA methylation status of EMT-related genes in the colonic mucosa in UC. METHODS: Colonic biopsies were obtained from the rectal inflammatory mucosa of 86 UC patients and the non-inflammatory proximal colonic mucosa of 10 paired patients. Bisulfite pyrosequencing was used to quantify the methylation of 5 candidate CpG island promoters (NEUROG1, CDX1, miR-1247, CDH1, and CDH13 and LINE1. RESULTS: Using an unsupervised hierarchical clustering analysis, inflamed rectal mucosa was well separated from mucosa that appeared normal. The CDH1 and CDH13 promoters were significantly associated with patient age (p = 0.04, 0.03, respectively. A similar trend was found between those genes and the duration of disease (CDH1: p = 0.07, CDH13: p = 0.0002, mean of both: p<0.00001. Several positive associations were found between hypermethylation and severe clinical phenotypes (CDX1 and miR-1247 and a refractory phenotype: p = 0.04 and 0.006, respectively. miR-1247 and CDH1 hyper methylation and a more severe Mayo endoscopic subscore: miR-1247: p = 0.0008, CDH1: p = 0.03, mean of both: p = 0.003. When the severe clinical phenotype was defined as having any of five phenotypes (hospitalized more than twice, highest Mayo endoscopic subscore, steroid dependence, refractory, or a history of surgery miR-1247 hypermethylation was associated with the same phenotype (p = 0.008. CONCLUSIONS: Our data suggest that variability in the methylation status of EMT-related genes is associated with more severe clinical phenotypes in UC.

  19. BDE-99 (2,2',4,4',5-pentabromodiphenyl ether) triggers epithelial-mesenchymal transition in colorectal cancer cells via PI3K/Akt/Snail signaling pathway.

    Science.gov (United States)

    Wang, Fei; Ruan, Xin-Jian; Zhang, Hong-Yan

    2015-01-01

    The gut is in direct contact with BDE-99 (2,2',4,4',5-pentabromodiphenyl ether), one of the most abundant PBDE congeners in the environment and in human tissues. The objective of the present study was to investigate the effects of BDE-99 on colorectal cancer (CRC) cells. The effects of BDE-99 on cell proliferation were measured by CCK-8 assay in the CRC cell line HCT-116. Wound healing and transwell migration/invasion assays were used to test the migration and invasion of CRC cells. Factors related to epithelial-to-mesenchymal transition (EMT) were measured by real-time PCR and Western blot analysis for mRNA and protein levels, respectively. BDE-99 was found to increase migration and invasion and trigger EMT in HCT-116 cells; EMT was characterized by cells acquiring mesenchymal spindle-like morphology and by increased expression of N-cadherin with a concomitant decrease in E-cadherin. BDE-99 treatment also increased the protein and mRNA levels of the transcription factor Snail, but not Slug, Twist, and ZEB1. Knockdown of Snail by siRNA significantly attenuated BDE-99-induced EMT in HCT-116 cells, suggesting that Snail plays a crucial role in BDE-99-induced EMT. The PI3K/Akt inhibitor LY294002 completely blocked BDE-99-induced Snail and invasion of HCT-116 cells. Our results revealed that BDE-99 can trigger the EMT of colon cancer cells via the PI3K/AKT/Snail signaling pathway. This study provides new insight into the tumorigenesis and metastasis of CRC stimulated by BDE-99 and possibly other PBDE congeners.

  20. Upregulation of kazrin F by miR-186 suppresses apoptosis but promotes epithelial-mesenchymal transition to contribute to malignancy in human cervical cancer cells.

    Science.gov (United States)

    Liu, Chang; Wang, Jinghua; Hu, Yang; Xie, Hong; Liu, Min; Tang, Hua

    2017-02-01

    Previous studies have identified that kazrin is a constituent of desmosome and influences intercellular adhesion, growing development and morphology. We previously cloned another new isoform, kazrin F and found that it has anti-apoptotic effects on human glioma cell line. To further explore whether kazrin F is involved in tumorigenesis, we investigated its expression and role in cervical cancer (CC) cells. The role of kazrin F and miR-186 in CC was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, colony formation, transwell, and apoptosis assays. Using enhanced green fluorescent protein (EGFP) reporter assays, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis, we identified kazrin F post-transcriptional regulation by miR-186. We demonstrate that kazrin F is highly expressed in CC tissues compared with the adjacent noncancerous tissues and promotes cell proliferation, colony formation, migration and invasion in HeLa and C33A cells by suppressing apoptosis and facilitating epithelial-to-mesenchymal transition (EMT). Furthermore, miR-186 was confirmed as a regulator of kazrin F dysregulation. An EGFP reporter assay proved that miR-186 directly targets the 3'-untranslated region (3'UTR) of kazrin F and downregulates its expression, and miR-186 expression showed an inverse correlation with kazrin F levels in CC tissues. In addition, overexpression of miR-186 suppressed the malignant behaviors of CC cells. The ectopic expression of kazrin F rescued the inhibitory effects of miR-186. Our findings indicate that the upregulation of kazrin F due to downregulated miR-186 levels contributes to malignancy, and highlight the significance of kazrin F in CC tumorigenesis.